RU2812474C2 - Caffeylpyruvate hydrolases and their use - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: invention relates to proteins of the biosynthesis of fungal luciferin. The invention discloses new enzymes of the biosynthesis of fungal luciferin and the nucleic acids encoding them, the use of proteins and nucleic acids in the biosynthesis of fungal luciferin.

EFFECT: disclosure of methods of obtaining luciferin and pre-luciferin from fungi in in vitro and in vivo systems, as well as the use of such substances as phosphors in various fields of science.

20 cl, 25 ex, 6 tbl, 9 dwg

Description

[001] The group of inventions relates to the field of biotechnology and genetic engineering. In particular, the invention relates to enzymes of the bioluminescent system of fungi.

State of the art

[002] Luciferases are enzymes that catalyze the oxidation of low molecular weight luciferin compounds, accompanied by the emission of light - bioluminescence. As a result of oxidation, oxyluciferin is formed from luciferin, which is released from the complex with the enzyme luciferase.

[003] Luciferases are widely used as reporter genes in a number of biomedical applications and in biotechnology. For example, luciferases are used to determine cell viability, activity of promoters and other components of living systems, in studies of carcinogenesis in animal models, in methods for identifying microorganisms and toxic agents in the environment, as indicators for determining the concentration of various substances, for visualizing the passage of signaling cascades, etc. .d. [Scott et al., Annu Rev Anal Chem, 2011, 4: 297-319; Badr and Tannous, Trends Biotechnol. 2011, 29: 624-33; Andreu et al., FEMS Microbiol Rev. 2011, 35: 360-94]. Many methods of using luciferases are described in reviews [Kaskova et al., Chem Soc Rev., 2016, 45: 6048-6077; Scott et al., Annu Rev Anal Chem, 2011, 4: 297-319; Widder and Falls, IEEE Journal of Selected Topics in Quantum Electronics, 2014, 20: 232-241]. All major applications of luciferases involve the detection of light emitted depending on the phenomenon or signal being studied. In most cases, detection is carried out using a luminometer or a modified optical microscope.

[004] Thousands of species capable of bioluminescence are known, for which about a dozen luciferins of different structures and several dozen corresponding luciferase enzymes have been described. It has been shown that in various organisms, bioluminescence systems arose independently in evolution more than forty times [Herring, Journal of Bioluminescence and Chemiluminescence, 1987, 1: 147-63; Haddock et al., Annual Review of Marine Science, 2010; 2: 443-93].

[005] A group of insect luciferases that catalyze the oxidation of D-luciferin has been described [de Wet et al., Proc. Natl. Acad. Sci. USA, 1985, 82: 7870-3; de Wet et al., Proc. Natl. Acad. Sci. USA, 1987, 7: 725-37]. A group of luciferases that catalyze the oxidation of coelenterazine [O. Shimomura, Bioluminescence: Chemical Principles and Methods, World Scientific Publishing Co. Pte. Ltd, Singapore, 2006, 470 p.]. Bioluminescent systems of ostracods of the genus Cypridina are known, which are characterized by chemically highly active luciferin and highly stable luciferase [Shimomura et al., Science, 1969, 164: 1299-300]. Bioluminescent systems of dinoflagellates and euphausiids are also known. Currently, genes encoding three luciferases from this group have been cloned [O. Shimomura, Bioluminescence: Chemical Principles and Methods, World Scientific Publishing Co. Pte. Ltd, Singapore, 2006]. A significant drawback of this system is its incomplete knowledge: complete luciferase sequences have not yet been established.

[006] Recently, a group of luciferases and the luciferin bioluminescent system of fungi have been described. Bioluminescence of fungi has been known for hundreds of years, but fungal luciferin was identified only in 2015: it turned out to be 3-hydroxyhispidin, a metabolite capable of penetrating cell membranes [Purtov et al., Angewandte Chemie, 2015, 54: 8124-28]. The same work confirmed the presence in fungal lysates of an enzyme that hydroxylates hispidin to produce luciferin, but the enzyme was not identified. Patent application No. 2017102986 dated January 30, 2017 described luciferase genes from several fungi using 3-hydroxyhispidin as luciferin, which has the structure:

[007] It has been shown that fungal luciferases can catalyze the light-emitting oxidation of other chemical compounds, the structures of which are shown in Table 1 [Kaskova et al., Sci. Adv. 2017;3:e1602847]. All of these compounds, which are fungal luciferins, including 3-hydroxyhispidin, belong to the group of 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-ones and have the general formula:

where R is aryl or heteroaryl.

[008] For the vast majority of luciferins, the enzymes that ensure their synthesis in a living organism, as well as the reduction of oxyluciferin back to luciferin, are unknown. Thus, most applications of bioluminescence involve the addition of exogenous luciferin to a system (e.g., cell culture or organism) containing luciferase. The use of bioluminescent systems is thus limited due to a number of reasons: many luciferins penetrate the cell membrane poorly, luciferins themselves are chemically unstable, and their chemical synthesis is a complex multi-step and expensive process.

[009] The only bioluminescent system for which enzymes for luciferin synthesis have been identified is described in marine bacteria. This system is significantly different from other bioluminescent systems. Bacterial luciferin (myristic aldehyde) is oxidized during the reaction, but is not an emitter of bioluminescence [O. Shimomura, Bioluminescence: Chemical Principles and Methods, World Scientific Publishing Co. Pte. Ltd, Singapore, 2006, 470 p.]. In addition to luciferin, the key components of the luminescent reaction are NADH (nicotinamide adenine dinucleotide) and FMN-H2 (flavin mononucleotide), the oxidized derivative of which acts as a true light source. The bioluminescent system of marine bacteria is the only one today that can be completely encoded in a heterologous expression system and can be considered as the closest analogue of the present invention. However, this system is applicable primarily to prokaryotic organisms. To obtain autonomous bioluminescence, the luxCDABE operon is used, encoding luciferases (heterodimers luxA and luxB) and proteins for the biosynthesis of the bioluminescence substrate - luciferin luxCDE (Meighen 1991). In 2010, it was possible to achieve autonomous luminescence using this system in human cells - however, the low intensity of bioluminescence, only 12 times higher than the signal emanating from non-bioluminescent cells, did not allow the developed system to be used to solve most applied problems [Close et al. PloS One, 2010, 5(8):e12441]. Work to increase the intensity of emitted light has not been successful due to the toxicity of the components of the bacterial system to eukaryotic cells [Hollis et al. FEBS Letters, 2001, 506(2):140-42].

[010] Identification of enzymes that provide the synthesis of luciferin from stable and/or widely distributed precursor compounds in cells and the reduction of oxyluciferin back to luciferin seems to be an urgent task. The identification of such enzymes makes it possible to provide a simpler and cheaper method for the synthesis of luciferin and opens the way to the creation of autonomous bioluminescent systems. Of particular interest are bioluminescent systems that are non-toxic to eukaryotic cells.

The essence of the invention

[011] Applicants have deciphered the steps of luciferin biosynthesis in the fungal bioluminescent system and identified the enzymes involved in fungal luciferin cycling and the nucleic acid sequences encoding them.

[012] The stages of fungal luciferin turnover are shown in the diagram:

[013] Thus, the present invention primarily provides isolated fungal luciferin biosynthetic proteins as well as nucleic acids encoding them.

[014] In advantageous embodiments, the present invention provides hispidin hydroxylases having an amino acid sequence selected from the group of SEQ ID NOs: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, as well as essentially similar proteins, homologues, mutants and derivatives of the indicated hispidin hydroxylases.

[015] In some embodiments, the hispidin hydroxylases of the present invention are characterized by an amino acid sequence that, over at least 350 amino acids, has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity , for example, at least 80% identical, at least 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NO: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28.

[016] In some embodiments, the amino acid sequence of the hispidin hydroxylase of the present invention is characterized by the presence of several consensus sequences separated by non-conserved amino acid inserts, shown in SEQ ID NOs: 29-33.

[017] The hispidin hydroxylases of the present invention catalyze the reaction of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

,

to 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl.

[018] Also provided are hispidin synthases having an amino acid sequence selected from the group of SEQ ID NOs: 35, 37, 39, 41, 43, 45,47,49, 51, 53, 55, as well as substantially similar proteins, mutants , homologs and derivatives of these hispidin synthases.

[019] In some embodiments, the amino acid sequence of the hispidin synthase of the present invention is characterized by the presence of several consensus sequences separated by non-conserved amino acid inserts, shown in SEQ ID NOs: 56-63.

[020] In some embodiments, the hispidin synthases of the present invention are characterized by an amino acid sequence that has at least 40% identity, such as at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85% identity, at least 90% identity (for example, at least 90% , 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NOs: 35, 37, 39, 41 , 43, 45,47,49, 51, 53, 55.

[021] The hispidin synthases of the present invention catalyze the conversion reaction of 3-arylacrylic acid with the structural formula

,

where R is selected from the group aryl or heteroaryl in 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one having the structural formula

,

where R is aryl or heteroaryl.

[022] Also provided are caffeoyl pyruvate hydrolases having an amino acid sequence selected from the group of SEQ ID NOs: 65, 67, 69, 71, 73, 75, as well as substantially similar proteins, mutants, homologues and derivatives of these caffeoyl pyruvate hydrolases.

[023] In some embodiments, the amino acid sequence of the caffeoyl pyruvate hydrolase of the present invention is characterized by the presence of several consensus sequences separated by non-conserved amino acid inserts, shown in SEQ ID NOs: 76-78.

[024] In some embodiments, the amino acid sequence of the caffeoyl pyruvate hydrolase of the present invention has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, not less than 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identical) with an amino acid sequence selected from the group SEQ ID NOs: 65, 67, 69, 71, 73, 75.

[025] The caffeylpyruvate hydrolases of the present invention catalyze the reaction of 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acids having the structural formula

,

where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula

.

[026] In advantageous embodiments, the hispidin hydroxylases of the present invention catalyze the reaction of converting pre-luciferin into fungal luciferin, for example, hispidin into 3-hydroxyhispidin.

[027] In advantageous embodiments, the hispidin synthases of the present invention catalyze the conversion of a pre-luciferin precursor to pre-luciferin, for example, the conversion of caffeic acid to hispidin.

[028] In advantageous embodiments, the caffeoyl pyruvate hydrolases of the present invention catalyze the conversion of fungal oxyluciferin to a pre-luciferin precursor, for example, the conversion of caffeoyl pyruvate to caffeic acid.

[029] Also provided is the use of a protein whose amino acid sequence over at least 350 amino acids has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identical, at least 85% identical, at least 90% identical (e.g. at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, and/or the amino acid sequence of which contains separated by non-conservative amino acid insertions, consensus sequences shown in SEQ ID NOs: 29-33, in in vitro or in vivo systems as a hispidin hydroxylase catalyzing the reaction of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2- it has a structural formula,

to 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula ,

where R is aryl or heteroaryl.

[030] Also provided is the use of a protein whose amino acid sequence has at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or at least 70 % identity, or at least 75% identity, for example, at least 80% identity, at least 85% identity, at least 90% identity (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, and/or the amino acid sequence of which contains, separated by non-conservative amino acid insertions, the consensus sequences shown in SEQ ID NOs: 56-63, in in vitro or in vivo systems as hispidin synthase, catalyzing the reaction of conversion of 3-arylacrylic acid with the structural formula

to 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula ,

where R is aryl or heteroaryl.

[031] Also provided is the use of a protein whose amino acid sequence has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85 % identity, at least 90% identity (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) with amino acid sequence selected from the group of SEQ ID NOs: 65, 67, 69, 71, 73, 75, and/or the amino acid sequence of which contains, separated by non-conservative amino acid inserts, the consensus sequences shown in SEQ ID NOs: 76-78, in in vitro systems or in vivo as caffeoylpyruvate hydrolase, catalyzing the reaction of conversion of 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acids, having the structural formula ,

where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula .

[032] Nucleic acids encoding the above hispidin hydroxylases, hispidin synthases and caffeoylpyruvate hydrolases are also provided.

[033] In some embodiments, nucleic acids are provided encoding hispidin hydroxylases, the amino acid sequence of which is selected from the group:

(a) the amino acid sequence is shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28;

(b) the amino acid sequence has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85% identity, at least 90 % identity (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) to the amino acid sequence selected from SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, over at least 350 amino acids;

(c) the amino acid sequence contains the consensus sequences shown in SEQ ID NOs: 29-33.

[034] In some embodiments, nucleic acids are provided encoding hispidin synthases, the amino acid sequence of which is selected from the group:

(a) the amino acid sequence is shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55;

(b) the amino acid sequence has at least 40% identity, for example, at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or not less than 70% identical, or at least 75% identical, for example, at least 80% identical, at least 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94 %, 95%, 96%, 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55;

(c) the amino acid sequence contains the consensus sequences shown in SEQ ID NOs: 56-63.

[035] In some embodiments, nucleic acids are provided encoding caffeoyl pyruvate hydrolases, the amino acid sequence of which is selected from the group:

(a) the amino acid sequence is shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75;

(b) the amino acid sequence has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85% identity, at least 90 % identity (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) to the amino acid sequence selected from SEQ ID NOs: 65, 67, 69, 71, 73, 75;

(c) the amino acid sequence contains, separated by non-conservative amino acid inserts, the consensus sequences shown in SEQ ID NOs: 76-78.

[036] Also provided is the use of a nucleic acid encoding a protein whose amino acid sequence over at least 350 amino acids has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, or at least 80% identical, or at least 85% identical, or at least 90% identical, or at least 95% identical, (for example, at least 96%, 97%, 98%, 98% or 99% identical) with an amino acid sequence selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, or the amino acid sequence of which contains consensus sequences separated by non-conservative amino acid inserts, shown in SEQ ID NOs: 29-33, for the production in vitro or in vivo of hispidin hydroxylase catalyzing the reaction of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

,

to 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl.

[037] Also provided is the use of a nucleic acid encoding a protein whose amino acid sequence has at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or at least 70% identical, or at least 75% identical, or at least 80% identical, or at least 85% identical, or at least 90% identical, or at least 95% identical, (for example, at least 96% , 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, or amino acid sequence which contains consensus sequences separated by non-conservative amino acid inserts, shown in SEQ ID NOs: 56-63, for the production in vitro or in vivo of hispidin synthase, catalyzing the reaction of conversion of 3-aryl acrylic acid with the structural formula

to 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula ,

where R is aryl or heteroaryl.

[038] Also provided is the use of a nucleic acid encoding a protein whose amino acid sequence has at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, or at least 80% identity, or at least 85% identity, or at least 90% identity, or at least 95% identity, (e.g., at least 96%, 97%, 98%, 98%, or 99% identity) to an amino acid sequence selected from SEQ ID NOs: 65, 67, 69, 71, 73, 75, or the amino acid sequence of which contains, separated by non-conservative amino acid inserts, the consensus sequences shown in SEQ ID NOs: 76-78, for the production in vitro or in vivo of caffeoyl pyruvate hydrolase, catalyzing the conversion reaction of 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acids, having the structural formula ,

where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula .

[039] Also provided is a fusion protein comprising, operably linked directly or via amino acid linkers, at least one hispidin hydroxylase of the invention, and/or at least one hispidin synthase of the invention, and/or at least one caffeoyl pyruvate hydrolase of the invention. the invention, and an intracellular localization signal and/or a signal peptide and/or luciferase capable of oxidizing fungal luciferin to release light.

[040] Luciferase capable of oxidizing fungal luciferin to release light is known in the art. In advantageous embodiments, it has an amino acid sequence substantially similar or identical to an amino acid sequence selected from the group of SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. For example, it may have the amino acid sequence which is at least 40% identical, for example, at least 45% identical, or at least 50% identical, or at least 55% identical, or at least 60% identical, or at least at least 70% identical, or at least 75% identical, or at least 80% identical, or at least 85% identical to an amino acid sequence selected from the group SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. In many embodiments, the amino acid sequence of said luciferase has at least 90% identity, or at least 95% identity, (e.g., at least 96%, 97%, 98%, 98% or 99% identity) with an amino acid sequence selected from the group SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98.

[041] In some embodiments, the fusion protein has the amino acid sequence shown in SEQ ID NO: 101.

[042] A nucleic acid encoding the above fusion protein is also provided.

[043] Also provided is an expression cassette comprising (a) a transcription initiation region functional in a host cell; (b) a nucleic acid encoding a fungal luciferin biosynthetic enzyme, i.e. hispidin synthase, hispidin hydroxylase or caffeoylpyruvate hydrolase or a fusion protein according to the invention (c) a transcription termination region functional in a host cell.

[044] Also provided is a vector for transferring a nucleic acid into a host cell containing a nucleic acid encoding a fungal luciferin biosynthetic enzyme of the invention, that is, hispidin synthase, hispidin hydroxylase or caffeoylpyruvate hydrolase or a fusion protein of the invention.

[045] Also provided is a host cell containing, as part of an extrachromosomal element or integrated into the genome of the cell as a result of the introduction of the specified cassette into the specified cell, an expression cassette, which includes a nucleic acid encoding hispidin synthase and/or hispidin hydroxylase and/or caffeoylpyruvate hydrolase of the present invention. Such a cell produces at least one of the above enzymes for the biosynthesis of fungal luciferin due to the expression of the nucleic acid introduced into it.

[046] An antibody produced using the protein of the invention is also provided.

[047] Also provided is a method for producing mushroom luciferin, which is 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl, in an in vitro or in vivo system, comprising combining, under physiological conditions, with at least one molecule of the hispidin hydroxylase of the invention with at least one molecule of 6-(2-arylvinyl)-4-hydroxy-2H -pyran-2-one with structural formula ,

with at least one molecule of NAD(P)H and with at least one molecule of molecular oxygen.

[048] Also provided is a method for preparing fungal pre-luciferin, which is 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one with the structural formula ,

where R is aryl or heteroaryl, in an in vitro or in vivo system, involving the association, under physiological conditions, of at least one molecule of 3-arylacrylic acid with the structural formula

with at least one molecule of hispidin synthase according to the invention, with at least one molecule of coenzyme A, with at least one molecule of ATP and with at least two molecules of malonyl-CoA.

[049] Also provided is a method for producing fungal luciferin, in an in vitro or in vivo system, comprising combining, under physiological conditions, with at least one hispidin hydroxylase molecule of the invention with at least one molecule of 3-arylacrylic acid, with at least one a hispidin synthase molecule according to the invention, with at least one molecule of coenzyme A, with at least one molecule of ATP, with at least two molecules of malonyl-CoA, with at least one molecule of NAD(P)H and with at least at least one molecule of molecular oxygen.

[050] Methods for producing fungal luciferin and pre-luciferin can be implemented in a cell or organism, in which case the methods include introducing into the cell nucleic acids encoding the corresponding enzymes of luciferin biosynthesis (hispidin synthases and/or hispidin hydroxylases) capable of expressing these enzymes in a cell or organism. In advantageous embodiments, nucleic acids are introduced into a cell or organism as part of an expression cassette or vector of the invention.

[051] In some embodiments, a nucleic acid encoding a 4'-phosphopantotheinyl transferase capable of transferring 4-phosphopantetheinyl from coenzyme A to serine in the acyl-transfer domain of polyketide synthases is further introduced into the cell or organism. In some embodiments, the 4'-phosphopantotheinyl transferase has an amino acid sequence substantially similar or identical to SEQ ID NO: 105.

[052] Also provided is the use of a polyketide synthase (PKS) whose amino acid sequence is at least 40% identical to the sequence selected from the group SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139 , or by at least 45%, or by at least 50%, or by at least 55%, or by at least 60%, by at least 65%, or by at least 70%, or by at least 80%, or at least 85%, or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least by 94%, or by at least 95%, or by at least 96%, or by at least 97%, or by at least 98%, or by at least 99%, for obtaining hispidin in the in system vitro or in vivo.

[053] In some embodiments, a method for producing hispidin comprises combining, under physiological conditions, at least one PKS molecule with at least two malonyl-CoA molecules and at least one caffeoyl-CoA molecule. In some embodiments, the method includes combining, under physiological conditions, at least one PKS molecule with at least two malonyl-CoA molecules, at least one caffeic acid molecule, at least one coenzyme A molecule, at least one coumarate-CoA molecule ligase and at least one ATP molecule.

[054] Any coumarate-CoA ligase that catalyzes the reaction of caffeic acid to caffeoyl-CoA can be used for the purposes of the present invention. For example, coumarate-CoA ligase has an amino acid sequence that is at least 40% identical, or at least 45%, or at least 50% identical to the sequence shown in SEQ ID NO:141. 55%, or at least 60%, at least 65%, or at least 70%, or at least 80%, or at least 85%, or at least 90%, or by at least 91%, or by at least 92%, or by at least 93%, or by at least 94%, or by at least 95%, or by at least 96%, or by at least 97%, or at least 98%, or at least 99%.

[055] The reaction can be used in any methods instead of the reaction for producing fungal pre-luciferin from pre-luciferin precursors using the hispidin synthase of the present invention. For example, the reaction may be carried out in a cell or organism and involve introducing into the cell or organism an expression cassette containing a nucleic acid encoding a PKS. If necessary, a nucleic acid encoding a coumarate-CoA ligase is also introduced into the cell or organism.

[056] In some embodiments, nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid are additionally introduced into the cell or organism, for example, nucleic acids encoding tyrosine ammonium lyase, the amino acid sequence of which is substantially similar or identical to the amino acid sequence of tyrosine ammonium lyase Rhodobacter capsulatus shown in SEQ ID NO: 107, and nucleic acids encoding the HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase components, the amino acid sequences of which are substantially similar to the sequences of the HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase E components .coli shown in SEQ ID NOs: 109 and 111. In some embodiments, a nucleic acid encoding a phenylalanine ammonium lyase is used, the amino acid sequence of which is substantially similar to the amino acid sequence shown in SEQ ID NOs: 117.

[057] Methods for producing transgenic bioluminescent cells or organisms, including plant or animal cells or organisms, or bacteria or fungi, are also provided.

[058] In preferred embodiments, methods for producing transgenic bioluminescent cells or organisms include introducing into the cell or organism at least one nucleic acid of the invention, as well as a nucleic acid encoding a luciferase capable of oxidizing fungal luciferin to release light. Nucleic acids are introduced into a cell or organism in a form that allows for their expression and production of functional protein products. For example, nucleic acids are contained within an expression cassette. Nucleic acids are present in cells as part of an extrachromosomal element or integrated into the genome of a cell as a result of the introduction of an expression cassette into the specified cell.

[059] In preferred embodiments, methods for producing transgenic bioluminescent cells or organisms include introducing into the cell or organism a nucleic acid encoding a hispidin hydroxylase of the invention and a nucleic acid encoding a luciferase capable of oxidizing fungal luciferin to release light. The specified cell or organism acquires the ability to bioluminescence in the presence of fungal pre-luciferin, which is 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one with the structural formula ,

where R is aryl or heteroaryl.

[060] In some embodiments, instead of nucleic acids encoding hispidin synthase and luciferase, a nucleic acid encoding a hispidin hydroxylase-luciferase fusion protein is introduced into the cell.

[061] In some embodiments, methods for producing transgenic bioluminescent cells or organisms also include introducing into the cell or organism a nucleic acid encoding the hispidin synthase of the invention. The specified cell or organism acquires the ability to bioluminescence in the presence of a precursor of fungal preluciferin, which is 3-arylacrylic acid with the structural formula

,

where R is aryl or heteroaryl.

[062] In some embodiments, instead of a nucleic acid encoding hispidin synthase, a nucleic acid encoding PKS is introduced into the cell.

[063] In some embodiments, methods for producing transgenic bioluminescent cells or organisms also include introducing into the cell or organism a nucleic acid encoding the caffeoyl pyruvate hydrolase of the invention, which results in an increase in the intensity of bioluminescence.

[064] In some embodiments, methods for producing transgenic bioluminescent cells or organisms also include introducing into the cell or organism a nucleic acid encoding a 4'-phosphopantotheinyl transferase.

[065] In some embodiments, methods for producing transgenic bioluminescent cells or organisms also include introducing into the cell or organism a nucleic acid encoding a coumarate CoA ligase.

[066] In some embodiments, methods for producing transgenic bioluminescent cells or organisms also include introducing into the cell or organism nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid.

[067] Transgenic bioluminescent cells and organisms obtained using the methods described above and containing one or more nucleic acids of the invention as part of an extrachromosomal element or integrated into the genome of the cell are also provided.

[068] In some embodiments, the transgenic bioluminescent cells and organisms of the invention are capable of autonomous bioluminescence without the exogenous addition of luciferin, pre-luciferin, and pre-luciferin precursor.

[069] Combinations of proteins and nucleic acids of the invention, articles and kits containing proteins and nucleic acids of the invention are also provided. For example, combinations of nucleic acids are provided to produce autonomously luminous cells, cell lines or transgenic organisms, combinations for assaying promoter activity, or combinations for cell labeling.

[070] In some embodiments, kits are provided for the production of fungal luciferin and/or fungal pre-luciferin, comprising the above-described hispidin hydroxylase and/or hispidin synthase and/or PKS or nucleic acids encoding them.

[071] In some embodiments, kits are provided for producing a bioluminescent cell or bioluminescent transgenic organism, comprising a nucleic acid encoding a hispidin hydroxylase and a nucleic acid encoding a luciferase capable of oxidizing fungal luciferin to release light. The kit may also contain a nucleic acid encoding caffeoyl pyruvate hydrolase. The kit may also contain a nucleic acid encoding hispidin synthase or PKS. The kit may also contain a nucleic acid encoding 4'-phosphopantotheinyl transferase and/or a nucleic acid encoding coumarate-CoA ligase and/or nucleic acids encoding 3-arylacrylic acid biosynthetic enzymes. The kit may also contain additional components: buffer solutions, antibodies, fungal luciferin, fungal pre-luciferin, fungal pre-luciferin precursor, etc. The kit may also contain instructions for using the kit. In some embodiments, the nucleic acids are contained in an expression cassette or vector for introduction into cells or organisms.

[072] In advantageous embodiments, the cells and transgenic organisms of the invention are capable of producing fungal luciferin from precursors. In some embodiments, cells and transgenic organisms of the invention are capable of bioluminescence in the presence of a fungal luciferin precursor. In some embodiments, the cells and transgenic organisms of the invention are capable of autonomous bioluminescence.

[073] In the above methods and applications, in advantageous embodiments, the preluciferin used is 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one selected from the group:

(E)-6-(3,4-dihydroxystyryl)-4-hydroxy-2H-pyran-2-one (hispidin),

(E)-4-hydroxy-6-styryl-2H-pyran-2-one,

(E)-4-hydroxy-6-(4-hydroxystyryl)-2H-pyran-2-one (bisnoryangonin),

(E)-4-hydroxy-6-(2-hydroxystyryl)-2H-pyran-2-one,

(E)-4-hydroxy-6-(2,4-dihydroxystyryl)-2H-pyran-2-one,

(E)-4-hydroxy-6-(4-hydroxy-3,5-dimethoxystyryl)-2H-pyran-2-one,

(E)-4-hydroxy-6-(4-hydroxy-3-methoxystyryl)-2H-pyran-2-one,

(E)-4-hydroxy-6-(2-(6-hydroxynaphthalene-2-yl)vinyl)-2H-pyran-2-one,

(E)-6-(4-aminostyryl)-4-hydroxy-2H-pyran-2-one,

(E)-6-(4-(diethylamino)styryl)-4-hydroxy-2H-pyran-2-one,

(E)-6-(2-(1H-indol-3-yl)vinyl)-4-hydroxy-2H-pyran-2-one,

(E)-4-hydroxy-6-(2,3,6,7-tetrahydro-1H,5H-pyrido[3,2,1-ij]quinolin-9-yl)vinyl)-2H-pyran-2- He.

[074] In advantageous embodiments, 3-arylacrylic acid selected from the group: caffeic acid, cinnamic acid, paracoumaric acid, coumaric acid, umbellic acid, sinapic acid, ferulic acid is applicable to the needs of the present invention.

[075] In preferred embodiments, 3-hydroxyhispidin is used as luciferin, hispidin is used as pre-luciferin, and caffeic acid is used as pre-luciferin.

[076] The technical result consists in creating an effective method for producing autonomous bioluminescent systems with visible luminescence, including those based on eukaryotic non-luminous cells and organisms.

[077] Also, the technical result consists in the development of a new effective method for the synthesis of hispidin and its functional analogues.

[078] Also, the technical result consists in the development of a new effective method for the synthesis of fungal luciferin and its functional analogues.

[079] Also, the technical result consists in obtaining autonomously luminous cells and organisms.

[080] The technical result is achieved by identifying the stages of luciferin transformation in bioluminescent mushrooms and identifying the amino acid and nucleotide sequences of proteins involved in the biosynthesis of luciferin. The function of all proteins was demonstrated for the first time.

Brief description of drawings

[081] FIG. 1 shows a multiple amino acid sequence alignment of hispidin hydroxylases. The FAD/NAD(P)-binding domain is indicated by underlining. Consensus sequences are shown below the alignment.

[082] FIG. 2 shows a multiple alignment of the amino acid sequences of hispidin synthases. Consensus sequences are shown below the alignment.

[083] FIG. Figure 3 shows a multiple amino acid sequence alignment of caffeoylpyruvate hydrolases. Consensus sequences are shown below the alignment.

[084] FIG. 4 shows the luminescence intensity of Pichia pastoris cells expressing hispidin hydroxylase and luciferase (A) or only luciferase (B), or wild-type yeast (I), when colonies were sprayed with 3-hydroxyhispidin (luciferin, left graph) and hispidin (preluciferin, right schedule).

[085] FIG. 5 shows a comparison of the fluorescence intensity of HEK293NT cells expressing hispidin hydroxylase and luciferase and HEK293NT cells expressing only luciferase when hispidin was added.

[086] FIG. 6 shows the luminescence curve of HEK293T cells expressing (1) hispidin hydroxylase and luciferase genes separately upon addition of hispidin; (2) a gene for a chimeric protein of hispidin hydroxylase and luciferase with the addition of hispidin; (3) the gene of a chimeric protein of hispidin hydroxylase and luciferase with the addition of 3-hydroxyhispidin.

[087] FIG. Figure 7 illustrates the ability of transfected Pichia pastoris cells to undergo autonomous bioluminescence, in contrast to wild-type cells: on the Petri dish on the left are cells under daylight, on the right are cells in the dark.

[088] FIG. 8 shows the fluorescence of a culture of transfected Pichia pastoris cells in the dark.

[089] Fig. 9 shows autonomously bioluminescent transgenic Nicotiana benthamiana plants. The photo on the left was taken in ambient light, the photo on the right was taken in the dark.

Carrying out the invention

Definitions

[090] Various terms related to the subject matter of the present invention are used above and also in the description and claims. As used herein, the terms “includes” and “including” are interpreted to mean “including, but not limited to.” These terms are not intended to be construed as “consisting only of.”

[091] The terms “luminescence” and “bioluminescence” for the purposes of the present invention are interchangeable and refer to the phenomenon of light emission during a chemical reaction catalyzed by the enzyme luciferase.

[092] The terms “reactive,” “reacting,” and the like, with respect to the activity of a protein, mean that the protein is an enzyme that catalyzes said reaction.

[093] As used herein, the term “luciferase” means a protein that has the ability to catalyze the oxidation by molecular oxygen of a chemical compound (luciferin), where the oxidation reaction is accompanied by the release of light (luminescence or bioluminescence) and the release of oxidized luciferin.

[094] As used herein, the term "fungal luciferin" means a chemical compound selected from the group of 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-ones having the structural formula

[095] ,

where R is aryl or heteroaryl.

[096] Fungal luciferin is oxidized by a group of luciferases, hereinafter referred to as “luciferases capable of oxidizing fungal luciferin to release light” or the like. These luciferases are found in bioluminescent fungi; for example, they are described in application RU No. 2017102986/10(005203) dated January 30, 2017. The amino acid sequences of the luciferases useful in the methods and combinations of the present invention are substantially similar or identical to the amino acid sequences selected from SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. In many embodiments, of the present invention, the luciferases useful for the purposes of the present invention are characterized by amino acid sequences that are at least 40% identical, for example, at least 45% identical, or at least 50% identical, or at least 55% identical, or at least 60% identical, or at least 70% identical, or at least 75% identical, or at least 80% identical, or at least 85% identical to an amino acid sequence selected from SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. Luciferases are often characterized by amino acid sequences that have an amino acid sequence selected from the group SEQ ID NOs: 80, 82, 84, 86, 88,90, 92, 94, 96, 98, at least 90% identity (for example, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least at least 96 %, not less than 97%, not less than 98%, not less than 99% identity or 100% identity).

[097] Oxidation of mushroom luciferin produces "mushroom oxyluciferin", which is 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acid with the structural formula

[098] .

[099] The terms "fungal pre-luciferin" or simply "pre-luciferin" are used herein to refer to compounds belonging to the group of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-ones having the structural formula where R is aryl or heteroaryl. In a chemical reaction catalyzed by the enzyme of the present invention, pre-luciferin is converted into fungal luciferin.

[0100] The term “pre-luciferin precursor” is used herein to refer to compounds belonging to the group of 3-arylacrylic acids with the structural formula

[0101] ,

where R is aryl or heteroaryl. Pre-luciferins are formed from 3-arylacrylic acids in a chemical reaction catalyzed by the enzyme of the present invention.

[0102] Examples of fungal luciferins are shown in Table 1. Fungal luciferin-related examples of pre-luciferins, oxyluciferins, and pre-luciferin precursors are shown in Table 2.

[0103] The term "aryl" or "aryl substituent" refers to an aromatic radical in a single or fused carbocyclic ring system containing from five to fourteen ring members. In a preferred embodiment, the ring system contains from six to ten ring members. One or more hydrogen atoms may also be replaced by a substituent selected from acyl, acylamino, acyloxy, alkenyl, alkoxy, alkyl, alkynyl, amino, aryl, aryloxy, azido, carbamoyl, carboalkoxy, carboxy, carboxyamido, carboxyamino, cyano, disubstituted amino , formyl, guanidino, halogen, heteroaryl, heterocyclyl, hydroxy, iminoamino, monosubstituted amino, nitro, oxo, phosphonamino, sulfinyl, sulfonamino, sulfonyl, thio, thioacylamino, thioureido or ureido. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, biphenyl, terphenyl. Additionally, the term "aryl" as used herein includes groups in which an aromatic ring is linked to one or more non-aromatic rings.

[0104] The term "heterocyclic aromatic substituent", "heteroaryl substituent" or "heteroaryl" refers to an aromatic radical that contains from one to four heteroatoms or heterogroups selected from O, N, S or SO, in a single or fused heterocyclic ring system, containing from five to fifteen ring members. In a preferred embodiment, the heteroaryl ring system contains from six to ten ring members. One or more hydrogen atoms may also be replaced by a substituent selected from acyl, acylamino, acyloxy, alkenyl, alkoxy, alkyl, alkynyl, amino, aryl, aryloxy, carbamoyl, carboalkoxy, carboxy, carboxyamido, carboxyamino, cyano, disubstituted amino, formyl , guanidino, halogen, heteroaryl, heterocyclyl, hydroxy, iminoamino, monosubstituted amino, nitro, oxo, phosphonamino, sulfinyl, sulfonamino, sulfonyl, thio, thioacylamino, thioureido or ureido. Examples of heteroaryl groups include, but are not limited to, pyridinyl, thiazolyl, thiadiazolyl, isoquinolinyl, pyrazolyl, oxazolyl, oxadiazoyl, triazolyl and pyrrolyl groups. Additionally, the term “heteroaryl” as used herein includes groups in which a heteroaromatic ring is linked to one or more non-aromatic rings.

[0105] In the present invention, to designate chemical compounds, in addition to traditional names (if available), names are used in accordance with the international IUPAC nomenclature.

[0106] The term “luciferin biosynthetic enzyme” or “luciferin cycling enzyme” or the like is used to refer to an enzyme that catalyzes the reactions of converting a pre-luciferin precursor into pre-luciferin, and/or pre-luciferin into fungal luciferin, and/or oxyluciferin into a pre-luciferin precursor in in vitro and/or in vivo systems. Unless otherwise stated, luciferases are not included in the term “fungal luciferin biosynthetic enzymes.”

[0107] The term “hispidin hydroxylase” is used here to describe an enzyme that catalyzes the reaction of pre-luciferin to fungal luciferin, for example, the synthesis of 3-hydroxyhispidin from hispidin.

[0108] The term “hispidin synthase” is used here to describe an enzyme capable of catalyzing the synthesis of fungal pre-luciferin from a pre-luciferin precursor, for example, the synthesis of hispidin from caffeic acid.

[0109] The term "PKS" is used here to describe an enzyme belonging to the group of type III polyketide synthases capable of catalyzing the synthesis of hispidin from caffeoyl-CoA.

[0110] The term "caffeyl pyruvate hydrolase" is used here to describe an enzyme capable of catalyzing the cleavage of fungal oxyluciferin into simpler compounds, for example, to form a pre-luciferin precursor. An example of such a reaction is the conversion of caffeoyl pyruvate to caffeic acid.

[0111] The term "functional analogue" is used in the present invention to describe chemical compounds or proteins that perform the same function and/or can be used for the same purpose. For example, all of the fungal luciferins listed in Table 1 are functional analogues of each other.

[0112] The term "ATP" refers to adenosine triphosphate, which is the main energy carrier in the cell and has the structural formula:

.

[0113] The term "NAD(P)H" is used herein to refer to the reduced nicotinamide adenine dinucleotide phosphate (NADPH) or nicotinamide adenine dinucleotide (NADH) molecule. The term "NAD(P)" is used to refer to the oxidized form of nicotinamide adenine dinucleotide phosphate (NADH) or nicotinamide adenine dinucleotide (NADH). Nicotinamide adenine dinucleotide:

and nicotinamide adenine dinucleotide phosphate:

are dinucleotides built from nicotinic acid amide and adenine, interconnected by a chain consisting of two D-ribose residues and two phosphoric acid residues. NADP differs from NAD by containing another phosphoric acid residue attached to the hydroxyl of one of the D-ribose residues. Both compounds are widespread in nature and participate in many redox reactions, serving as carriers of electrons and hydrogen, which they accept from oxidized substances. The reduced forms transfer the resulting electrons and hydrogen to other substances.

[0114] The terms “coenzyme A” or “CoA” refers to a coenzyme well known from the prior art involved in the processes of oxidation and synthesis of fatty acids, biosynthesis of fats, oxidative transformations of carbohydrate breakdown products, with the structural formula:

[0115] The term "malonyl-CoA" refers to a derivative of coenzyme A formed during the synthesis of fatty acids and containing a malonic acid residue:

.

[0116] The term "coumaryl-CoA" refers to the thioester of coenzyme A and coumaric acid:

[0117] The term "caffeyl-CoA" refers to the thioester of coenzyme A and caffeic acid:

[0118] As used herein, the term “mutant” or “derivative” refers to a protein disclosed herein in which one or more amino acids have been added and/or substituted and/or deleted and/or inserted into the N- end and/or C-terminus, and/or within the native amino acid sequences of the proteins of the present invention. As used here, the term "mutant" refers to a nucleic acid molecule that encodes a mutant protein. In addition, the term "mutant" herein refers to any variant that is shorter or longer than the protein or nucleic acid disclosed in the present invention.

[0119] The term “homology” is used to describe the relationship of nucleotide or amino acid sequences to other nucleotide or amino acid sequences, which is determined by the degree of identity and/or similarity between the sequences being compared.

[0120] As used herein, an amino acid or nucleotide sequence is “substantially identical” or “substantially the same” as a reference sequence if the amino acid or nucleotide sequence has at least 40% identity with the specified sequence within the region selected for comparison. Thus, substantially similar sequences include those that have, for example, at least 40% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 62% identical, or at least 65% identical, or at least 70% identical, or at least 75% identical, e.g. at least 80% identical, at least 85% identity, at least 90% identity (for example, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity). Two sequences that are identical to each other are also essentially similar. For the purposes of the present invention, the length of the compared sequences is at least 100 amino acids or more, preferably at least 200 amino acids, for example 300 amino acids or more amino acids. In particular, it is possible to compare the amino acid sequences of full-length proteins. For nucleic acids, the length of the compared sequences is generally at least 300 nucleotides or more; preferably at least 600 nucleotides, including 900 or more nucleotides.

[0121] One example of an algorithm useful for determining percent sequence identity and sequence similarity is the BLAST algorithm described in Altschul et al., J. Mol. Biol. 215:403-410 (1990). Software for performing BLAST analyzes is available through the National Center for Biotechnology Information (http://www.ncbi.nlm.nih.gov/). This algorithm involves first finding pairs with the highest degree of identity (HIP) by identifying short words of length W in the test sequence that either completely match or satisfy some threshold positive value T when combined with a word of the same length from the sequence obtained in the database. T is the word proximity threshold (Altschul et al., 1990). These initial findings of word proximity (matches) serve as a seed to initiate a search for longer PVIs containing these words. These word matches are then expanded in both directions along each sequence as far as the cumulative match score can increase. Aggregates are calculated using (for nucleotide sequences) the parameters M (the bonus point awarded for a pair of matching residues; it is always >0) and N (the penalty point for mismatched residues; it is always <0). A scoring matrix is used to calculate the cumulative score for amino acid sequences. Expansion of word matches in each direction stops when the cumulative match score falls from the maximum achieved by an amount of X, when the cumulative score falls to zero or below zero due to the accumulation of one or more negative match scores, or when the end of any of the matches is reached. sequences. The W, T and X parameters of the BLAST algorithm determine the sensitivity and speed of registration. In the BLASTN program (for nucleotide sequences), the default word length (W) is set to 11, the expected value (E) is 10, the cutoff is 100, M=5, N=-4, and the comparison is performed on both strands. For amino acid sequences, BLASTP defaults to a word length (W) of 3, an expected value (E) of 10, and uses the BLOSUM62 scoring matrix (see Henikoff and Henikoff, Proc. Natl. Acad. Sci. USA 89: 10915 (1989)).

[0122] In addition to calculating percent sequence identity, the BLAST algorithm also performs statistical analysis of similarity between two sequences (see, for example, Karlin and Altschul, Proc. Nat'l. Acad. Sci. USA 90: 5873-5787 (1993)). One of the similarity definitions provided by the BLAST algorithm is the lowest cumulative probability (P(N)), which measures the probability with which a match between two nucleotide or amino acid sequences could occur by chance. For example, a test nucleic acid sequence is considered to be similar to a reference sequence if the lowest overall probability when comparing the test nucleic acid sequence to the reference nucleic acid sequence is less than 0.1, more preferably less than 0.01, and most preferably less than 0.001.

[0123] The term “consensus sequence” refers to the archetypal amino acid sequence to which all variants of a particular protein or sequence of interest are compared. Consensus sequences and methods for their determination are well known to those skilled in the art. For example, a consensus sequence is determined by multiple comparisons of known homologous proteins by identifying the amino acids that occur most frequently at a given position across the entire population of related sequences.

[0124] The term “conserved sequence” is used to refer to a nucleotide sequence in nucleic acids or an amino acid sequence in a polypeptide chain that does not change at all or changes slightly in different organisms during evolution. Accordingly, a “non-conserved sequence” is a sequence that varies significantly among the organisms being compared.

[0125] The term "amino acid insert" means one or more amino acids within a polypeptide chain that are located between the protein fragments in question (protein domains, linkers, consensus sequences). It will be apparent to those skilled in the art that the fragments and amino acid insertions discussed are operatively linked to form a single polypeptide chain.

[0126] Any software known in the art can be used to determine the domain structure of a protein. For example, SMART (Simple Modular Architecture Research Tool) software, available on the Internet at http://smart.embl-heidelberg.de [Schultz et al., PNAS 1998; 95:5857-5864; Letunic I, Doerks T, Bork P Nucleic Acids Res 2014; doi:10.1093/nar/gku949].

[0127] The term "operably linked" or the like when describing fusion proteins refers to polypeptide sequences that are in physical and functional relationship with one another. In most preferred embodiments, the functions of the polypeptide components of the chimeric molecule are not altered compared to the functional properties of the isolated polypeptide components. For example, the hispidin hydroxylase of the present invention can be operatively ligated to a fusion partner of interest, such as luciferase. In this case, the fusion protein retains the properties of hispidin hydroxylase, and the polypeptide of interest retains its original biological activity - for example, the ability to oxidize luciferin to release light. In some embodiments of the present invention, the activities of the fusion partners may be reduced compared to the activities of the isolated proteins. Such fusion proteins also find use within the scope of the present invention.

[0128] The term "operably linked" or the like when describing nucleic acids means that the nucleic acids are covalently linked such that there are no reading frame errors or stop codons at their junctions. As is obvious to anyone skilled in the art, the nucleotide sequences encoding a fusion protein including "operably linked" components (proteins, polypeptides, linker sequences, amino acid inserts, protein domains, etc.) consist of fragments encoding these components , where these fragments are covalently linked in such a way that during transcription and translation of the nucleotide sequence a full-length fusion protein is produced.

[0129] When describing the association of a nucleic acid with regulatory coding sequences (promoters, enhancers, transcription terminators), the term "operably linked" means that the sequences are arranged and linked in such a way that the regulatory sequence will affect the level of expression of the coding or nucleic acid sequence.

[0130] In the context of the present invention, “linking” nucleic acids means that two or more nucleic acids are joined together using any methods known in the art. By way of non-limiting example, nucleic acids can be ligated together using, for example, a DNA ligase or annealed using PCR. Nucleic acids can also be joined by chemical nucleic acid synthesis using a sequence of two or more individual nucleic acids.

[0131] The terms “regulatory elements” or “regulatory sequences” refer to sequences included in controlling the expression of an encoding nucleic acid. Regulatory elements include a promoter, termination signals, and other sequences that influence nucleic acid expression. Typically they also cover sequences required for proper translation of the nucleotide sequence.

[0132] The term “promoter” is used to describe the non-translated and non-transcribed DNA sequence upstream of the coding region and containing the RNA polymerase binding site and initiating DNA transcription. The promoter region may also include other elements that act as regulators of gene expression.

[0133] As used here, the term “functional” means that the nucleotide or amino acid sequence can function for the specified test or task. The term "functional" used to describe luciferases means that the protein has the ability to produce the oxidation reaction of luciferin accompanied by luminescence. When describing hispidin hydroxylases, the term “functional” means that the protein has the ability to catalyze the reaction of converting at least one of the pre-luciferins shown in Table 2 into the corresponding luciferin. When describing hispidin synthases, the term "functional" means that the protein has the ability to catalyze the conversion of at least one of the precursors of pre-luciferins to pre-luciferin, for example, the conversion of caffeic acid to hispidin. When describing caffeoylpyruvate hydrolases, the term “functional” means that the protein has the ability to catalyze the reaction of at least one of the oxyluciferins to a pre-luciferin precursor (eg, the conversion of caffeoylpyruvate to caffeic acid).

[0134] As used herein, the term “enzymatic properties” refers to the ability of a protein to catalyze a particular chemical reaction.

[0135] As used herein, the term “biochemical properties” refers to protein folding and maturation rate, half-life, rate of catalysis, pH and temperature stability, and other similar properties.

[0136] As used herein, “spectral properties” refer to spectra, quantum yield and luminescence intensity and other similar properties.

[0137] Reference to a nucleotide sequence “coding” a polypeptide means that the nucleotide sequence produces the polypeptide during mRNA transcription and translation. In this case, both a coding strand, identical to mRNA and usually used in the sequence list, and a complementary strand, which is used as a template for transcription, can be indicated. As will be apparent to anyone skilled in the art, the term also includes any degenerate nucleotide sequences encoding the same amino acid sequence. The nucleotide sequences encoding the polypeptide include sequences containing introns.

[0138] The terms "expression cassette" or "expression cassette" are used herein to mean a nucleic acid sequence capable of directing the expression of a particular nucleotide sequence in a corresponding host cell. Typically, an “expression cassette” contains a heterologous nucleic acid encoding a protein or its functional fragment, operably linked to a promoter and termination signals. Typically it also contains sequences required for proper translation of the significant nucleotide sequence. The expression cassette may be one that occurs naturally (including in host cells) but has been produced in a recombinant form useful for the expression of a heterologous nucleic acid. Often the "expression cassette" is heterologous to the host, i.e. the particular nucleic acid sequence of this expression cassette does not occur naturally in the host cell, and must be introduced into the host cell or host cell precursor by transformation. Expression of a nucleotide sequence may be under the control of a constitutive promoter or an inducible promoter, which initiates transcription only when the host cell is exposed to a particular external stimulus. In the case of a multicellular organism, the promoter may also be specific to a particular tissue or organ or developmental stage.

[0139] “Heterologous” or “exogenous” nucleic acid means a nucleic acid not present in a wild-type host cell.

[0140] The term “endogenous” refers to a native protein or nucleic acid in its natural position in the genome of an organism.

[0141] As used herein, the term "specifically hybridizes" refers to the association between two single-stranded nucleic acid molecules or sufficiently complementary sequences to permit such hybridization under predetermined conditions commonly used in the art (the term "substantially complementary" is sometimes used ).

[0142] An “isolated” nucleic acid molecule or isolated protein is a nucleic acid molecule or protein that, due to human action, exists separately from its natural environment and is therefore not a product of nature. An isolated nucleic acid molecule or an isolated protein may exist in a purified form or may exist in a non-natural environment, such as (but not limited to) a recombinant prokaryotic cell, a plant cell, an animal cell, a non-bioluminescent fungal cell, a transgenic organism (fungus, plant , animal), etc.

[0143] "Transformation" is a process for introducing a heterologous nucleic acid into a host cell or organism. Specifically, “transformation” refers to the stable integration of a DNA molecule into the genome of the organism of interest.

[0144] "Transformed/transgenic/recombinant" refers to a host organism, such as a bacterium, or plant, or fungus, or animal, into which a heterologous nucleic acid molecule has been introduced. This nucleic acid molecule may be stably integrated into the host genome, or this nucleic acid molecule may also be present as an extrachromosomal molecule. Such an extrachromosomal molecule may be capable of self-replication. It should be understood that transgenic or stably transformed cells, tissues or organisms include not only the end products of the transformation process, but also their transgenic progeny. The terms “non-transformed,” “non-transgenic,” or “non-recombinant,” or “wild type” refer to a natural host organism or host cell, such as a bacterium or plant, that does not contain a heterologous nucleic acid molecule.

[0145] The term "autonomously luminous" or "autonomously bioluminescent" refers to transgenic organisms and host cells that have the ability to bioluminesce without exogenous addition of luciferins, pre-luciferins, or pre-luciferin precursors.

[0146] The term "4'-phosphopantotheinyl transferase" is used herein to refer to the enzyme that transfers 4-phosphopantotheinyl from coenzyme A to serine in the acyl-transfer domain of polyketide synthase. 4'-phosphopantotheinyl transferases are expressed in nature by many plants and fungi and are known from the prior art [Gao Menghao et al., Microbial Cell Factories 2013, 12:77]. It will be apparent to one skilled in the art that any functional variant of 4'-phosphopantotheinyl transferase can be used for the purposes of the present invention. For example, the 4'-phosphopantotheinyl transferase NpgA from Aspergillus nidulans (SEQ ID NO 104, 105) described in [Gao Menghao et al., Microbial Cell Factories 2013, 12:77] or its homolog or mutant, i.e. a protein whose amino acid sequence is substantially similar or identical to the sequence shown in SEQ ID NO: 105. For example, a 4'-phosphopantotheinyl transferase having at least 40% identity with the sequence of SEQ ID NO: 105 can be used, including at least 50% identical, or at least 55% identical, or at least 60% identical, or at least 62% identical, or at least 65% identical, or at least 70% identity, or at least 75% identity, for example, or at least 80% identity, or at least 85% identity, or at least 90% identity (for example, 90%, 91%, 92% , 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity).

[0147] Nucleotides are designated by their bases using the following standard abbreviations: adenine (A), cytosine (C), thymine (T), and guanine (G). Similarly, amino acids are designated by the following standard abbreviations: alanine (Ala; A), arginine (Arg; R), asparagine (Asn; N), aspartic acid (Asp; D), cysteine (Cys; C), glutamine (Gln; Q ), glutamic acid (Glu; E), glycine (Gly; G), histidine (His; H), isoleucine (He; 1), leucine (Leu; L), lysine (Lys; K), methionine (Met; M ), phenylalanine (Phe; F), proline (Pro; P), serine (Ser; S), threonine (Thr; T), tryptophan (Trp; W), tyrosine (Tyr; Y) and valine (Val; V) .

[0148] The present invention is directed to new enzymes for the biosynthesis of fungal luciferin, encoding their nucleic acids, the use of proteins as enzymes that catalyze steps in the biosynthesis of fungal luciferin. The invention also provides the use of nucleic acids to obtain said enzymes in a cell or organism. Methods for producing fungal luciferin and pre-luciferin chemical compounds in in vitro and in vivo systems are also provided. Also provided are vectors comprising the nucleic acid described in the present invention. In addition, the present invention provides expression cassettes comprising the nucleic acid of the present invention and regulatory elements necessary for expression of the nucleic acid in a selected host cell. Also provided are cells, stable cell lines, transgenic organisms (eg, plants, animals, fungi, microorganisms) incorporating the nucleic acids, vectors, or expression cassettes of the present invention. Combinations of nucleic acids to produce autonomously luminescent cells, cell lines or transgenic organisms are also provided. In advantageous embodiments, cells and transgenic organisms are capable of producing fungal luciferin from precursors. In some embodiments, cells and transgenic organisms are capable of producing fungal pre-luciferin from precursors. In some embodiments, cells and transgenic organisms are capable of bioluminescence in the presence of a fungal luciferin precursor. In some embodiments, cells and transgenic organisms are capable of autonomous bioluminescence. Protein combinations are also provided to produce luciferin and its precursors from simpler chemical compounds. Also provided is a kit containing nucleic acids or vectors or expression cassettes of the present invention for producing luminous cells, cell lines or transgenic organisms.

Squirrels

[0149] As stated above, the present invention provides proteins involved as enzymes in the biosynthesis (cyclic system of transformations) of fungal luciferin.

[0150] The proteins of the invention can be obtained from natural sources or produced using recombinant technologies. For example, wild-type proteins can be isolated from bioluminescent fungi, for example from fungi belonging to the phylum Basidiomycota, predominantly the class Basidiomycetes, in particular the order Agaricales. For example, wild-type proteins can be isolated from the fungi Neonothopanus nambi, Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos, etc. The proteins of the present invention can also be produced by expressing a recombinant nucleic acid coding sequence of the protein in an appropriate host or cell-free expression system, as described in the Nucleic Acids section. In some embodiments, the proteins are used within host cells into which expressible nucleic acids encoding the proteins have been introduced.

[0151] In advantageous embodiments, the claimed proteins fold rapidly after expression in a host cell. Rapid folding means that proteins reach their tertiary structure, which provides their enzymatic properties, after a short period of time. In these embodiments, the proteins are folded over a period of time that generally does not exceed about 3 days, typically does not exceed about 2 days, and more often does not exceed about 12-24 hours.

[0152] In some embodiments, the proteins are used in isolated form. Any conventional techniques may be used for protein purification, where suitable methods for protein purification are described in the Guide to Protein Purification, (Deuthser ed., Academic Press, 1990). For example, the lysate can be prepared from the original source and purified using HPLC, size exclusion chromatography, gel electrophoresis, affinity chromatography, and the like.

[0153] When the proteins of the present invention are in isolated form, this means that the protein is substantially free from the presence of other proteins or other natural biological molecules, such as oligosaccharides, nucleic acids and fragments thereof, etc., where the term "substantially free" as used herein means that less than 70%, typically less than 60%, and more often less than 50% of the composition containing the isolated protein is another naturally occurring biological molecule. In some embodiments, the proteins are present in substantially purified form, wherein the term "substantially purified form" means a purity of at least 95%, typically at least 97%, and more typically at least 99%.

[0154] The proteins of the present invention remain active at temperatures below 50°C, more often at temperatures up to 45°C, that is, they remain active at temperatures of 20-42°C and can be used in heterologous expression systems in vitro and in vivo.

[0155] The claimed proteins have a pH stability in the range from 4 to 10, more often in the range from 6.5 to 9.5. The optimum pH stability of the claimed proteins lies in the range between 6.8 and 8.5, for example between 7.3-8.3.

[0156] The claimed proteins are active under physiological conditions. The term "physiological conditions" in this invention means an environment having a temperature in the range of 20 to 42°C, a pH in the range of 6.8 to 8.5, saline and an osmolarity of 300-400 mOsmol/L. In particular, the term "physiological conditions" includes the intracellular environment, cell extract and fluids extracted from living organisms, such as blood plasma. "Physiological conditions" can be created artificially. For example, by combining known chemical compounds, reaction mixtures can be created that provide “physiological conditions”. Methods for creating such environments are well known in the art. Non-limiting examples include:

[0157] 1) Ringer's solution, isotonic to mammalian blood plasma.

[0158] Ringer's solution consists of 6.5 g NaCl, 0.42 g KCl and 0.25 g CaCl ₂ dissolved in 1 liter of double-distilled water. When preparing a solution, salts are added sequentially, each subsequent salt is added only after the previous one has dissolved. To prevent precipitation of calcium carbonate, it is recommended to pass carbon dioxide through the sodium bicarbonate solution. The solution is prepared with fresh distilled water.

[0159] 2) Versene solution

[0160] Versene solution is a mixture of EDTA and inorganic salts, dissolved in distilled water or water for injection, sterilized by membrane filtration using filters with a final pore size of 0.22 microns. 1 liter of Versene solution contains NaCl 8.0 g, KCl 0.2 g, sodium phosphate disubstituted 12-aqueous 1.45 g, potassium phosphate disubstituted 0.2 g, sodium salt of ethylenediaminetetraacetic acid 0.2 g, bidistilled water - up to 1 l. Versene solution must have a buffer capacity of at least 1.4 ml. The content of chlorine ion is from 4.4 to 5.4 g/l, EDTA is not less than 0.6 mmol/l.

[0161] 3) phosphate buffered saline (PBS, Na-phosphate buffer)

[0162] Na-phosphate buffer consists of 137 mM NaCl, 10 mM Na ₂ HPO ₄ , 1.76 mM KH ₂ PO ₄ . The buffer may also contain KCl at concentrations up to 2.7 mM. To prepare 1 liter of one-time sodium phosphate buffer, use: 8.00 g NaCl, 1.44 g Na ₂ HPO ₄ , 0.24 g KH ₂ PO ₄ , 0.20 g KCl (optional). Dissolve in 800 ml of distilled water. The required pH is adjusted with hydrochloric acid or sodium hydroxide. Next, bring the volume to 1 liter with distilled water.

[0163] Specific proteins of interest are enzymes involved in the cyclic biosynthesis of fungal luciferin, their mutants, homologues and derivatives. Each of these specific types of polypeptide structures of interest will now be discussed individually in more detail.

[0164] Hispidin hydroxylases

[0165] The hispidin hydroxylases of the present invention are proteins that have the ability to catalyze the synthesis of luciferin from pre-luciferin. In other words, these are enzymes that catalyze the conversion reaction of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, which has the structural formula

,

to 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl.

[0166] The reaction is carried out under physiological conditions in in vitro and in vivo systems in the presence of at least one molecule of NAD(P)H and at least one molecule of molecular oxygen (O2) per molecule of 6-(2-arylvinyl)-4 -hydroxy-2H-pyran-2-one:

[0167] Hispidin hydroxylases of interest include proteins from the bioluminescent fungi Neonothopanus nambi, Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos, amino acids ny sequences which are shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, as well as their functional mutants, homologues and derivatives.

[0168] In advantageous embodiments, the hispidin hydroxylases of the present invention are characterized by the presence of a FAD/NAD(P) binding domain, IPR002938 - code from the InterPro public database, available on the Internet at http://www. ebi.ac.uk/interpro). This domain is involved in the binding of flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD) in a variety of hydroxyl-substrate addition enzymes found in the metabolic pathways of many organisms. The hispidin hydroxylases of the present invention contain a specified domain of 350-385 amino acids in length, usually 360-380 amino acids, for example 364-377 amino acids, flanked by N- and C-terminal non-conservative amino acid sequences having a lower percentage identity with each other. The position of the FAD/NAD binding domain of the claimed hispidin hydroxylases is indicated in the multiple amino acid sequence alignment of the individual proteins in Figure 1.

[0169] Also provided are homologs or mutants of hispidin hydroxylases, the sequence of which differs from the above-described specific amino acid sequences claimed in the invention, that is, SEQ ID NO: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28. The homologues or mutants of interest have at least 40% identity, e.g., 45% identity, or 50% identity, or 55% identity, or 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85% identity, at least 90% identity (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity) with a protein whose amino acid sequence is selected from the group SEQ ID NOs: 2, 4 , 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, over at least 350 amino acids. This especially applies to the amino acid sequences that provide the functional regions of the protein, that is, to the sequence of the FAD/NAD-binding domain included in the hispidin hydroxylases.

[0170] In advantageous embodiments, the amino acid sequence of the hispidin hydroxylase of the present invention is characterized by the presence of several conserved amino acid motifs (consensus sequences) characteristic only of this group of enzymes. These consensus sequences are shown in SEQ ID NOs: 29-33. Consensus regions within the amino acid sequences of hispidin hydroxylases are operatively linked through amino acid insertions with less conservation.

[0171] Hispidin synthases

[0172] The hispidin synthases of the present invention are proteins that have the ability to catalyze the synthesis of pre-luciferin from its precursors. In other words, these are enzymes that catalyze the reaction of conversion of 3-arylacrylic acid with the structural formula

,

where R is aryl or heteroaryl in 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl.

[0173] Examples of 3-arylacrylic acids that serve as precursors for pre-luciferins are shown in Table 2.

[0174] The reaction is carried out under physiological conditions in in vitro and in vivo systems in the presence of at least one molecule of coenzyme A, at least one molecule of ATP and at least two molecules of malonyl-CoA:

[0175] Hispidin synthases of interest include proteins from the bioluminescent fungi Neonothopanus nambi, Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos, amino acids ny sequences which are shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, as well as their functional mutants, homologues and derivatives.

[0176] In advantageous embodiments, the amino acid sequence of the hispidin synthase of the present invention is characterized by the presence of several conserved amino acid motifs (consensus sequences) characteristic only of this group of enzymes. These consensus sequences are shown in SEQ ID NOs: 56-63. Consensus regions within the amino acid sequences of hispidin synthases are operatively linked through amino acid insertions with less conservation.

[0177] In many embodiments of the present invention, the amino acid sequences of interest of homologues and mutants of specific hispidin synthases have significant identity to the sequences shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51. 53, 55, which is, for example, at least 40% identity, for example, at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity , or at least 70% identical, or at least 75% identical, for example, at least 80% identical, at least 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) throughout the amino acid sequence of the protein.

[0178] In advantageous embodiments, the hispidin synthases of the present invention are multidomain proteins belonging to the polyketide synthase superfamily. In advantageous embodiments, the hispidin synthases of the present invention are subject to post-translational modification, namely, their maturation requires the transfer of 4-phosphopantetheinyl from coenzyme A to serine in the acyl-transfer domain of the polyketide synthase. Enzymes known from the prior art are 4'-phosphopantotheinyl transferases that carry out such modification [Gao Menghao et al., Microbial Cell Factories 2013, 12:77]. 4'-phosphopantotheinyl transferases are expressed in nature by many plants and fungi, in the cells of which the maturation of the functional hispidin synthase of the present invention occurs without the introduction of additional enzymes or nucleic acids encoding them. At the same time, the maturation of hispidin synthase in the cells of a number of lower fungi (for example, yeast) and animals requires the introduction of the 4'-phosphopantotheinyl transferase coding sequence into the host cells. It will be apparent to one skilled in the art that any functional variant of 4'-phosphopantotheinyl transferase known in the art can be used for the purposes of the present invention. For example, the 4'-phosphopantotheinyl transferase NpgA from Aspergillus nidulans (SEQ ID NO 104, 105) described in [Gao Menghao et al., Microbial Cell Factories 2013, 12:77], any homolog or mutant with confirmed activity, can be used .

[0179] Caffeylpyruvate hydrolases

[0180] The caffeylpyruvate hydrolases of the present invention are proteins that have the ability to catalyze the conversion of oxyluciferin, which is a 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acid having the structural formula

[0181] ,

where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula

[0182] ,

where R is aryl or heteroaryl.

[0183] Examples of oxyluciferins are shown in Table 2.

[0184] The reaction is carried out under physiological conditions in in vitro and in vivo systems:

[0185] In advantageous embodiments, the caffeoyl pyruvate hydrolases of the present invention convert caffeoyl pyruvate to caffeic acid. In advantageous embodiments, they convert the oxyluciferin shown in Table 2 into a pre-luciferin precursor.

[0186] Caffeylpyruvate hydrolases of interest include proteins from the bioluminescent fungi Neonothopanus nambi, Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos , amino acid sequences which are shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75, as well as their functional mutants, homologues and derivatives.

[0187] In advantageous embodiments, the amino acid sequence of the caffeoylpyruvate hydrolases of the present invention (including homologs and mutants of interest) is characterized by the presence of several conserved amino acid motifs (consensus sequences) unique to this group of enzymes. These consensus sequences are shown in SEQ ID NOs: 76-78. Consensus regions within the amino acid sequences of caffeoylpyruvate hydrolases are operatively linked through amino acid insertions with less conservation.

[0188] In many embodiments of the present invention, the amino acid sequences of interest for caffeoyl pyruvate hydrolases have significant identity to the sequences shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75, which is, for example, at least 40% identity , for example, at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity , for example, at least 80% identical, at least 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) throughout the entire amino acid sequence of the protein.

[0189] Homologs of the specific proteins described above (i.e., proteins with amino acid sequences SEQ ID NO: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 35, 37, 39 , 41, 43, 45,47,49, 51, 53, 55, 65, 67, 69, 71, 73, 75) can be isolated from natural sources. Homologues can be found in many organisms (fungi, plants, microorganisms, animals). In particular, homologs can be found in various species of bioluminescent fungi, in particular from fungi belonging to the phylum Basidiomycota, mainly to the class Basidiomycetes, in particular to the order Agaricales. Also of particular interest as a source of protein homologs of the present invention are non-bioluminescent hispidin-producing fungi and plants, such as Pteris ensiformis [Yung-Husan Chen et al., “Identification of phenolic antioxidants from Sword Brake fern (Pteris ensiformis Burm.),” Food Chemistry, Volume 105, Issue 1, 2007, pp.48-56], Inonotus xeranticus [In-Kyoung Lee et al., “Hispidin Derivatives from the Mushroom Inonotus xeranticus and Their Antioxidant Activity,” J. Nat. Prod., 2006, 69 (2), pp.299-301], Phellinus sp. [In-Kyoung Lee et al., “Highly oxygenated and unsaturated metabolites providing a diversity of hispidin class antioxidants in the medicinal mushrooms Inonotus and Phellinus” . Bioorganic & Medicinal Chemistry. 15 (10): 3309-14.], Equisetum arvense [Markus Herderich et al., “Establishing styrylpyrone synthase activity in cell free extracts obtained from gametophytes of Equisetum arvense L. by high performance liquid chromatography-tandem mass spectrometry.” Phytochem. Anal., 8: 194-197].

[0190] Also provided are proteins that are derivatives or mutants of the above-described naturally occurring proteins. Mutants and derivatives may retain the biological properties of wild-type proteins (eg, those found in nature), or may have biological properties that differ from wild-type proteins. Mutations include substitution of one or more amino acids, deletion or insertion of one or more amino acids, substitution or truncation or extension of the N-terminus, substitution or truncation or extension of the C-terminus, and the like. Mutants and derivatives can be generated using standard molecular biology techniques, as detailed in the Nucleic Acids section. Mutants are essentially identical to wild-type proteins, that is, they have at least 40% identity with them within the region chosen for comparison. Thus, substantially similar sequences include those that have, for example, at least 40% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 62% identical, or at least 65% identical, or at least 70% identical, or at least 75% identical, e.g. at least 80% identical, or at least 85 % identity, or at least 90% identity (for example, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) within the selected for regional comparison. In many embodiments, the homologs of interest have much higher sequence identity, e.g., 70%, 75%, 80%, 85%, 90% (e.g., 92%, 93%, 94%) or higher, e.g., 95% , 96%, 97%, 98%, 99%, 99.5%, especially for amino acid sequences that provide functional regions of the protein.

[0191] Derivatives can also be made using standard methods and include modification by RNA, chemical modifications, post-translation and post-transcription modifications, and the like. For example, derivatives can be made by methods such as altered phosphorylation or glycosylation, or acetylation , or lipidation, or various types of cleavage during ripening, etc.

[0192] Methods well known to those skilled in the art are used to search for functional mutants, homologues and derivatives. For example, functional screening of an expression library containing variants (eg, mutant forms of proteins, or homologous proteins or protein derivatives) can be used. An expression library is prepared by cloning the nucleic acids encoding the protein variants being tested into an expression vector and introducing them into suitable host cells. Methods for working with nucleic acids are described in more detail in the “Nucleic Acids” section. To detect the functional enzymes of the present invention, a suitable substrate is added to cells expressing the nucleic acids being tested. The occurrence of the expected reaction product catalyzed by the functional enzyme can be detected by high performance liquid chromatography techniques using synthetic versions of the expected reaction products as standards. For example, to identify functional hispidin hydroxylases, hispidin or another pre-luciferin shown in Table 2 can be used as a substrate. The expected reaction product is fungal luciferin. To detect hispidin synthases, a pre-luciferin precursor (for example, caffeic acid) can be used as a substrate, and the reaction product is the corresponding fungal pre-luciferin. It should be taken into account that to screen for functional hispidin synthases, host cells must express 4'-phosphopantotheinyl transferase, which ensures post-translational modification of the protein.

[0193] To search for functional caffeoylpyruvate hydrolases, oxyluciferin is used as a reaction substrate (Table 2), and the reaction product tested is a pre-luciferin precursor, 3-arylacrylic acid.

[0194] In many embodiments of the present invention, a bioluminescence reaction can be used to search for functional enzymes of the present invention. In this case, to prepare an expression library, cells are used that produce luciferase, which is capable of oxidizing fungal luciferin with the release of light, and functional enzymes that ensure the production of fungal luciferin from the product of the enzymatic reaction carried out by the protein being analyzed.

[0195] Thus, to screen for functional hispidin hydroxylases, host cells that produce functional luciferase, the subrate of which is fungal luciferin, are used. When pre-luciferin is added to cells containing a functional variant of hispidin hydroxylase, fungal luciferin is formed and luminescence occurs due to the oxidation reaction of fungal luciferin by luciferase.

[0196] Screening for functional hispidin synthases uses host cells that additionally produce a functional luciferase, the substrate of which is fungal luciferin, and a functional hispidin hydroxylase. When a luciferin precursor is added to such cells, fungal luciferin is formed and luminescence occurs due to the oxidation reaction of fungal luciferin by luciferase.

[0197] Screening for functional caffeoylpyruvate hydrolases uses host cells that produce a functional luciferase whose substrate is fungal luciferin, a functional hispidin hydroxylase, and a functional hispidin synthase. When oxyluciferin is added to such cells, fungal luciferin is formed and luminescence occurs due to the oxidation reaction of fungal luciferin by luciferase.

[0198] Any luciferase capable of oxidizing, releasing light, luciferin selected from the group of 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-ones with the general formula can be used for screening ,

where R is aryl or heteroaryl. Non-limiting examples of luciferins are shown in Table 1. Non-limiting examples of suitable luciferases are described in the "Uses, Combinations and Methods of Use" section below.

[0199] The oxidation reaction of luciferins by luciferase is accompanied by the release of detectable light. The light produced by the reaction can be detected by conventional methods (e.g., visual inspection, night vision, spectrophotometry, spectrofluorimetry, photographic image recording, specialized luminescence and fluorescence detection equipment such as e.g. IVIS Spectrum In Vivo Imaging System (Perkin Elmer), etc.). The detected light can be emitted in a range of intensities from one photon to light easily visible to the eye, for example, with an intensity of 1 cd, and bright light with an intensity, for example, 100 cd, or more. The light emitted during the oxidation of 3-hydroxyhispidine is in the range from 400 to 700 nm, most often in the range from 450 to 650 nm, with a maximum emission at 520-590 nm. The light emitted by the oxidation of other 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-ones may have a shift in the emission maximum (Table 3).

Table 3. Emission maxima for a number of fungal luciferins Substance Emission maximum, nm 3-hydroxyhispidin 538 (E)-3,4-dihydroxy-6-(4-hydroxystyryl)-2H-pyran-2-one 520 (E)-6-(2-(1H-indol-3-yl)vinyl)-3,4-dihydroxy-2H-pyran-2-one, 480 (E)-6-(4-(diethylamino)styryl)-3,4-dihydroxy-2H-pyran-2-one 504 (E)-3,4-dihydroxy-6-(2-(2,3,6,7-tetrahydro-1H,5H-pyrido[3,2,1-ij]quinolin-9-yl)vinyl)-2H -piran-2-on, 534 (E)-3,4-dihydroxy-6-(2-(6-hydroxynaphthalene-2-yl)vinyl)-2H-pyran-2-one 564

[0200] Examples of performing functional screening using bioluminescence are described in the experimental section below.

[0201] The invention also covers fusion proteins including the protein of the present invention. Its homologue, mutant, including a shortened or elongated form. The protein of the invention can be operatively fused to an intracellular localization signal (for example, a nuclear localization signal, a localization signal in mitochondria, or peroxisomes, or lysosomes, or the Golgi apparatus, or in other cellular organelles), a signal peptide that ensures the release of the protein into the intercellular space , transmembrane domain, or with any protein or polypeptide (fusion partner) of interest. Fusion proteins may include operably linked, for example, hispidin hydroxylase and/or hispidin synthase and/or caffeoylpyruvate hydrolase of the invention with a fusion partner attached at the C- or N-terminus. Non-limiting examples of fusion partners may include proteins of the present invention having other enzymatic function, antibodies or binding fragments thereof, ligands or receptors, luciferases capable of using fungal luciferins as substrates in a bioluminescent reaction. In some embodiments, the fusion partner and the protein of the invention are operatively linked through a linker sequence (peptide linker) allowing independent folding and function of the fusion protein. Methods for making fusion proteins are well known to those skilled in the art.

[0202] In some embodiments, the fusion proteins include a hispidin hydroxylase of the invention operably linked through a short peptide linker and a luciferase capable of oxidizing fungal luciferin to release light. Such a fusion protein can be used to produce bioluminescence in in vitro and in vivo systems in the presence of pre-luciferin (eg, in the presence of hispidin). One skilled in the art will appreciate that any functional hispidin hydroxylase described above can be used with any functional luciferase to create a fusion protein. Specific examples of fusion proteins are described below in the Experimental Section. Examples of luciferases that can be used in the creation of fusion proteins are described in the Applications, Combinations and Uses section below.

Nucleic acids

[0203] The present invention provides nucleic acids encoding fungal luciferin biosynthetic enzymes, homologs and mutants of these proteins, including truncated and extended forms.

[0204] A nucleic acid as used herein is an isolated DNA molecule, such as a genomic DNA molecule or cDNA molecule, or an RNA molecule, such as an mRNA molecule. In particular, said nucleic acids are cDNA molecules having an open reading frame that encodes the luciferin biosynthetic enzyme of the invention and is capable, under suitable conditions, of causing expression of the enzyme of the invention.

[0205] The term "cDNA" is intended to describe nucleic acids that reflect the arrangement of sequence elements found in native mature mRNA, where the sequence elements are exons and 5' and 3' non-coding regions. Immature mRNA may have exons separated by intervening introns, which, if present, are removed by post-translational RNA splicing to produce mature mRNA having an open reading frame.

[0206] The genomic sequence of interest may include the nucleic acid present between the start codon and the stop codon, as defined in the listed sequences, including all introns that are normally present on the native chromosome. The genomic sequence of interest may further include the 5' and 3' untranslated regions found in the mature mRNA, as well as specific transcriptional and translational control sequences such as promoters, enhancers, etc., including flanking genomic DNA approximately 1 kb in size, but possibly larger, at the 5' or 3' end of the transcribed region.

[0207] The invention also covers nucleic acids that are homologous, substantially similar, identical, derivatives or mimetics of the nucleic acids encoding proteins of the present invention.

[0208] The claimed nucleic acids are present in an environment other than their natural environment; for example, they are isolated, present in enriched quantities, or present or expressed in vitro or in a cell or organism other than their naturally occurring environment.

[0209] Specific nucleic acids of interest include nucleic acids that encode hispidin hydroxylase, or hispidin synthase, or caffeoylpyruvate hydrolase, described in the "Proteins" section above. Each of these specific types of nucleic acids of interest is individually disclosed in more detail.

[0210] Nucleic acids encoding hispidin hydroxylases.

[0211] In advantageous embodiments, the nucleic acids of the invention encode proteins capable of catalyzing the conversion reaction of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one (pre-luciferin) having the structural formula

,

c 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one (fungal luciferin), having the structural formula

,

[0212] where R is aryl or heteroaryl.

[0213] In preferred embodiments, the nucleic acids encode hispidin hydroxylases, the amino acid sequences of which are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NO: 29-33.

[0214] Specific examples of nucleic acids include nucleic acids encoding hispidin hydroxylases, the amino acid sequences of which are shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28. Examples of nucleic acids encoding these proteins are given in SEQ ID NOs: 1, 3.5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27. Also of interest are functional mutants homologues and derivatives of the above specific nucleic acids.

[0215] In advantageous embodiments, the nucleic acids of the invention encode proteins whose amino acid sequences are at least 60%, or at least 65%, or at least 70%, or at least 80%, or at least at least 85%, or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99% identical to the sequences shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, over at least 350 amino acids.

[0216] Nucleic acids encoding hispidin synthases

[0217] In advantageous embodiments, the nucleic acids of the invention encode proteins capable of catalyzing the conversion reaction of 3-arylacrylic acid with the structural formula

[0218] ,

where R is aryl or heteroaryl in 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

[0219] ,

where R is aryl or heteroaryl.

[0220] In preferred embodiments, the nucleic acids encode hispidin synthases, the amino acid sequences of which are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 56-63.

[0221] Specific examples of nucleic acids include nucleic acids encoding the hispidin synthases of the invention, the amino acid sequences of which are shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55. Examples nucleic acids encoding these proteins are given in SEQ ID NOs: 34, 36,38,40,42, 44, 46, 48, 50, 52, 54.

[0222] Also of interest are functional mutants, homologues and derivatives of the above specific nucleic acids.

[0223] In advantageous embodiments, the nucleic acids of the invention encode proteins that are at least 45%, typically at least 50%, such as at least 55%, or at least 60%, or at least 65%, or at least 70%, or at least 80%, or at least 85%, or at least 90%, or at least 91%, or at least by 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98 %, or at least 99% identical to the sequences shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45,47,49, 51, 53, 55, throughout the entire polypeptide chain of the protein.

[0224] Nucleic acids encoding caffeoylpyruvate hydrolases

[0225] In advantageous embodiments, the nucleic acids of the invention encode proteins capable of catalyzing the conversion reaction of oxyluciferin having the structural formula

[0226] ,

where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula

[0227] ,

where R is selected from the group aryl, heteroaryl.

[0228] In preferred embodiments, the nucleic acids encode caffeoyl pyruvate hydrolases, the amino acid sequences of which are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 76-78. Specific examples of nucleic acids include nucleic acids encoding caffeoyl pyruvate hydrolases, the amino acid sequences of which are shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75. Examples of nucleic acids encoding these proteins are shown in SEQ ID NOs: 64. 66, 68, 70,72, 74.

[0229] Also of interest are nucleic acids encoding functional mutants, homologs and derivatives of the above proteins.

[0230] In advantageous embodiments, the nucleic acids of the invention encode proteins whose amino acid sequences are at least 60%, or at least 65%, or at least 70%, or at least 80%, or at least at least 85%, or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99% identical to the sequences shown in SEQ ID NOs: 65, 67, 69, 71 , 73, 75, throughout the entire polypeptide chain.

[0231] Nucleic acids of interest (e.g., nucleic acids encoding homologues of proteins characterized by the amino acid sequences shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 35, 37, 39, 41, 43, 45,47,49, 51, 53,55, 65, 67, 69, 71, 73, 75), can be isolated from any organisms (fungi, plants, microorganisms, animals), in particular from various types of bioluminescent fungi, for example, from fungi belonging to the phylum Basidiomycota, mainly to the class Basidiomycetes, in particular to the order Agaricales, for example, from bioluminescent fungi Neonothopanus nambi, Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos, etc. Also of particular interest as a source of nucleic acids encoding homologs of the proteins of the present invention are non-bioluminescent hispidin-producing fungi and plants, such as Pteris ensiformis [Yung-Husan Chen et al., “Identification of phenolic antioxidants from Sword Brake fern (Pteris ensiformis Burm .)", Food Chemistry, Volume 105, Issue 1, 2007, pp.48-56], Inonotus xeranticus [In-Kyoung Lee et al., "Hispidin Derivatives from the Mushroom Inonotus xeranticus and Their Antioxidant Activity", J. Nat . Prod., 2006, 69 (2), pp.299-301], Phellinus sp. [In-Kyoung Lee et al., “Highly oxygenated and unsaturated metabolites providing a diversity of hispidin class antioxidants in the medicinal mushrooms Inonotus and Phellinus” . Bioorganic & Medicinal Chemistry. 15 (10): 3309-14.], Equisetum arvense [Markus Herderich et al., “Establishing styrylpyrone synthase activity in cell free extracts obtained from gametophytes of Equisetum arvense L. by high performance liquid chromatography-tandem mass spectrometry.” Phytochem. Anal., 8: 194-197].

[0232] Homologs are identified in any of a variety of ways. The cDNA fragment of the present invention can be used as a hybridization probe against a cDNA library from a target organism using low stringency conditions. The probe may be a large fragment or one or more short degenerate primers. Nucleic acids having sequence similarity are detected by hybridization under low stringency conditions, for example, at 50°C and 6xSSC (0.9M sodium chloride/0.09M sodium citrate) followed by washing at 55xC in 1xSSC (01 .15M sodium chloride/0.015M sodium citrate). Sequence identity can be determined by hybridization under high stringency conditions, for example, 50°C or higher and 0.1×SSC (15 mM sodium chloride/1.5 mM sodium citrate). Nucleic acids having a region substantially identical to the presented sequences, for example, allelic variants, genetically altered nucleic acid variants, etc., bind to the presented sequences under conditions of high hybridization stringency. Using probes, in particular labeled DNA sequence probes, homologous or similar genes can be isolated.

[0233] Homologues can be identified by polymerase chain reaction from a genomic or cDNA library. Oligonucleotide primers representing fragments of known sequences of specific nucleic acids can be used as primers for PCR. In a preferred aspect, the oligonucleotide primers have a degenerate structure and correspond to nucleic acid fragments encoding conserved regions of the amino acid sequence of a protein, for example. consensus sequences shown in SEQ ID NOs: 29-33, 56-63, 76-78. Full-length coding sequences can then be detected using 3' and 5' RACE methods well known in the art.

[0234] Homologues can also be identified in the results of whole genome sequencing of organisms by comparing the amino acid sequences predicted from the sequencing with the amino acid sequences SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 35, 37, 39, 41, 43, 45,47,49, 51, 53,55, 65, 67, 69, 71, 73, 75. Sequence identity is determined based on the reference sequences. Algorithms for sequence analysis are known in the art, such as BLAST, described in Altschul et al., J. Mol. Biol., 215, pp.403-10 (1990). For purposes of the present invention, comparison of nucleotide and amino acid sequences using the Blast software package available from the National Center for Biotechnology Information (http://www.ncbi.nlm.nih) can be used to determine the level of identity and similarity between nucleotide sequences and amino acid sequences. .gov/blast) using a gap-containing alignment with standard parameters.

[0235] Also provided are nucleic acids that hybridize with the above-described nucleic acids under stringent conditions, preferably under high stringency conditions (ie, complementary to the previously described nucleic acids). An example of hybridization under stringent conditions is at 50°C or higher and 0.1xSSC (15 mM sodium chloride/1.5 mM sodium citrate). Another example of hybridization under high stringency conditions is overnight incubation at 42°C in 50% formamide, 5x SSC (150 mM NaCl, 15 mM trisodium citrate), 50 mM sodium phosphate (pH 7.6), 5x solution Denhardt, 10% dextran sulfate, and 20 μg/ml denatured, sheared salmon seminal fluid DNA, prewashed in 0.1 x SSC at approximately 65°C. Other high stringency hybridization conditions are known in the art and may also be used to detect the nucleic acids of the invention.

[0236] Nucleic acids encoding variants, mutants or derivatives of the proteins of the invention are also provided. Mutants or derivatives can be produced on a template nucleic acid selected from the nucleic acids described above by modifying, deleting or adding one or more nucleotides in the template sequence or a combination thereof to produce a variant template nucleic acid. Modifications, additions or deletions can be made by any method known in the art (see, for example, Gustin et al., Biotechniques (1993) 14: 22; Barany, Gene (1985) 37: 111-123; and Colicelli et al. ., Mol. Gen. Genet. (1985) 199:537-539, Sambrook et al., Molecular Cloning: A Laboratory Manual, (1989), CSH Press, pp.15.3-15.108), including error-prone PCR, permutation, oligonucleotide-directed mutagenesis, assembly PCR, pairwise PCR mutagenesis, in vivo mutagenesis, cassette mutagenesis, recursive ensemble mutagenesis, exponential multiple mutagenesis, site-specific mutagenesis, nonspecific mutagenesis, gene reassembly, gene site-saturating mutagenesis (GSSM), artificial reassembly with ligation (SLR) or combinations thereof. Modifications, additions or deletions can also be made by methods including recombination, recursive sequence recombination, phosphorothioate-modified DNA mutagenesis, uracil-containing template mutagenesis, double-skip mutagenesis, mismatch repair point mutagenesis, repair-deficient strain mutagenesis, chemical mutagenesis, radioactive mutagenesis, deletion mutagenesis, restriction selective mutagenesis, restriction mutagenesis with purification, synthesis with artificial genes, multiple mutagenesis, creation of chimeric multiple nucleic acids and combinations thereof. Nucleic acids encoding shortened and extended versions of these luciferases are also included in the scope of the present invention. As used here, these protein variants contain amino acid sequences with altered C-, N-, or both ends of the polypeptide chain.

[0237] In advantageous embodiments, the homologs and mutants discussed are functional enzymes capable of performing fungal luciferin biosynthetic reactions, e.g., fungal luciferin. Homologues and mutants of interest may have altered properties, such as the rate of maturation in the host cell, the ability to aggregate or dimerize, half-life or other biochemical properties, including substrate binding constant, thermostability, pH stability, temperature optimum activity, pH optimum of activity, Michaelis-Menten constant, substrate specificity, range of by-products. In some embodiments, homologs and mutants have the same properties as the claimed proteins.

[0238] Nucleic acids encoding functional homologs and mutants of the present invention can be determined through functional tests, for example, in the functional expression library screen described above in the "Proteins" section.

[0239] In addition, degenerate nucleic acid variants that encode the proteins of the present invention are also provided. Degenerate nucleic acid variants involve substitutions of nucleic acid codons with other codons encoding the same amino acids. In particular, degenerate nucleic acid variants are created to increase expression in the host cell. In this embodiment, nucleic acid codons that are not or are less preferred in genes in the host cell are replaced by codons that are overrepresented in coding sequences in genes in the host cell, wherein said replaced codons encode the same amino acid. In particular, humanized versions of the nucleic acids of the present invention are of interest. As used herein, the term “humanized” refers to substitutions made in a nucleic acid sequence to optimize codons for protein expression in mammalian cells (Yang et al., Nucleic Acids Research (1996) 24: 4592-4593). See also US Patent No. 5,795,737, which describes the humanization of proteins, the disclosure of which is incorporated by reference herein. Of particular interest are nucleic acid variants optimized for expression in plant cells. Examples of such nucleic acids encoding proteins of the present invention are shown in SEQ ID NOs: 103, 113 and 114.

[0240] The subject nucleic acids can be isolated and obtained in substantially purified form. Essentially, purified form means that the nucleic acid is at least about 50% pure, typically at least about 90% pure, and is typically “recombinant,” that is, flanked by one or more nucleotides to which it does not normally bind on a chromosome found naturally in its natural host.

[0241] The subject nucleic acids may be artificially synthesized. Methods for producing nucleic acids are well known in the art. For example, the availability of amino acid sequence information or nucleotide sequence information makes it possible to produce the isolated nucleic acid molecules of the present invention using oligonucleotide synthesis. In the case of amino acid sequence information, several nucleic acids can be synthesized that differ from each other due to the degeneracy of the genetic code. Methods for selecting codon variants for a desired host are well known in the art.

[0242] Synthetic oligonucleotides can be prepared using the phosphoramidite method, and the resulting constructs can be purified using methods well known in the art, such as high performance liquid chromatography (HPLC) or other methods as described, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual, ^2nd Ed., (1989) Cold Spring Harbor Press, Cold Spring Harbor, NY, and as described in, for example, United States Dept. of HHS, National Institute of Health (NIH) Guidelines for Recombinant DNA Research. Long double-stranded DNA molecules of the present invention can be synthesized as follows: several smaller fragments with the required complementarity can be synthesized, which contain suitable ends capable of cohesion with an adjacent fragment. Adjacent fragments can be stitched together using DNA ligase, recombination-based methods, or PCR-based methods.

[0243] Nucleic acids encoding fusion proteins are also provided, including the proteins of the present invention. Examples of such proteins are given above in the “Proteins” section. Nucleic acids encoding fusion proteins can be synthesized artificially as described above.

[0244] Also provided are expression cassettes or systems used inter alia to produce the claimed proteins (i.e., hispidin hydroxylases, hispidin synthases, and caffeoylpyruvate hydrolases) or fusion proteins based thereon, or to replicate the claimed nucleic acid molecules. The expression cassette may exist as an extrachromosomal element or may be incorporated into the genome of a cell by introducing the expression cassette into the cell. When an expression cassette is introduced into a cell, a protein product encoded by the nucleic acid of the invention is formed; in this case the protein is said to be "produced" or "expressed" by the cell. Any expression system is suitable, including, for example, bacterial systems, yeast, plants, insects, amphibians or mammalian cells. In an expression cassette, the target nucleic acid is operably linked to regulatory sequences, which may include promoters, enhancers, termination sequences, operators, repressors and inducers. Typically, the expression cassette contains at least (a) a transcription initiation region functional in the host cell; (b) a nucleic acid according to the invention; and (c) a transcription termination region functional in the host cell. Methods for preparing expression cassettes or systems capable of expressing a desired product are known to those skilled in the art.

[0245] Vector and other nucleic acid constructs containing the claimed nucleic acids are also provided. Suitable vectors include viral and non-viral vectors, plasmids, cosmids, phages, etc., preferably plasmids, and are used to clone, amplify, express, transfer, etc., the nucleic acid sequence of the present invention into a suitable host. The selection of a suitable vector is readily apparent to one skilled in the art. The full-length nucleic acid or a portion thereof is typically inserted into a vector by DNA ligase attachment to a restriction enzyme-cleaved site in the vector. Alternatively, the desired nucleotide sequence can be inserted by homologous recombination in vivo, typically by joining homologous regions to the vector flanking the desired nucleotide sequence. Homologous regions are added by ligation of oligonucleotides or polymerase chain reaction, using primers including, for example, both homologous regions and part of the desired nucleotide sequence. The vector, as a rule, has an origin of replication that ensures its reproduction in host cells as a result of its introduction into the cell as an extrachromosomal element. The vector may also contain regulatory elements that ensure expression of the nucleic acid in the host cell and production of a recombinant functional protein. In an expression vector, said nucleic acid is operably linked to a regulatory sequence, which may include promoters, enhancers, terminators, operators, repressors, silencers, insulators and inducers. To express functional proteins or their truncated forms, the nucleic acids encoding them are operatively linked to nucleic acids containing at least regulatory sequences and a transcription start site. These nucleic acids may also contain sequences encoding a histidine tag (6 His tag), signal peptide, or functional protein domains. In many embodiments, the vectors enable the integration of nucleic acid operably linked to regulatory elements into the genome of the host cell. The vector may contain an expression cassette for a selectable marker, such as a fluorescent protein (for example, gfp), an antibiotic resistance gene (for example, a gene for resistance to amphicillin, or kanamycin, or neomycin or hygromycin, etc.), genes conferring resistance to herbicides, such as genes conferring resistance to phosphinothricin and sulfonamide herbicides, or other selectable marker known in the art.

[0246] The vector may contain additional expression cassettes including nucleic acids encoding 4'-phosphopantotheinyl transferase, 3-arylacrylic acid synthesis proteins (for example, those described in the Applications, Combinations and Uses section), luciferases, etc.

[0247] The expression systems described above can be used in prokaryotic or eukaryotic hosts. To obtain the protein, host cells such as E. coli, B. subtilis, S. cerevisiae, insect cells, or cells of a higher organism other than human embryonic cells, such as yeast, plants (for example, Arabidopsis thaliana, Nicotiana benthamiana) can be used , Physcomitrella patens), vertebrates, e.g. COS 7 cells, HEK 293, CHO, Xenopus oocytes, etc.

[0248] Cell lines that stably produce the proteins of the present invention can be selected by methods known in the art (for example, co-transfection with a selectable marker such as dhfr, gpt, antibiotic resistance genes (ampicillin, or kanamycin, or neomycin or hygromycin, etc.), which makes it possible to identify and isolate transfected cells that contain a gene included in the genome or existing as part of an extrachromosomal element.

[0249] If any of the above host cells or other suitable host cells or organisms for replicating and/or expressing the nucleic acids of the invention are used, then the resulting replicated nucleic acid, expressed protein or polypeptide is within the scope of the invention as a product of the host cell or organism . The product can be isolated by a suitable method known in the art.

[0250] In many embodiments of the present invention, multiple expression cassettes containing nucleic acids of the invention encoding various fungal luciferin biosynthetic enzymes are co-transfected into a cell. In some embodiments, an expression cassette containing a nucleic acid encoding a luciferase capable of oxidizing fungal luciferin to release light is further introduced into the cell. In some cases, expression cassettes are combined into a single vector, which is used to transform cells. In some embodiments, nucleic acids encoding 4'-phosphopantotheinyl transferase and/or 3-arylacrylic acid synthesis proteins are additionally introduced into the cell.

[0251] Short DNA fragments of the claimed nucleic acids are also provided, which are used as primers for PCR, rolling circle amplification, hybridization screening assays, etc. Long DNA fragments are used to obtain encoded polypeptides as previously described. However, for use in geometric amplification reactions such as PCR, a pair of short DNA fragments, i.e. primers, are used. The exact sequence of the primer is not critical to the invention, but for most applications the primers will hybridize to the claimed sequence under conditions of stringency as known in the art. It is preferable to select a primer pair that will produce an amplification product of at least about 50 nucleotides, preferably at least about 100 nucleotides, and may span the entire nucleic acid sequence. Algorithms for selecting primer sequences are generally known and available in commercial software packages. Amplification primers hybridize to complementary DNA strands and will prime counter amplification reactions.

[0252] The nucleic acid molecules of the present invention can also be used to determine gene expression in a biological sample. The technique, which examines cells for the presence of specific nucleotide sequences, such as genomic DNA or RNA, is well established in the field. Briefly, DNA or mRNA is isolated from a cell sample. The mRNA can be amplified by RT-PCR, using reverse transcriptase to form a complementary DNA strand, followed by polymerase chain reaction amplification using primers specific to the reported DNA sequences. Alternatively, the mRNA sample is separated by gel electrophoresis, transferred to a suitable support such as nitrocellulose, nylon, etc., and then tested with a fragment of the claimed DNA as a probe. Other methods may also be used, such as oligonucleotide cross-linking assays, in situ hybridization, and hybridization with DNA probes immobilized on a solid chip. Detection of mRNA hybridizing to the reported sequence indicates gene expression in the sample.

Transgenic organisms

[0253] Transgenic organisms, transgenic cells, and transgenic cell lines expressing the nucleic acids of the present invention are also provided. Transgenic cells according to the present invention include one or more nucleic acids contemplated in the present invention, which are present as a transgene. For purposes of the present invention, any suitable host cell can be used, including prokaryotic (eg, Escherichia coli, Streptomyces sp., Bacillus subtilis, Lactobacillus acidophilus, etc.) or eukaryotic host cells other than human embryonic cells. Transgenic organisms of the present invention may be prokaryotic or eukaryotic organisms, including bacteria, cyanobacteria, fungi, plants and animals, into which one or more cells of the organism containing a heterologous nucleic acid of the present invention is introduced by insertion through human manipulation, e.g. , within the framework of transgenic techniques known in the art.

[0254] In one embodiment of the present invention, the transgenic organism may be a prokaryotic organism. Methods for transforming prokaryotic host cells are well known in the art (see, for example, Sambrook et al. (Molecular Cloning: A Laboratory Manual, 2nd Ed., (1989) Cold Spring Harbor Laboratory Press and Ausubel et al., Current Protocols in Molecular Biology (1995) John Wiley & Sons, Inc).

[0255] In another embodiment of the present invention, said transgenic organism may be a fungus, such as yeast. Yeast is widely used as a vehicle for heterologous gene expression (see, for example, Goodey et al., Yeast biotechnology, D R Berry et al., eds, (1987) Allen and Unwin, London, pp. 401-429, and Kong et al., Molecular and Cell Biology of Yeasts, E. F. Walton and G. T. Yarronton, eds, Blackie, Glasgow (1989) pp.107-133). Several types of yeast vectors are available, including integrating vectors that require recombination with the host genome for maintenance, as well as autonomously replicating plasmid vectors.

[0256] Another host organism is an animal organism. Transgenic animals can be produced using transgenic techniques known in the art and described in standard manuals (such as: Pinkert, Transgenic Animal Technology: A Laboratory Handbook, 2nd edition (2003) San Diego: Academic Press; Gersenstein and Vinterstein, Manipulating the Mouse Embryo: A Laboratory Manual, 3rd ed, (2002) Nagy A. (Ed), Cold Spring Harbor Laboratory; Blau et al., Laboratory Animal Medicine, 2nd Ed., (2002) Fox J.G., Anderson L.C., Loew F.M., Quimby F.W. (Eds), American Medical Association, American Psychological Association; Gene Targeting: A Ptactical Approach by Alexandra L. Joyner (Ed.) Oxford University Press; 2nd edition (2000)). For example, transgenic animals can be produced through homologous recombination, which changes the endogenous locus. Alternatively, the nucleic acid construct is randomly integrated into the genome. Vectors for stable integration include plasmids, retroviruses and other animal viruses, YAC, and the like.

[0257] The nucleic acid can be introduced into a cell, directly or indirectly, by introduction into a cell precursor, by careful genetic manipulation such as microinjection, or by infection with a recombinant virus or using a recombinant viral vector, transfection, transformation, delivery using a gene gun or transconjugation. Techniques for transferring nucleic acid molecules (eg, DNA) into such organisms are well known and are described in standard manuals such as Sambrook et al. (Molecular Cloning: A Laboratory Manual, 3rd Ed., (2001) Cold Spring Harbor Press, Cold Spring Harbor, NY).

[0258] The term "genetic manipulation" does not include classical crossbreeding or in vitro fertilization, but rather refers to the introduction of a recombinant nucleic acid molecule. Said nucleic acid molecule may be integrated into a chromosome or may be extrachromosomally replicating DNA.

[0259] DNA constructs for homologous recombination include at least a portion of a nucleic acid of the present invention, wherein the nucleic acid of the invention is operatively linked to regions of homology to a target locus. The DNA design for random integration does not need to include regions of homology to facilitate recombination. Positive and negative selection markers may also be included. Methods for producing cells containing target gene modifications through homologous recombination are known in the art. Various methods for transfecting mammalian cells are described, for example, in Keown et al., Meth. Enzymol. (1990) 185:527-537).

[0260] In the case of embryonic stem (ES) cells, an ES cell line may be used or embryonic cells may be obtained fresh from a host such as a mouse, rat, guinea pig, etc. Such cells are grown in an appropriate fibroblast culture layer or grown in the presence of leukemia inhibitory factor (LIF). Transformed ES or embryonic cells can be used to create transgenic animals using appropriate techniques known in the art.

[0261] Transgenic animals can be any non-human animals, including a non-human mammal (eg, mouse, rat), bird or amphibian, etc., and are used in functional studies, drug screening, etc. .P.

[0262] Transgenic plants can also be produced. Methods for obtaining transgenic plant cells and plants are described in patents No. US 5767367, US5750870, US5739409, US5689049, US5689045, US5674731, US5656466, US5633155, US5629470, US5595896, US5576198, US55388 79 and US5484956, the description of which is incorporated herein by reference. Methods for producing transgenic plants are also summarized in the following reviews: Plant Biochemistry and Molecular Biology (eds. Lea and Leegood, John Wiley & Sons (1993) pp. 275-295 and in Plant Biotechnology and Transgenic Plants (eds. Oksman-Caldentey and Barz) (2002) 719 p.

[0263] To obtain a transgenic host organism, for example, embryogenic explants containing somatic cells can be used. Once the cells or tissues are collected, the exogenous DNA of interest is introduced into the plant cells, and many different techniques are available for such introduction. Once isolated protoplasts are available, the possibility of introduction using DNA-mediated gene transfer protocols arises, involving incubation of protoplasts with deproteinized DNA, such as a plasmid, including an exogenous coding sequence of interest, in the presence of multivalent cations (eg, PEG or poly-L-ornithine) ; or by electroporation of protoplasts in the presence of naked DNA containing the exogenous sequence of interest. Next, protoplasts that have successfully taken up exogenous DNA are selected, grown to form callus, and ultimately transgenic plants are produced by contact with appropriate quantities and proportions of stimulating factors such as auxins and cytokinins.

[0264] Other suitable plant production methods may be used, such as gene gun approaches or Agrobacterium-mediated transformation, known to those skilled in the art.

Antibodies

[0265] The term “antibody” is used herein to refer to a polypeptide or group of polypeptides comprising at least one active site of an antibody (antigen-binding site). The term “antigen-binding site” refers to the spatial structure, surface parameters and charge distribution of which are complementary to the parameters of the antigen epitope: this ensures the binding of the antibody to the corresponding antigen. The term "antibody" includes, for example, vertebrate antibodies, chimeric antibodies, hybrid antibodies, humanized antibodies, modified antibodies, monovalent antibodies, Fab fragments and single domain antibodies.

[0266] Antibodies specific for the proteins of the present invention are useful in affinity chromatography, immunoassays, and in the detection and identification of proteins of the invention (hispidin hydroxylases, hispidin synthases, and caffeoyl pyruvate hydrolases). Antibodies of interest bind to antigenic polypeptides or proteins, or protein fragments, which are described above under “Proteins.” Antibodies of the present invention can be immobilized on a carrier and used in an immunoassay or affinity chromatography column to detect and/or separate polypeptides, proteins or protein fragments, or cells containing such polypeptides, proteins or protein fragments. Alternatively, such polypeptides, proteins or protein fragments can be immobilized so as to elicit antibodies capable of binding specifically to them.

[0267] Antibodies specific for the proteins of the present invention, either polyclonal or monoclonal, can be produced using standard methods. In general, the protein is first used to immunize a suitable mammal, preferably a mouse, rat, rabbit or goat. Rabbits and goats are the preferred targets for the production of polyclonal sera due to the significant volume of blood serum produced and the availability of labeled anti-rabbit and anti-goat antibodies. Immunization is typically accomplished by mixing or emulsifying the specific protein in saline, preferably with an adjuvant such as Freund's complete adjuvant, and then administering the resulting mixture or emulsion parenterally (usually by subcutaneous or intramuscular injection). Typically, 50-200 mcg per injection is sufficient.

[0268] In various embodiments of the invention, recombinant or natural proteins in native or denatured form are used for immunization. Protein fragments or synthetic polypeptides containing part of the amino acid sequence of the protein according to the invention can also be used for immunization.

[0269] Immunization is usually boosted after 2-6 weeks with one or more additional injections of protein in saline, preferably including Freund's incomplete adjuvant. Antibodies can also be produced alternatively by in vitro immunization using methods known in the art, which for purposes of the present invention are equivalent to in vivo immunization. Polyclonal antisera are obtained by collecting blood from immunized animals in a glass or plastic container, followed by incubation of the blood at 25 C for 1 hour and then incubation at 4 C for 2-18 hours. Serum is isolated by centrifugation (for example, at 1000 g for 10 minutes). In rabbits, 20-50 ml of blood can be obtained at a time.

[0270] Monoclonal antibodies are prepared using the standard Kohler-Milstein method (Kohler & Milstein, 1975, Nature, 256, 495-496) or modifications thereof. Typically, a mouse or rat is immunized as described above. However, unlike drawing blood from animals to obtain serum, this method involves removing the spleen (and optionally some large lymph nodes) and macerating the tissue to separate individual cells. If desired, spleen cells can be screened (after removing non-specifically adherent cells) by applying a cell suspension to a plate or a separate well coated with antigen protein. B lymphocytes expressing membrane-bound immunoglobulin specific for the test antigen are bound to the plate in such a way that they are not washed off from the remaining suspension. The resulting B lymphocytes or all macerated splenocytes are then fused with the myeloma cells to form hybridomas, which are then cultured in a selective medium (eg HAT medium containing hypoxanthine, aminopterin and thymidine). The resulting hybridomas are plated at a limiting dilution and tested for the production of antibodies that specifically bind to the antigen used for immunization (and that do not bind to foreign antigens). Selected monoclonal antibody (mAb)-secreting hybridomas are then cultured either in vitro (eg, in hollow fiber bundle fermenters or glass tissue culture flasks) or in vivo (in ascites fluid in mice).

[0271] Antibodies (both polyclonal and monoclonal) can be labeled using standard methods. Suitable labels include fluorophores, chromophores, radionuclides (particularly 32P and 125I), electron-dense reagents, enzymes and ligands for which specific binding partners are known. Enzymes are usually identified by their catalytic activity. For example, horseradish peroxidase is typically identified by its ability to convert 3,3',5,5'-tetramethylbenzidine (TMB) into a blue pigment, quantified by a spectrophotometer. The term “specific binding partner” refers to a protein that is capable of binding a ligand molecule with a high level of specificity, such as in the case of an antigen and its specific monoclonal antibody. Other examples of specific binding partners are biotin and avidin (or streptavidin), immunoglobulin-G and protein-A, as well as numerous pairs of receptors and their ligands known in the art. Other variations and possibilities will be apparent to those skilled in the art and are considered equivalent within the scope of the present invention.

[0272] Antigens, immunogens, polypeptides, proteins or protein fragments of the present invention cause the formation of specific binding partners - antibodies. Said antigens, immunogens, polypeptides, proteins or protein fragments of the present invention include the immunogenic compositions of the present invention. Such immunogenic compositions may further contain or include adjuvants, carriers or other compositions that stimulate or enhance or stabilize the antigens, polypeptides, proteins or protein fragments of the present invention. Such adjuvants and carriers will be apparent to those skilled in the art.

Applications, combinations and methods of use

[0273] The present invention provides the use of fungal luciferin biosynthetic proteins as enzymes that catalyze the reactions of (1) the synthesis of luciferin (namely 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one) having the structural formula

,

where R is aryl or heteroaryl, from pre-luciferin (namely 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one) having the structural formula

;

or the synthesis of pre-luciferin from 3-arylacrylic acid (a precursor of pre-luciferin) with the structural formula

,

where R is selected from the group aryl or heteroaryl; or the synthesis of 3-aryl acrylic acid from 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acid (oxyluciferin) with the structural formula .

[0274] Fungal luciferin biosynthetic proteins find use in many methods of implementing the present invention, non-limiting examples of which are provided below in this chapter.

[0275] Fungal luciferin biosynthetic proteins useful in the present invention can be obtained from a variety of natural sources or produced by recombinant technologies. For example, wild-type proteins can be isolated from bioluminescent fungi, for example from fungi belonging to the phylum Basidiomycota, predominantly the class Basidiomycetes, in particular the order Agaricales. For example, wild-type proteins can be isolated from bioluminescent fungi Neonothopanus nambi, Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos, etc. They can also be produced by expressing the recombinant nucleic acid coding sequence of the protein in an appropriate host or cell-free expression system.

[0276] In some embodiments, the proteins are used within host cells where they are expressed and carry out fungal luciferin cycling reactions. In other embodiments, isolated recombinant or natural proteins or extracts containing proteins of use are used.

[0277] Fungal luciferin biosynthetic proteins are active under physiological conditions.

[0278] Some embodiments provide the use of hispidin hydroxylase proteins in in vitro and in vivo systems to produce luciferin, which is oxidized by bioluminescent fungal luciferases, their homologues and light-emitting mutants. Thus, the present invention provides the use of the hispidin hydroxylases of the present invention to catalyze the conversion reaction of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one (pre-luciferin) having the structural formula

,

c 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one (fungal luciferin), having the structural formula

,

where R is aryl or heteroaryl.

[0279] A method for producing mushroom luciferin from pre-luciferin comprises combining at least one molecule of hispidin hydroxylase with at least one molecule of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, with at least one a molecule of NAD(P)H and at least one molecule of molecular oxygen (O2). The reaction is carried out under physiological conditions in in vitro and in vivo systems at temperatures from 20 to 42 degrees Celsius, including the reaction can be carried out in cells, tissues and host organisms expressing hispidin hydroxylase. In advantageous embodiments, said cells, tissues and organisms contain sufficient amounts of NAD(P)H and molecular oxygen to carry out the reaction. The reaction can use exogenously added 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, or endogenous 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one produced in cells, tissues or organisms.

[0280] In advantageous embodiments of the hispidin hydroxylase of the present invention, 3-hydroxyhispidin is synthesized from hispidin. In advantageous embodiments, they synthesize at least one functional analog of 3-hydroxyhispidine from the corresponding preluciferin shown in Table 2. In some embodiments, the hispidin hydroxylases of the present invention synthesize 6-(2-arylvinyl)-3,4-dihydroxy-2H- pyran-2-one from any corresponding 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one having the structural formula ,

where R is aryl or heteroaryl.

[0281] The resulting 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one is used to produce luminescence in in vitro and in vivo systems containing a functional luciferase that recognizes fungal luciferin as a substrate.

[0282] Applicable for the needs of the present invention as hispidin hydroxylases are proteins whose amino acid sequences are shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, as well as their mutants, homologues and derivatives. For example, functional hispidin hydroxylases 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28 can be used, or at least 60%, or at least 65% %, or at least 70%, or at least 80%, or at least 85%, or at least 90%, or at least 91%, or at least 92%, or by at least 93%, or by at least 94%, or by at least 95%, or by at least 96%, or by at least 97%, or by at least 98%, or by at least 99% over at least 350 amino acids. For example, they may be 60% identical, or at least 65% identical, or at least 70% identical, or at least 80% identical, or at least 85% identical, or at least 90% identical. or by at least 91%, or by at least 92%, or by at least 93%, or by at least 94%, or by at least 95%, or by at least 96%, or by at least 97%, or at least 98%, or at least 99% throughout the entire polypeptide chain of the protein.

[0283] In advantageous embodiments, those useful for the purposes of the present invention as hispidin hydroxylase are proteins whose amino acid sequences are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 29-33. Consensus regions within the amino acid sequences of hispidin hydroxylases are operatively linked through amino acid insertions with less conservation (Figure 1).

[0284] In some embodiments, the use of hispidin synthase proteins in in vitro and in vivo systems is provided for the production of fungal pre-luciferin from its precursor, that is, the use for catalyzing the conversion reaction of 3-arylacrylic acids with the structural formula

,

where R is selected from the group aryl, heteroaryl, 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl.

[0285] A method for producing pre-luciferin includes combining at least one molecule of hispidin synthase with at least one molecule of 3-arylacrylic acid with at least one molecule of coenzyme A, with at least one molecule of ATP and with at least two molecules of malonyl -CoA.

[0286] The reaction is carried out under physiological conditions at a temperature of from 20 to 42 degrees Celsius, including the reaction can be carried out in cells, tissues and host organisms expressing a functional hispidin synthase. In advantageous embodiments, said cells, tissues and organisms contain sufficient amounts of coenzyme A, malonyl-CoA and ATP to carry out the reaction.

[0287] The reaction may use exogenously added 3-arylacrylic acid or 3-arylacrylic acid produced in cells, tissues or organisms.

[0288] For example, the hispidin synthases of the present invention can be used to produce hispidin from caffeic acid. In advantageous embodiments, they synthesize a functional hispidin analogue (6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one) from the corresponding 3-arylacrylic acid shown in Table 2.

[0289] The resulting 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one is used for the production of fungal luciferin in the presence of the hispidin hydroxylase of the present invention. Hispidin and its functional analogues are also used in medicine, as they exhibit antioxidant and antitumor properties; According to some data, hispidin can prevent obesity [Be Tu et al., Drug Discov Ther. 2015 Jun; 9 (3): 197-204; Nguyen et al., Drug Discov Ther. 2014 Dec; 8 (6): 238-44; Yousfi et al., Phytother Res. 2009 Sep; 23 (9): 1237-42].

[0290] Applicable for the needs of the present invention as hispidin synthases are proteins whose amino acid sequences are shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45,47,49, 51, 53, 55, as well as their mutants, homologs and derivatives. For example, functional hispidin synthases can be used having an amino acid sequence that is identical to a sequence selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45,47,49, 51, 53, 55, at least by 40%, usually by at least 45%, usually by at least 50%, for example by at least 55%, or at least 60%, or at least 65%, or at least 70%, or at least 80%, or at least 85%, or at least 90%, or at least 91%, or at least 92%, or at least 93% , or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%.

[0291] In advantageous embodiments, hispidin synthases useful in the present invention are proteins whose amino acid sequences are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 56-63. Consensus regions within the amino acid sequences of hispidin synthases are operatively linked through amino acid insertions with less conservation (Fig. 2).

[0292] Some embodiments provide the use of caffeoylpyruvate hydrolase proteins in in vitro and in vivo systems to produce 3-arylacrylic acids with the structural formula

[0293] ,

where R is selected from the group aryl, heteroaryl, 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acid with the structural formula

where R is aryl or heteroaryl. The reaction is carried out under physiological conditions in in vitro and in vivo systems. The caffeyl pyruvate hydrolases of the present invention find use in autonomous bioluminescence systems, described in detail below.

[0294] Proteins whose amino acid sequences are shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75, as well as their mutants, homologues and derivatives, are applicable as caffeoylpyruvate hydrolases for the purposes of the present invention. For example, functional caffeoylpyruvate hydrolases may be used having an amino acid sequence that is at least 60% identical to the sequence selected from SEQ ID NOs: 65, 67, 69, 71, 73, 75, at least 65% identical. , at least 70%, at least 80%, at least 85%, at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99% .

[0295] In advantageous embodiments, useful caffeoylpyruvate hydrolases for the purposes of the present invention are proteins whose amino acid sequences are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 76-78. Consensus regions within the amino acid sequences of caffeoylpyruvate hydrolases are operatively linked through amino acid insertions with less conservation (Figure 3).

[0296] Combinations of proteins useful in the methods of the present invention are also provided. In preferred embodiments, the combinations include a functional hispidin hydroxylase and a functional hispidin synthase. The combination finds application in the preparation of 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one from 3-aryl acrylic acid with the structural formula

,

where R is aryl or heteroaryl. For example, the combination can be used to produce caffeic acid hydroxyhispidin. The reaction is carried out under physiological conditions in the presence of at least one molecule of hispidin hydroxylase, at least one molecule of hispidin synthase, at least one molecule of 3-arylacrylic acid, at least one molecule of coenzyme A, at least one molecule of ATP, at least two molecules of malonyl-CoA, at least one molecule of NAD(P)H and at least one molecule of molecular oxygen (O ₂ ).

[0297] In some embodiments, the combination also includes a luciferase capable of using 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one with the structural formula

,

where R is aryl or heteroaryl, as luciferin. Oxidation of the said 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one by such luciferase is accompanied by bioluminescence and the formation of oxyluciferin (6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acid) .

[0298] Any protein characterized by the above activity can be used as luciferase. For example, known luciferases from bioluminescent fungi can be used, including those described in application RU No. 2017102986/10(005203) dated January 30, 2017, as well as their homologues, mutants and chimeric proteins with luciferase activity.

[0299] In many embodiments of the present invention, useful luciferases have amino acid sequences that are at least 40% identical, such as at least 45% identical, or at least 50% identical, or at least at least 55% identical, or at least 60% identical, or at least 70% identical, or at least 75% identical, or at least 80% identical, or at least 85% identical amino acid sequence selected from the group SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. Often luciferases are characterized by amino acid sequences that have an amino acid sequence selected from the group SEQ ID NOs: 80 , 82, 84, 86, 88,90, 92, 94, 96, 98, at least 90% identity (for example, at least 91%, at least 92%, at least 93%, at least 94%, at least 95 %, no less than no less than 96%, no less than 97%, no less than 98%, no less than 99% identity or 100% identity).

[0300] Mutants may retain the biological properties of the wild-type luciferase from which they were derived, or may have biological properties that differ from those of the wild-type proteins. The term "biological property" of luciferases according to the present invention refers, without limitation, to the ability to oxidize various luciferins; biochemical properties such as in vivo and/or in vitro stability (eg half-life); speed of maturation; tendency to aggregation or oligomerization, as well as other similar properties. Mutations include changes to one or more amino acids, deletions or insertions of one or more amino acids; N-terminal truncations or extensions, C-terminal truncations or extensions, etc.

[0301] In some embodiments of the present invention, luciferases are used in isolated form, that is, they are essentially free from the presence of other proteins or other naturally occurring biological molecules, such as oligosaccharides, nucleic acids and fragments thereof, and the like, wherein the term “substantially free” as used herein means that less than 70%, typically less than 60%, and more often less than 50% of the composition containing the isolated protein is another naturally occurring biological molecule. In some embodiments, the proteins are present in substantially purified form, wherein the term "substantially purified form" means a purity of at least 95%, typically at least 97%, and more typically at least 99%.

[0302] In some embodiments, luciferases are used in extracts obtained from bioluminescent fungi or from host cells containing nucleic acids encoding recombinant luciferases.

[0303] In many embodiments, the luciferases are found in heterologous expression systems (in cells or organisms of the present invention) that contain nucleic acids encoding recombinant luciferases.

[0304] Methods for producing recombinant proteins, in particular luciferases, either in isolated form, or in extracts, or in heterologous expression systems are well known in the art and are described in the Nucleic Acids section. Methods for purifying proteins are described in the “Proteins” section.

[0305] In advantageous embodiments, luciferases remain active at temperatures below 50°C, more often at temperatures up to 45°C, that is, they remain active at temperatures of 20-42°C and can be used in heterologous expression systems in vitro and in vivo. As a rule, the described luciferases have pH stability in the range from 4 to 10, more often in the range from 6.5 to 9.5. The optimum pH stability of the claimed proteins lies in the range between 7.0 and 8.5, for example between 7.3-8.0. In advantageous embodiments, said luciferases are active under physiological conditions.

[0306] The combination of hispidin hydroxylase and luciferase, which oxidizes fungal luciferin to release light, is used in methods for detecting hispidin and its functional analogues in biological objects: cells, tissues, or organisms. The method involves contacting the biological object under study or an extract obtained from it with a combination of isolated hispidin hydroxylase and said luciferase in a suitable reaction buffer that creates physiological conditions and contains the necessary components for carrying out the reactions. One skilled in the art can formulate a variety of reaction buffers that satisfy this condition. A non-limiting example of a reaction buffer is 0.2 M sodium phosphate buffer (pH 7.0-8.0) with the addition of 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM), 1 mM NADPH.

[0307] The presence of hispidin or its functional analogue is determined by the appearance of detectable light - bioluminescence. Methods for detecting detectable light are described above in the section “Proteins” when describing functional screening methods.

[0308] The combination of hispidin hydroxylase, hispidin synthase and luciferase, which oxidizes fungal luciferin to release light, finds use in methods for detecting 3-aryl acrylic acid with the structural formula

,

where R is aryl or heteroaryl, in biological objects. The method involves contact of the biological object under study or an extract obtained from it with a combination of isolated hispidin hydroxylase, hispidin synthase and luciferase, which creates physiological conditions and contains the necessary components for the reactions. One skilled in the art can formulate a variety of reaction buffers that satisfy this condition. A non-limiting example of a reaction buffer is 0.2 M sodium phosphate buffer (pH 7.0-8.0) with the addition of 0.5 M Na ₂ SO ₄ , 0.1% dodecyl maltoside (DDM), 1 mM NADPH, 10 mM ATP, 1 mM CoA, 1 mM Malonyl- CoA.

[0309] The presence of 3-arylacrylic acid is determined by the appearance of detectable light - bioluminescence. Methods for detecting detectable light are described above in the section “Proteins” when describing functional screening methods.

[0310] In some embodiments, instead of the combination of hispidin hydroxylase and luciferase that oxidizes fungal luciferin to release light, the fusion protein described in the "Proteins" section above can be used. The fusion protein simultaneously exhibits hispidin hydroxylase activity and luciferase activity and can be used in any method instead of a combination of the two enzymes.

[0311] In some embodiments, instead of the hispidin synthase described above, a type III polyketide synthase is used, characterized by an amino acid sequence substantially similar to an amino acid sequence selected from the group of SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139. Applicable for the needs of the present invention are type III polyketide synthases having an amino acid sequence that is identical to the sequence selected from the group SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137 , 139 by at least 40%, usually by at least 45%, usually by at least 50%, for example, or at least 55%, or at least 60%, or at least 65% , or by at least 70%, or by at least 80%, by at least 85%, or by at least 90%, or by at least 91%, or by at least 92%, or by at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least at 99%.

[0312] Representatives of these polyketide synthases (PKS) have been identified in many plant organisms; their ability and/or the ability of their mutants to catalyze the synthesis of bisnoryangonin from coumaryl-CoA is known from the prior art [Lim et al., Molecules, 2016 Jun 22;21(6) ]. Applicants have demonstrated that these enzymes are also capable of catalyzing the synthesis of hispidin from caffeoyl-CoA in in vitro and in vivo systems:

[0313] Thus, the use of these proteins for the synthesis of hispidin is also within the scope of the present invention.

[0314] In some embodiments of the present invention, PKS are used in isolated form, that is, they are essentially free from the presence of other proteins or other naturally occurring biological molecules, such as oligosaccharides, nucleic acids and fragments thereof, and the like, where the term “substantially free” here means that less than 70%, typically less than 60%, and more often less than 50% of the composition containing the isolated protein is another naturally occurring biological molecule. In some embodiments, the proteins are present in substantially purified form, wherein the term "substantially purified form" means a purity of at least 95%, typically at least 97%, and more typically at least 99%.

[0315] In many embodiments, PKS are found in heterologous expression systems (in cells or organisms of the present invention) that contain nucleic acids encoding recombinant enzymes.

[0316] Methods for producing recombinant proteins either in isolated form, or as part of extracts, or in heterologous expression systems are well known in the art and are described in the Nucleic Acids section. Methods for purifying proteins are described in the “Proteins” section.

[0317] In advantageous embodiments, PKS remain active at temperatures below 50°C, more often at temperatures up to 45°C, that is, they remain active at temperatures of 20-42°C and can be used in heterologous expression systems in vitro and in vivo. Typically, the described PKS have a pH stability in the range from 4 to 10, more often in the range from 6.0 to 9.0. The optimum pH stability of the claimed proteins lies in the range between 6.5 and 8.5, for example, between 7.0-7.5. In advantageous embodiments, said PKSs are active under physiological conditions.

[0318] A method for producing hispidin involves combining at least one molecule of the type III polyketide synthase described above with at least two molecules of malonyl-CoA and at least one molecule of caffeoyl-CoA.

[0319] In some embodiments, the method includes producing caffeyl-CoA from caffeic acid through an enzymatic reaction catalyzed by coumarate-CoA ligase. In this case, the method includes combining at least one molecule of the type III polyketide synthase described above with at least one molecule of caffeic acid, with at least one molecule of coenzyme A, with at least one molecule of coumarate-CoA ligase, with at least one molecule of ATP and at least two molecules of malonyl-CoA.

[0320] For the needs of the present invention, any coumarate-CoA ligase enzymes known from the prior art that carry out the reaction of adding coenzyme A to caffeic acid to form caffeoyl-CoA can be used:

[0321] In particular, coumarate CoA ligase 1 from Arabidopsis thaliana having the amino acid and nucleotide sequences shown in SEQ ID NO: 141, as well as functional mutants and homologues thereof, can be used. For example, suitable for the purposes of the present invention is a functional coumarate-CoA ligase, the amino acid sequence of which has at least 40% identity, for example, at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85% identity, at least 90% identity (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identity) with the amino acid sequence shown in SEQ ID NO: 141.

[0322] All of these reactions are carried out under physiological conditions at temperatures from 20 to 50 degrees Celsius, including the reaction can be carried out in cells, tissues and host organisms expressing functional enzymes.

[0323] In combination with the hispidin hydroxylase of the present invention, PKS and coumarate-CoA ligase can be used to produce 3-hydroxyhispidin from caffeic acid. The reaction is carried out under physiological conditions in the presence of at least one molecule of hispidin hydroxylase, at least one molecule of PKS, at least one molecule of coumarate-CoA ligase, at least one molecule of caffeic acid or caffeyl-CoA, at least one a molecule of coenzyme A, with at least one molecule of ATP, with at least one molecule of NAD(P)H, with at least one molecule of oxygen and at least two molecules of malonyl-CoA.

[0324] The present invention also provides the use of nucleic acids encoding enzymes for the biosynthesis of fungal luciferin, homologues and mutants of these proteins, including shortened and elongated forms and fusion proteins, for the production of enzymes involved in the biosynthesis of fungal luciferin in in vitro and\ or in vivo.

[0325] Advantageous embodiments provide the use of nucleic acids encoding the hispidin hydroxylases of the invention, namely proteins characterized by the amino acid sequences shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20 , 22, 24, 26, 28, as well as their functional homologues, mutants and derivatives. In preferred embodiments, the nucleic acids encode proteins that are at least 40%, more typically at least 45%, typically at least 50%, e.g., at least 55%, at least 60%, amino acid in sequence. at least 65%, at least 70%, at least 80%, at least 85%, at least 90%, or at least 91%, or at least 92% , or at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99% identical to the sequences shown in SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, over at least 350 amino acids. In preferred embodiments, the nucleic acids encode proteins whose amino acid sequences are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 29-33.

[0326] Also provided is the use of nucleic acids encoding hispidin synthases, namely proteins characterized by the amino acid sequences shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, as well as their functional homologues, mutants and derivatives. In preferred embodiments, the nucleic acids encode proteins whose amino acid sequences are at least 40%, more typically at least 45%, typically at least 50%, such as at least 55%, or at least 60 %, or at least 65%, or at least 70%, or at least 80%, or at least 85%, or at least 90%, or at least 91%, or by at least 92%, or by at least 93%, or by at least 94%, or by at least 95%, or by at least 96%, or by at least 97%, or by at least 98%, or at least 99% identical to the sequences shown in SEQ ID NOs: 35, 37, 39, 41, 43, 45,47,49, 51, 53, 55, throughout the entire polypeptide chain of the protein . In preferred embodiments, the nucleic acids encode proteins whose amino acid sequences are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 56-63.

[0327] Also provided is the use of nucleic acids encoding caffeoylpyruvate hydrolases, namely proteins characterized by the amino acid sequences shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75, as well as functional homologues, mutants and derivatives thereof. In preferred embodiments, the nucleic acids encode proteins that are at least 40%, more typically at least 45%, typically at least 50%, e.g., at least 55%, or at least 60%, amino acid in sequence. or by at least 65%, or by at least 70%, or by at least 80%, or by at least 85%, or by at least 90%, or by at least 91%, or by at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99% identical to the sequences shown in SEQ ID NOs: 65, 67, 69, 71, 73, 75, throughout the entire polypeptide chain. In preferred embodiments, the nucleic acids encode proteins whose amino acid sequences are characterized by the presence of several conserved amino acid motifs (consensus sequences) shown in SEQ ID NOs: 76-78.

[0328] The above groups of nucleic acids are used for the production of recombinant proteins of hispidin hydroxylases, hispidin synthases and caffeoylpyruvate hydrolases, as well as for the expression of these proteins in heterologous expression systems.

[0329] In particular, nucleic acids encoding hispidin hydroxylases are used to produce cells producing 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula

[0330] from exogenous or endogenous 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one having the structural formula

,

where R is aryl or heteroaryl.

[0331] Nucleic acids encoding caffeoylpyruvate hydrolases are used to obtain cells and organisms capable of converting oxyluciferin into a precursor of preluciferins.

[0332] Nucleic acids encoding hispidin synthases are used to produce cells producing the above 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one from the corresponding 3-arylacrylic acid. For example, cells expressing hispidin synthase are used to produce hispidin from caffeic acid.

[0333] In some embodiments, nucleic acids encoding hispidin synthases are used to produce hispidin from tyrosine. In these embodiments, nucleic acids encoding enzymes that provide the synthesis of caffeic acid from tyrosine are additionally introduced into the cells. Such enzymes are known from the prior art. For example, a combination of nucleic acids encoding Rhodobacter capsulatus tyrosine ammonium lyase and E. coli 4-hydroxyphenylacetate 3-monooxygenase reductase components HpaB and HpaC can be used as described in [Lin and Yan. Microbial Cell Fact. 2012, 4;11:42]. It will be apparent to one skilled in the art that alternatively, any other enzymes known in the art that carry out the reaction of converting tyrosine into caffeic acid can be used, for example, enzymes whose amino acid sequences are substantially similar to the amino acid sequences of the Rhodobacter capsulatus tyrosine ammonium lyase and the components of HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase from E. coli, shown in SEQ ID NOs: 107, 109 and 111. For example, these enzymes may have amino acid sequences that have at least 40% identity, for example, at least 45% identity, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, or at least 80% identity , or or at least 85% identical, or at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity) with the sequences shown in SEQ ID NOs: 107, 109 and 111, respectively.

[0334] In some embodiments, nucleic acids encoding hispidin synthases are used to produce cells producing functional hispidin analogues from aromatic compounds, including aromatic amino acids and derivatives thereof. In these embodiments, nucleic acids are additionally introduced into the cells, encoding enzymes that provide the synthesis of 3-arylacrylic acids, from which, in turn, the biosynthesis of functional hispidin analogues occurs. Such enzymes are known from the prior art. For example, for the biosynthesis of cinnamic acid, a nucleic acid encoding the phenylalanine ammonium lyase of Streptomyces maritimus can be used, as described in [Bang, H.B., Lee, Y.H., Kim, S.C. et al. Microb Cell Fact (2016) 15: 16. https://doi.org/10.1186/s12934-016-0415-9]. It will be apparent to one skilled in the art that alternatively, any other enzymes known in the art that carry out reactions that convert aromatic amino acids and other aromatic compounds into 3-aryl acrylic acids can be used. For example, any functional phenylalanine ammonium lyase can be used to synthesize cinnamic acid, for example, a phenylalanine ammonium lyase, the amino acid sequence of which is substantially similar to the sequence shown in SEQ ID NO: 117, for example, the sequence of which is identical to the sequence of SEQ ID NO: 117 at least 40%, including having at least 45% identity with it, or at least 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity , or at least 70% identical, or at least 75% identical, for example, at least 80% identical, at least 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93 %, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity).

[0335] In some embodiments, obtaining host cells expressing a functional hispidin synthase requires co-transfecting them with a nucleic acid encoding a hispidin synthase of the invention and a nucleic acid encoding a 4'-phosphopantotheinyl transferase capable of transferring 4-phosphopantetheinyl from coenzyme A to serine in the acyl-transfer domain of polyketide synthases. In other embodiments, the selected host cells, for example, plant cells or cells of certain lower fungi (eg, those belonging to the genus Aspergillus) contain endogenous 4'-phosphopantotheinyl transferase and co-transfection is not required.

[0336] The use of nucleic acid combinations of the invention is also provided. Thus, a combination of nucleic acids encoding hispidin hydroxylase and hispidin synthase is used to obtain cells producing 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one from 3-arylacrylic acid, for example, to obtain 3-hydroxyhispidine from caffeic acid and/or tyrosine. In some embodiments, the combination of nucleic acids includes a nucleic acid encoding a 4'-phosphopantotheinyl transferase. In some embodiments, the combination of nucleic acids includes nucleic acids encoding enzymes for the synthesis of 3-arylacrylic acid from cellular metabolites, for example, enzymes for the synthesis of caffeic acid from tyrosine or the synthesis of cinnamic acid from phenylalanine.

[0337] In some embodiments, a combination of nucleic acids encoding PKS and coumarate CoA ligases is used to produce hispidin producing cells from caffeic acid. Suitable for the purposes of the present invention are a nucleic acid encoding a functional PKS, the amino acid sequence of which is essentially similar or identical to a sequence selected from the group of SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139; for example, PKS, the amino acid sequence of which is identical to the sequence selected from the group SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, by at least 40%, more often by at least by 45%, usually by at least 50%, for example by at least 55%, or by at least 60%, or by at least 65%, or by at least 70%, or by at least 80%, or at least 85%, or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94% , or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%. Also suitable for the purposes of the present invention is a nucleic acid encoding a functional coumarate-CoA ligase that catalyzes the reaction of coenzyme A adding to caffeic acid to form caffeoyl-CoA. For example, a nucleic acid encoding a functional coumarate-CoA ligase may be used, the amino acid sequence of which is identical to or has at least 40% identity with the sequence shown in SEQ ID NO: 141, for example, at least 45% identity, or not less than 50% identity, or at least 55% identity, or at least 60% identity, or at least 65% identity, or at least 70% identity, or at least 75% identity, for example, at least 80% identity, at least 85% identical, at least 90% identical (for example, at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98%, or 99% identical).

[0338] In some embodiments, a combination of a nucleic acid encoding a hispidin hydroxylase of the invention and a nucleic acid encoding a PKS is used. In preferred embodiments, the combination also includes a nucleic acid encoding a coumarate CoA ligase. The combination is used to obtain 3-hydroxyhispidin-producing cells from caffeic acid and/or caffeyl-CoA.

[0339] In some embodiments, the combination of nucleic acids includes nucleic acids encoding enzymes for the synthesis of caffeic acid from tyrosine.

[0340] Of particular interest are the nucleic acid combinations of the invention when used together with a nucleic acid encoding a luciferase capable of oxidizing fungal luciferins to release light. Nucleic acid molecules encoding luciferases for the purposes of the present invention can be cloned from biological sources, for example, from a fungus belonging to the phylum Basidiomycota, preferably the class Basidiomycetes, in particular the order Agaricales, or obtained using genetic engineering methods. Luciferase mutants having luciferase activity can be generated using standard molecular biology techniques, such as those detailed above in the Nucleic Acids section. Mutations include changes in one or more amino acids, deletions or insertions of one or more amino acids; N-terminal truncations or extensions, C-terminal truncations or extensions, etc. In preferred embodiments, these nucleic acids encode luciferases whose amino acid sequences are at least 40% identical, for example, at least 45% identical, or at least 50% identical, or at least 55% identical, or at least 60% identical, or at least 70% identical, or at least 75% identical, or at least 80% identical identical, or at least 85% identical, to an amino acid sequence selected from the group of SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. For example, they may have amino acid sequences that have at least 90% identity (for example, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least at least 96%, at least 97%, at least 98%, at least 99% identity or 100% identity) with an amino acid sequence selected from the group of SEQ ID NOs: 80, 82, 84, 86, 88,90, 92, 94, 96, 98. Non-limiting examples of nucleic acids encoding luciferases are shown in SEQ ID NOs: 79, 81, 83, 85, 87, 89, 91, 93 and 95.

[0341] In some embodiments, a combination of a nucleic acid encoding a hispidin hydroxylase of the invention and a nucleic acid encoding a luciferase described above is used. The combination finds use in a wide range of applications when labeling organisms, tissues, cells, cellular organelles or proteins using bioluminescence. Methods for labeling organisms, tissues, cells, cellular organelles, or proteins with luciferase are well known in the art and involve introducing into a host cell a nucleic acid encoding luciferase, for example, as part of an expression cassette that allows expression of luciferase in said cell, tissue, or organism. . When a suitable luciferin is added to cells, tissue or organism expressing luciferase, detectable light is generated. When labeling cellular organelles or proteins, a nucleic acid encoding luciferase is operatively linked to a nucleic acid encoding, respectively, a localization signal to the cellular organelle of interest or the protein of interest. When co-expressing luciferase and hispidin synthase of the present invention in cells, biological objects (cells, tissues, organisms, cellular organelles or proteins) acquire the ability to glow in the presence of not only fungal luciferin, but also pre-luciferin (the latter in most cases is more stable in the presence of air oxygen).

[0342] Also, the combination of nucleic acids finds use in studying the dependence of the activity of two promoters in heterologous expression systems. In this case, a nucleic acid encoding luciferase, operatively linked to promoter “A,” and a nucleic acid encoding hispidin hydroxylase, operatively linked to promoter “B,” are introduced into the host cell. By adding luciferin, or pre-luciferin, or a mixture of pre-luciferin with luciferase to aliquots of cells (or cell extracts), the activity of one promoter “A” (luminescence is detected only in the presence of luciferin), one promoter “B” (luminescence is detected in the presence of a mixture of preluciferin and luciferase), or both promoters (luminescence is detected in all cases).

[0343] In some embodiments, the combination also contains a nucleic acid encoding hispidin synthase. In some embodiments, the combination further comprises a nucleic acid encoding a 4'-phosphopantotheinyl transferase.

[0344] In some embodiments, the combination comprises a nucleic acid encoding hispidin synthase, a nucleic acid encoding luciferase, a nucleic acid encoding PKS, and a nucleic acid encoding coumarate CoA ligase.

[0345] The combinations find use in a wide range of applications when labeling organisms, tissues, cells, cellular organelles or proteins using bioluminescence. In this embodiment, a suitable pre-luciferin precursor, such as caffeic acid or coumaric acid, is added to biological entities expressing hispidin hydroxylase, luciferase and hispidin synthase or hispidin hydroxylase, luciferase, PKS and coumarate-CoA ligase to produce luminescence.

[0346] The combinations also find use in methods for studying the activity dependence of three promoters in heterologous expression systems. The methods involve introducing to the host cell a nucleic acid encoding luciferase under the control of promoter “A”, a nucleic acid encoding hispidin hydroxylase under the control of promoter “B” and a nucleic acid encoding hispidin synthase (or PKS) under the control of promoter “ IN". If co-expression of 4'-phosphopantotheinyl transferase is necessary for the maturation of a functional hispidin synthase, it is also introduced into the cell under the control of any suitable constitutive or inducible promoter. Simultaneous activation of all three promoters will be indicated by the appearance of detectable light when a suitable pre-luciferin precursor is added to the cells (or their extracts).

[0347] Combinations also find use in the production of transgenic luminous organisms. In advantageous embodiments, the transgenic organisms are obtained from organisms whose wild type does not exhibit bioluminescence. Nucleic acids encoding the target proteins are introduced into the transgenic organism as part of an expression cassette or vector, which exists in the body as an extrachromosomal element or is integrated into the genome of the organism, as described above in the section “Transgenic Organisms,” and provides expression of the target proteins. The transgenic organisms of the invention are distinguished by the fact that they express, in addition to luciferase, the substrate of which is fungal luciferin, at least hispidin hydroxylase. In advantageous embodiments they also express hispidin synthase. In other advantageous embodiments they also express PKS. In other advantageous embodiments they also express PKS. In some embodiments, they also express coumarate CoA ligase. It is known that endogenous coumarate-CoA ligase is present in many plant organisms, so its additional introduction is carried out in cases where endogenous coumarate-CoA ligase is absent.

[0348] In some embodiments, they also express caffeoyl pyruvate hydrolase. Unlike organisms expressing only luciferase, transgenic organisms obtained using the nucleic acids of the present invention acquire the ability to glow in the presence of pre-luciferins and/or pre-luciferins - 3-arylacrylic acids (most often caffeic acid) - which are the cheapest and most stable a substrate for producing bioluminescence, which can be added to water for watering plants, or to a medium for cultivating microorganisms, or to the feed or habitat of animals (for example, fish). Bioluminescent transgenic organisms (plants, or animals, or fungi) are used as light sources and are also used for decorative purposes. Bioluminescent transgenic organisms, cells and cell cultures can also be used in various screens in which the intensity of bioluminescence changes depending on external influence. For example, they can be used to analyze the influence of various factors on the activity of promoters that regulate the expression of exogenous nucleic acids.

[0349] Of particular interest are autonomously bioluminescent transgenic organisms, which are also provided by the present invention.

[0350] In some embodiments, at least one 3-arylacrylic acid with the structural formula is present as a metabolite in these organisms

,

where R is aryl or heteroaryl.

[0351] Non-limiting examples of such organisms include higher and lower plants, including flowering plants and mosses. To obtain autonomously luminous transgenic plants, nucleic acids capable of expressing the corresponding enzymes are introduced into them, encoding hispidin synthase, hispidin hydroxylase and luciferase, capable of oxidizing fungal luciferin to release light. Since plants, as a rule, contain endogenous 4'-phosphopantotheinyl transferase, additional introduction of the nucleic acid encoding this enzyme is usually not required to obtain autonomously luminous plants.

[0352] In some embodiments, organisms that do not naturally produce 3-arylacrylic acids are used to produce autonomously bioluminescent transgenic organisms. Examples of such organisms include animals and many microorganisms, such as yeast and bacteria. To obtain autonomous bioluminescence, in this case, nucleic acids capable of expression are additionally introduced into organisms, encoding enzymes that provide the biosynthesis of 3-arylacrylic acids from cell metabolites, for example, caffeic acid from tyrosine. If necessary, a nucleic acid encoding 4'-phosphopantotheinyl transferase is also introduced into the organisms.

[0353] In some embodiments, to obtain autonomously luminous organisms, nucleic acids capable of expressing the corresponding enzymes are introduced into them, encoding PKS, hispidin hydroxylase, and luciferase capable of oxidizing fungal luciferin to release light. In advantageous embodiments, said cells, tissues and organisms contain sufficient amounts of caffeoyl-CoA and malonyl-CoA to carry out the hispidin synthesis reaction.

[0354] In cases where the transgenic organism does not produce sufficient amounts of caffeyl-CoA during normal metabolic processes, the nucleic acid encoding coumarate-CoA ligase, as well as, if necessary, enzymes for the biosynthesis of caffeic acid from tyrosine

[0355] In advantageous embodiments, the combination of nucleic acids to produce autonomously bioluminescent cells or transgenic organisms also includes a nucleic acid encoding caffeoyl pyruvate hydrolase. As demonstrated in the experimental section below, expression of caffeoylpyruvate hydrolase results in increased bioluminescence intensity of autonomously luminescent bioluminescent cells or transgenic organisms. In advantageous embodiments, the bioluminescence intensity is increased by at least 1.5 times, typically by at least 2 times, typically by at least five times, for example 7-9 times, for example 8 times or more.

[0356] Autonomously bioluminescent transgenic organisms (plants, or animals, or fungi), as well as cells and cell cultures, differ from transgenic organisms, cells and cell cultures expressing only luciferase and known in the art in that they do not require exogenous addition of luciferin or its precursor to them.

[0357] In some embodiments, instead of a combination of nucleic acids encoding hispidin hydroxylase and luciferase, a nucleic acid encoding a fusion protein of these two enzymes is used. It will be apparent to one skilled in the art that said fusion protein and the combination of nucleic acids encoding hispidin hydroxylase and luciferase are interchangeable entities in all modes of use. It is also apparent that other fusion proteins can be made from the nucleic acids of the invention which will retain the properties of the fusion partners; such fusion proteins and the nucleic acids encoding them can be used without limitation in place of combinations of individual proteins and nucleic acids.

[0358] In all of the applications and methods described above, the nucleic acids may be in the form of expression cassettes that can be used to cause expression of a coding sequence in a host cell. The nucleic acid may be introduced into a host cell as part of a vector to effect expression in a suitable host cell, or not included in the vector, for example, it may be incorporated into a liposome or viral particle. Alternatively, the purified nucleic acid molecule can be incorporated directly into the host cell by suitable means, for example, by direct endocytic uptake. The genetic construct can be introduced directly into the cells of a host organism (eg, a plant) by transfection, infection, microinjection, cell fusion, protoplast fusion, microprojectile bombardment, or using a “gene gun” (a gun that fires microparticles carrying the genetic construct).

[0359] The use of polyclonal and monoclonal antibodies according to the invention is also provided. They find use in methods for staining preparations of tissues, cells or organisms to reveal the localization of expressed or naturally occurring hispidin hydrolases, hispidin synthases and caffeoylpyruvate hydrolases according to the invention. Methods of staining using specific antibodies are well known in the art and are described, for example, in [Bykov V. L. Cytology and general histology]. The direct immunohistochemical method is based on the reaction of specific binding of labeled antibodies directly to the detected substance; the indirect immunohistochemical method is based on the fact that unlabeled primary antibodies bind to the detected substance, and then they are detected using secondary labeled antibodies, while the primary antibodies serve as antigens for the secondary antibodies. Antibodies are also used to stop enzymatic reactions. Contact of an antibody with a specific binding partner results in inhibition of the enzymatic reaction. Antibodies also find use in methods for purifying recombinant and natural proteins according to the invention using affinity chromatography. Affinity chromatography methods are known in the art and are described, for example, in [Ninfa et al (2009). Fundamental Laboratory Approaches for Biochemistry and Biotechnology (2 ed.). Wiley. p.133.; Cuatrecasas (1970). J.B.C. Retrieved November 22, 2017].

Sets and products

[0360] A further embodiment of the invention is an article that includes the hispidin hydroxylase or hispidin synthase or caffeoyl pyruvate hydrolase described above, or a nucleic acid encoding the above enzyme, preferably with elements to ensure expression of the target protein in a host cell, for example , a vector or expression cassette containing a nucleic acid encoding a target protein. Alternatively, the nucleic acid may contain flanking sequences for insertion into the target vector. Nucleic acids can be incorporated into promoterless vectors for convenient cloning of target regulatory elements. Recombinant proteins can be in a lyophilized state or dissolved in a buffer solution. Nucleic acids can be in a lyophilized state or as a precipitate in an alcohol solution or dissolved in water or a buffer solution.

[0361] In some embodiments, the article includes cells expressing one or more of the above nucleic acids.

[0362] In some embodiments, the article includes a transgenic organism expressing one or more of the above nucleic acids.

[0363] In some embodiments, the article includes antibodies for staining and/or inhibition and/or affinity chromatography of the above enzymes.

[0364] The product is a container with a label and instructions attached to it. Suitable containers include, for example, vials, ampoules, tubes, syringes, cell plates, petri dishes, etc. The container can be made of various materials, such as glass or polymer materials. The selection of a suitable container will be obvious to one skilled in the art.

[0365] In addition, the product may include other products required from a commercial and consumer point of view, for example, a reaction buffer, or components for its preparation, a buffer for reconstitution and/or dissolution and/or storage of proteins or nucleic acids, or components for its manufacture, deionized water, secondary antibodies to specific antibodies according to the invention, cell culture medium or components for its manufacture, food for the transgenic organism.

[0366] The products also include instructions for implementing the proposed methods. Instructions may be present in different forms, and one or more such forms may be attached to the product, for example, the instructions may be presented in the form of a file on electronic media and/or presented on printed media.

[0367] The invention also relates to kits that can be used for various purposes. The kit may include a combination of proteins of the invention or a combination of nucleic acids of the invention, preferably with elements for promoting expression of the target proteins in a host cell, for example, a vector or expression cassette containing a nucleic acid encoding the target protein. In some embodiments, the kit may also contain a nucleic acid encoding a luciferase capable of oxidizing fungal luciferin to release light. In some embodiments, the kit may contain nucleic acids encoding enzymes involved in the biosynthesis of caffeic acid from tyrosine. In some embodiments, the kit may also contain a nucleic acid encoding a 4'-phosphopantotheinyl transferase. In some embodiments, the kit may also contain a nucleic acid encoding a PKS. In some embodiments, the kit may also contain a nucleic acid encoding a coumarate CoA ligase.

[0368] In some embodiments, the kit may also contain antibodies for purifying recombinant proteins or staining expressed proteins in host cells. In some embodiments, the kit may also contain primers complementary to regions of said nucleic acid to amplify the nucleic acid or fragment thereof. In some embodiments, the kit may also contain one or more fungal luciferins and/or pre-luciferins and/or pre-luciferins. These compounds can be in the form of a dry powder, in the form of a solution in an organic solvent, or in the form of a solution in water. In some embodiments, the kit may include cells containing one or more of the above nucleic acids. In some embodiments, the kit may include a transgenic organism of the invention, for example, a producer strain or a transgenic autonomously bioluminescent plant. All kit components are placed in suitable containers. Kits usually also include instructions for use.

[0369] For a better understanding of the invention, the following examples are provided. These examples are provided for illustrative purposes only and should not be construed as limiting the scope of the invention in any form.

[0370] All publications, patents, and patent applications referenced in this specification are incorporated herein by reference. Although the foregoing invention has been described in some detail by way of illustration and example in order to avoid ambiguity, those skilled in the art will appreciate from the teachings disclosed herein that certain changes and modifications may be made without departing from the spirit and scope of the appended embodiments. implementation of the invention.

Experimental part (Examples)

Example 1. Isolation of hispidin hydroxylase sequences

[0371] Total RNA from the mycelium of Neonothopanus nambi was isolated according to the method described in [Chomczynski and Sacchi, Anal. Biochem., 1987, 162, 156-159]. cDNA was amplified using SMART PCR cDNA Synthesis Kit (Clontech, USA) according to the manufacturer's protocol. The resulting cDNA was used to amplify the coding sequence of luciferase, the nucleotide and amino acid sequences of which are shown in SEQ ID NOs: 79, 80. The coding sequence was cloned into the pGAPZ vector (Invitrogen, USA) according to the manufacturer's protocol and transformed into competent cells of E. coli strain XL1 Blue. Bacteria were grown on Petri dishes in the presence of the antibiotic zeocin. After 16 hours, the colonies were washed off the plates, mixed vigorously, and plasmid DNA was isolated from them using a plasmid DNA isolation kit (Evrogen, Russia). The isolated plasmid DNA was linearized at the AvrII restriction site and used to transform Pichia pastoris GS115 cells. Electroporation was carried out using the lithium acetate-dithiothreitol method described in [Wu and Letchworth, Biotechniques, 2004, 36:152-4]. Electroporated cells were scattered onto Petri dishes containing RDB medium containing 1 M sorbitol, 2% (w/v) glucose, 1.34% (w/v) yeast base nitrogen agar (YNB), 0.005% (w/v) amino acid mixture , 0.00004% (w/v) biotin and 2% (w/v) agar. The resulting colonies were sprayed with a solution of 3-hydroxyhispidin, detecting the presence of luciferase in the cells by the appearance of light. The light emitted by the colonies was detected using IVIS Spectrum CT (PerkinElmer, USA). Colonies in which luminescence was detected in response to the addition of 3-hydroxyhispidin were selected for further work.

[0372] Next, the amplified total cDNA from Neonothopanus nambi was cloned into the pGAPZ vector and transformed into competent E. coli strain XL1 Blue cells. Bacteria were grown on Petri dishes in the presence of the antibiotic zeocin. After 16 hours, the colonies were washed off the plates, mixed vigorously, and plasmid DNA was isolated from them using a plasmid DNA isolation kit (Evrogen, Russia). The isolated plasmid DNA was linearized at the AvrII restriction site and used to transform yeast cells Pichia pastoris GS115, constitutively expressing Neonothopanus nambi luciferase. Transformation was carried out by electroporation as described above. Cells were plated onto Petri dishes containing RDB medium containing 1 M sorbitol, 2% (w/v) glucose, 1.34% (w/v) yeast base nitrogen agar (YNB), 0.005% (w/v) amino acid mixture, 0.00004 % (w/v) biotin and 2% (w/v) agar. The diversity of the final Neonothopanus nambi cDNA library in yeast was on the order of one million clones. The resulting colonies were sprayed with a hispidin solution, detecting the presence of hispidin hydroxylase in the cells by the appearance of light. The light emitted by the colonies was detected using IVIS Spectrum CT (Perkin Elmer, USA). Cells expressing only luciferase and wild-type yeast cells were used as negative controls. When screening the library, colonies in which luminescence was detected were selected and used for PCR as a template with standard plasmid primers. PCR products were sequenced using the Sanger method to determine the sequence of the expressed gene. The resulting nucleic acid sequence of hispidin hydroxylase is shown in SEQ ID NO: 1. The amino acid sequence encoded by it is shown in SEQ ID NO: 2.

[0373] In FIG. Figure 4 shows the fluorescence of Pichia pastoris cells expressing hispidin hydroxylase and luciferase, or luciferase alone, or wild-type yeast, when colonies were sprayed with 3-hydroxyhispidin (luciferin) and hispidin (pre-luciferin). Data show that in the presence of hispidin hydroxylase, luciferin is formed in cells.

[0374] At the next stage, genomic DNA was isolated from the fungi Armillaria fuscipes, Armillaria gallica, Armillaria ostoyae, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Mycena chlorophos, Neonothopanus nambi, Neonothopanus gardneri, Omphalotus olearius and Panellus stipticus and full-genome sequencing was carried out using technology Illumina HiSeq (Illumina, USA) according to the manufacturer's recommendations. The sequencing results were used to predict the amino acid sequences of hypothetical proteins and used to search for homologs of hispidin hydroxylase from Neonothopanus nambi. Homologue searches were performed using software provided by the National Center for Biotechnology Information. Fungal genomic sequencing data were also searched for amino acid sequences in the NCBI Genbank database. The search used standard blastp search parameters. As a result, sequences of hispidin hydroxylase homologs from Neonothopanus nambi were identified in Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos.

[0375] The nucleotide and amino acid sequences of the Neonothopanus nambi hispidin synthase homologs are shown in SEQ ID NOs: 3-28.

[0376] All identified enzymes are essentially similar to each other. The degree of amino acid sequence identity is shown in Table 4.

Table 4. Percentage of identity of amino acid sequences of full-length natural proteins of hispidin hydroxylases. SEQ ID NO:2 SEQ ID NO:4 SEQ ID NO:6 SEQ ID NO:16 SEQ ID NO:8 SEQ ID NO:14 SEQ ID NO:20 SEQ ID NO:22 SEQ ID NO:24 SEQ ID NO:26 SEQ ID NO:28 SEQ ID NO:2 100 61 71 72 70 89 72 73 73 73 69 SEQ ID NO:4 61 100 50 48 48 63 49 51 50 50 45 SEQ ID NO:6 71 50 100 69 69 71 85 86 87 88 67 SEQ ID NO:6 72 48 69 100 76 72 71 72 72 72 71 SEQ ID NO:18 72 48 70 99 76 72 71 73 73 73 71 SEQ ID NO:8 70 48 69 76 100 71 70 70 71 71 73 SEQ ID NO:10 70 48 69 76 99 71 70 70 71 71 73 SEQ ID NO:14 89 63 71 72 71 100 72 75 73 74 71 SEQ ID NO:20 72 49 85 71 70 72 100 91 92 93 68 SEQ ID NO:22 73 51 86 72 70 75 91 100 93 93 69 SEQ ID NO:24 73 50 87 72 71 73 92 93 100 95 69 SEQ ID No:26 73 50 88 72 71 74 93 93 95 100 69 SEQ ID NO:28 69 45 67 71 73 71 68 69 69 69 100

[0377] Several highly homologous hispidin hydroxylase amino acid sequences, differing by single amino acid substitutions, have been isolated from Panellus stipticus and Mycena citricolor. Their nucleotide and amino acid sequences are shown in SEQ ID NOs 7-13 (Panellus stipticus) and SEQ ID NOs 15-18 (Mycena citricolor). Further study of the properties of these proteins did not reveal the effect of these substitutions on enzymatic properties.

[0378] The coding sequences of the identified homologues (SEQ ID NOs: 3-28) were cloned and transformed into Pichia pastoris GS115 cells constitutively expressing Neonothopanus nambi luciferase using the protocol described above. The resulting colonies were sprayed with a hispidin solution, detecting the presence of hispidin hydroxylase in the cells by the appearance of light. The light emitted by the colonies was detected using IVIS Spectrum CT (PerkinElmer, USA). All colonies expressing the tested genes (SEQ ID NOs: 3,5,7,9,11,13,15,17,19, 21, 23, 25,27) produced 1000-100000000 times more light when sprayed with hispidin solution than control cells, which confirms the ability of the enzymes encoded by the tested genes to catalyze the conversion of hispidin to 3-hydroxyhispidin (fungal luciferin).

[0379] Structural analysis of the amino acid sequences of the identified enzymes was performed. Analysis using SMART (Simple Modular Architecture Research Tool) software, available on the Internet at http://smart.embl-heidelberg.de [Schultz et al., PNAS 1998; 95:5857-5864; Letunic I, Doerks T, Bork P Nucleic Acids Res 2014; doi:10.1093/nar/gku949] revealed that all identified proteins include a FAD/NAD(P) binding domain (FAD/NAD(P) binding domain, IPR002938 - code according to the InterPro public database, available on the Internet at at http://www.ebi.ac.uk/interpro). This domain is involved in the binding of FAD and NAD in a number of enzymes, in particular monooxygenases, members of a large superfamily of enzymes that add a hydroxyl group to a substrate and are found in the metabolic pathways of many organisms. The identified hispidin hydroxylases, in addition to the FAD/NAD(P)-binding domain, contain N- and C-terminal amino acid sequences operatively associated with it (Fig. 1). Using multiple alignment and comparison of the amino acid sequences of the identified hispidin hydroxylases (Fig. 1), it was found that they contain several conserved amino acid motifs (consensus sequences) characteristic only for this group of enzymes (SEQ ID NOs: 29-33 ). Consensus regions within amino acid sequences are operatively linked through amino acid insertions.

Example 2: Expression of hispidin hydroxylase and fungal luciferase in mammalian cells and their combined use for cell labeling

[0380] The hispidin hydroxylase and luciferase coding sequences from Neonothopanus nambi, obtained as described in Example 1, were optimized for expression in mammalian cells (humanized). Optimized nucleic acids (SEQ ID NOs: 99 and 100) were obtained synthetically. The coding sequence of hispidin hydroxylase was cloned into the pmKate2-keratin vector (Evrogen, Russia) using NheI and NotI restriction sites instead of the sequence encoding the mKate2-keratin fusion protein. The luciferase sequence was amplified by PCR, digested with restriction endonucleases NheI and EcoRV (New England Biolabs, Ipswich, MA), and ligated into the lentiviral vector pRRLSIN.cPPT.EF1. Plasmid DNA was purified using plasmid DNA purification kits (Evrogen). Plasmid DNA containing the luciferase gene was used to generate stably expressing HEK293NT lines. Vector particles were produced by calcium phosphate transfection (Invitrogen, Carlsbad, CA) of HELK293T cells according to the protocol specified on the manufacturer's website. 24 hours before transfection, 1,500,000 cells were seeded into a 60 mm culture dish. For transfection, approximately 4 and 1.2 μg of packaging plasmids pR8.91 and pMD.G, as well as 5 μg of a transfer plasmid containing the luciferase sequence, were used. Viral particles were collected 24 hours after transfection, concentrated 10-fold, and used to transduce HEK293NT cells. About 100% of HEK293NT cells stably expressed Neonothopanus nambi luciferase.

[0381] The resulting cells were re-transfected with a vector containing the hispidin hydroxylase coding sequence using FuGENE HD transfection reagent (Promega, USA) according to the manufacturer's protocol. 24 hours after transfection, hispidin was added to the medium at a concentration of 800 μg/ml, and cell fluorescence was detected using IVIS Spectrum CT (PerkinElmer). The resulting cells emitted light with an intensity more than two orders of magnitude higher than the signal emanating from untransfected control cells (Fig. 5).

[0382] Cells were imaged under transmitted light in a green luminescence detection channel. Expression of hispidin hydroxylase from Neonothopanus nambi in human cells resulted in the appearance of a distinct light signal in the green region of the spectrum in the presence of hispidin, which made it possible to distinguish transfected cells from nontransfected ones.

Example 3: Use of hispidin hydroxylase with hispidin analogues in cell lysate

[0383] HEK293NT cells expressing Neonothopanus nambi luciferase and hispidin hydroxylase, obtained as described in Example 2, were washed from Petri dishes 24 hours after transfection using Versene solution supplemented with 0.025% trypsin, and the medium was changed to phosphate-buffered saline pH 8.0 by centrifugation, the cells were resuspended, lysed by ultrasound in a Bioruptor device (Diagenode, Belgium) for 7 minutes at a temperature of 0°C under the conditions recommended by the manufacturer, and 1 mM NADPH (Sigma-Aldrich, USA), as well as hispidin or one of its analogues:

(E)-4-hydroxy-6-(4-hydroxystyryl)-2H-pyran-2-one, (E)-6-(2-(1H-indol-3-yl)vinyl)-4-hydroxy-2H -pyran-2-one, (E)-6-(2-(1,2,3,5,6,7-hexahydropyrido[3,2,1-ij]quinolin-9-yl)vinyl)-4- hydroxy-2H-pyran-2-one, E)-6-(4-(diethylamino)styryl)-4-hydroxy-2H-pyran-2-one, or (E)-4-hydroxy-6-(2- (6-hydroxynaphthalene-2-yl)vinyl)-2H-pyran-2-one at a concentration of 660 μg/ml. Bioluminescence spectra were detected using a Varian Cary Eclipse spectrofluorimeter. Light emission in lysates was observed upon addition of all of the indicated functional hispidin analogues. Depending on the luciferin used, the expected shift in the luminescence maximum was observed.

Example 4. Preparation of recombinant hispidin hydroxylases

[0384] A polyhistidine sequence (His-Tag) was operatively appended to the 5' end of the nucleic acids encoding hispidin 3-hydroxylase and luciferase from Neonothopanus nambi, prepared as described in Examples 1 and 2. The resulting constructs were cloned into the pET-23 vector using BamHI and HindIII restriction endonucleases. The vector was used to transform Escherichia coli strain BL21-DE3 cells. Cells were seeded on Petri dishes with LB medium containing 1.5% agar, ampicillin 100 μg/ml, and incubated overnight at 37°C. Escherichia coli colonies were then transferred to 4 ml of liquid LB medium with the addition of ampicillin, incubated overnight at rocking at 37oC. 1 ml of overnight culture was transferred to 100 ml of Overnight Express Autoinduction medium (Novagen), to which ampicillin was previously added. The culture was grown at 37oC for 2.5 hours until an optical density of 0.6 OU was reached at 600 nm, and then grown at room temperature for 16 hours. Then the cells were pelleted by centrifugation at 4500 rpm for 20 minutes in an Eppendorf 5810R centrifuge and resuspended in 35 ml of buffer (50 mM Tris HCl pH 8.0, 150 mM NaCl). Cells were disrupted by ultrasound and centrifuged again. Metal affinity chromatography on TALON resin (Clontech, USA) was used to purify recombinant proteins. The presence of the expected recombinant product was confirmed by electrophoresis.

[0385] Aliquots of isolated recombinant hispidin hydroxylases were used to test functionality and stability. To determine the functionality, 15 μl of a solution of the isolated recombinant protein was added to a test tube containing 100 μl of buffer (0.2 M Na-phosphate buffer, 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM) pH 8.0), 0.5 μl of purified recombinant luciferase from Neonothopanus nambi, 1 mM NADPH and 0.2 μM hispidin. The test tube was placed in the luminometer. The activity of the isolated recombinant proteins resulted in the emission of light when combined with hispidin and its analogues described in example 3, in the presence of Neonothopanus nambi luciferase. In all cases, the intensity of emitted light using hispidin was greatest with the Neonothopanus nambi hispidin hydroxylase and lowest with the Armillaria mellea hispidin hydroxylase.

Example 5. Preparation of 3-hydroxyhispidine, (E)-3,4-dihydroxy-6-styryl-2H-pyran-2-one and (E)-3,4-dihydroxy-6-(4-hydroxystyryl)-2H- pyran-2-one using recombinant hispidin hydroxylase

[0386] Isolated recombinant hispidin hydroxylase from Neonothopanus nambi, prepared as described in Example 4, was added to reaction mixtures containing 1 mM NADPH and 0.2 μM hispidin, (E)-4-hydroxy-6-styryl-2H-pyran-2 -one or (E)-4-hydroxy-6-(4-hydroxystyryl)-2H-pyran-2-one in 100 μl of buffer (0.2 M Na-phosphate buffer, 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM) pH 8.0). After 30 minutes, the reaction mixture was analyzed by high performance liquid chromatography using synthetic luciferins as standards. Chromatography demonstrated the appearance of peaks corresponding to derivatives hydroxylated at the third position: 3-hydroxyhispidine, (E)-3,4-dihydroxy-6-styryl-2H-pyran-2-one and (E)-3,4-dihydroxy-6- (4-hydroxy-styryl)-2H-pyran-2-one.

Example 6 Detection of bioluminescence using hispidin hydroxylase-luciferase fusion protein

[0387] Humanized DNA sequences encoding Neonothopanus nambi hispidin hydroxylase and luciferase, prepared as described in Example 2, were operatively linked to each other via a flexible short peptide linker with the amino acid sequence GGSGGSGGS (SEQ ID NOs: 115). The nucleotide and amino acid sequences of the resulting chimeric protein are shown in SEQ ID NOs 101 and 102. The nucleic acid encoding the chimeric protein was cloned into the pEGFP-N1 vector (Clontech, USA) instead of the EGFP gene under the control of the cytomegalovirus promoter. The resulting construct was transfected into HEK293T cells. Also, as a control, similar vectors containing the hispidin hydroxylase and luciferase genes separately were cotransfected. Transfection was carried out using the FuGENE HD transfection agent (Promega, USA) according to the manufacturer's protocol. 24 hours after transfection, 1 million cells were resuspended in 0.5 ml PBS and luminescence was recorded without the addition of hispidin and with the addition of hispidin (10 μg per 1 million cells) using a luminometer. The addition of hispidin led to the appearance of bioluminescence in the cells in the green region of the spectrum (Fig. 6). The addition of 3-hydroxyhispidine also resulted in the appearance of a bioluminescent signal. Expression of hispidin hydroxylase fusion proteins with luciferase instead of luciferase alone allows the use of more stable luciferin precursors (hispidin, bisnoryangonin, and others) for bioluminescent cell labeling and does not require cotransfection of two nucleic acids into cells.

Example 7 Preparation of polyclonal antibodies

[0388] The coding sequences of the hispidin hydroxylases of Neonothopanus nambi (SEQ ID NO: 1) and Armillaria mellea (SEQ ID NO: 19) were synthetically obtained as linear double-stranded DNA and cloned into pQE-30 expression vectors (Qiagen, Germany) in this way that the recombinant proteins contained a histidine tag at the N-terminus. After expression in E. coli, the recombinant proteins were purified using TALON metal affinity resin (Clontech, USA) under denaturing conditions. Purified protein preparations emulsified in Freund's adjuvant were used for four immunizations of rabbits at monthly intervals. The blood of rabbits was collected on the tenth or eleventh day after immunization. The activity of the resulting polyclonal antisera was tested using ELISA and Western immunoblotting methods on a panel of purified recombinant hispidin hydroxylases obtained as described in example 4.

[0389] Antibodies obtained by immunizing a rabbit with a protein from Neonothopanus nambi demonstrated activity against denatured and undenatured Neonothopanus nambi hispidin hydroxylase and against denatured Neonothopanus gardneri hispidin hydroxylase.

Antibodies obtained by immunizing a rabbit with a protein from Armillaria mellea were active against denatured and undenatured hispidin hydroxylases from Armillaria mellea, Armillaria gallica, Armillaria ostoyae and Armillaria fuscipes.

Example 8. Production of transgenic plants expressing hispidin hydroxylase and luciferase from Neonothopanus nambi

[0390] The coding sequences for Neonothopanus nambi hispidin hydroxylase and luciferase have been optimized for expression in cells of the moss Physcomitrella patens. An expression cassette was then created in silico containing the rice aktI gene promoter, the 5' untranslated region of human cytomegalovirus, a hispidin hydroxylase coding sequence optimized for expression in plant cells (SEQ ID NO 103), a stop codon, a terminator sequence from the osc gene agrobacterium Agrobacterium tumefaciens, rice ubiquitin promoter, Neonothopanus nambi luciferase coding sequence optimized for expression in moss cells (SEQ ID NO 112), terminator from the nos gene of the agrobacterium Agrobacterium tumefaciens.

[0391] The resulting sequence was synthesized in such a way that all of the specified fragments were quickly stitched together and cloned using the Gibson assembly technique [Gibson et al., Nat Methods, 2009, 6:343-5] into the expression vector pLand# 1 (Institut Jean-Pierre Bourgin, France), between DNA fragments coinciding with the genomic DNA locus of the moss Physcomitrella patens between the sequences of highly expressed moss genes Pp3c16_6440V3.1 and Pp3c16_6460V3.1. The pLand#1 vector also contained a guide RNA (sgRNA) sequence for the Cas9 nuclease, complementary to a region of the same DNA locus.

[0392] The plasmid DNA preparation was cotransformed along with an expression vector containing the Cas9 nuclease sequence under the Arabidopsis thaliana ubiquitin promoter into protoplasts of the moss Physcomitrella patens according to the polyethylene glycol transformation protocol described in [Cove et al., Cold Spring Harb Protoc., 2009, 2 ]. The protoplasts were then incubated in BCD medium for two days in the dark with shaking at 50 rpm to regenerate the cell wall. The protoplasts were then transferred to Petri dishes containing agar and BCD medium and grown under 16-hour light for a week. Transformed moss colonies were screened with external genomic primers using PCR to determine the success of integration of the genetic construct into the genome, transferred to fresh Petri dishes and grown under the same lighting conditions for 30 days.

[0393] The resulting moss gametophytes were soaked in BCD medium containing hispidin at a concentration of 900 μg/ml and analyzed using the IVIS Spectrum In Vivo Imaging System (Perkin Elmer). All transgenic plants analyzed exhibited bioluminescence with an intensity at least two orders of magnitude higher than the signal intensity of control plants expressing only luciferase, incubated in the same solution with hispidin.

Example 9. Identification of hispidin synthases and caffeoylpyruvate hydrolases

[0394] Fungal luciferin precursors, such as hispidin, belong to a large group of polyketide-derived chemical compounds. Such compounds can theoretically be prepared from 3-arylacrylic acids in which the substituent at the third position is aromatic substituents, including aryl or heteroaryl. It is known from the prior art that the enzymes involved in the synthesis of polyketides and their derivatives in various organisms are multi-domain complexes belonging to the protein superfamily of polyketide synthases. At the same time, no polyketide synthase was known from the prior art capable of catalyzing the reaction of converting 3-arylacrylic acid into substituted 4-hydroxy-2H-pyran-2-one. To search for the target polyketide synthase, a cDNA library from Neonothopanus nambi was screened.

[0395] It is known that in order to obtain functional polyketide synthases in a yeast heterologous expression system, it is necessary to additionally introduce into the culture a gene expressing 4'-phosphopantotheinyl transferase, an enzyme that transfers 4-phosphopantheinyl from coenzyme A to serine in the acyl-transfer domain of polyketide synthase [Gao Menghao et al ., Microbial Cell Factories 2013, 12:77]. The 4'-phosphopantotheinyl transferase gene NpgA from Aspergillus nidulans (SEQ ID NOs 104, 105), known in the art, was obtained synthetically and cloned into the pGAPZ vector. The plasmid was linearized at the AvrII restriction site and used to transform the yeast line Pichia pastoris GS115, constitutively expressing Neonothopanus nambi luciferase and hispidin hydroxylase, obtained as described in Example 1. The diversity of the resulting Neonothopanus nambi cDNA library in yeast was about one million clones.

[0396] A cDNA library from Neonothopanus nambi expressed in the specified yeast line Pichia pastoris was prepared according to the protocol described in Example 1 and used to identify hispidin synthases and caffeoyl pyruvate hydrolases. Cells were scattered onto Petri dishes containing RDB medium containing 1 M sorbitol, 2% (w/v) glucose, 1.34% (w/v) yeast base nitrogen agar (YNB), 0.005% (w/v) amino acid mixture, 0.00004% (w/v) biotin and 2% (w/v) agar.

[0397] To identify the hispidin synthase gene, the resulting colonies were sprayed with a solution of caffeic acid (a potential precursor of hispidin), detecting the presence of hispidin synthase in the cells by the appearance of light. The light emitted by the colonies was detected using IVIS Spectrum CT (PerkinElmer, USA). Cells expressing only luciferase and hispidin hydroxylase and wild-type yeast cells were used as negative controls. When screening the library, colonies in which luminescence was detected were selected and used for PCR as a template with standard plasmid primers. PCR products were sequenced using the Sanger method to determine the sequence of the expressed gene. The resulting nucleic acid sequence of hispidin synthase is shown in SEQ ID NO: 34. The amino acid sequence encoded by it is shown in SEQ ID NO: 35.

[0398] The resulting line of yeast Pichia pastoris, containing the luciferase, hispidin hydroxylase and hispidin synthase genes integrated into the genome of Neonothopanus nambi, as well as the 4'-phosphopantotheinyl transferase gene NpgA from Aspergillus nidulans, was further used to identify the enzyme catalyzing the conversion of oxyluciferin (( 2E,5E)-6-(3,4-dihydroxyphenyl)-2-hydroxy-4-oxohexa-2,5-dienoic acid) to caffeic acid. The cell line was again transformed with the linearized plasmid library of Neonothopanus nambi genes obtained at the first stage of the work. Colonies were sprayed with a solution of caffeoylpyruvate, detecting the presence of the target enzyme in the cells by the appearance of light. The light emitted by the colonies was detected using IVIS Spectrum CT (PerkinElmer, USA). Cells expressing only luciferase and hispidin hydroxylase and wild-type yeast cells were used as negative controls. When screening the library, colonies in which luminescence was detected were selected and used for PCR as a template with standard plasmid primers. PCR products were sequenced using the Sanger method to determine the sequence of the expressed gene. The resulting nucleic acid sequence of the isolated enzyme is shown in SEQ ID NO: 64. The amino acid sequence encoded by it is shown in SEQ ID NO: 65. The identified enzyme was named caffeoylpyruvate hydrolase.

Example 10. Identification of homologs of Neonothopanus nambi hispidin synthase and Neonothopanus nambi caffeoylpyruvate hydrolase

[0399] To search for homologs of the hispidin synthase and caffeoyl pyruvate hydrolase of Neonothopanus nambi, we used whole genome sequencing data from bioluminescent fungi obtained as described in Example 1. The search for homologues was carried out using software provided by the National Center for Biotechnology Information. Fungal genomic sequencing data were also searched for amino acid sequences in the NCBI Genbank database. The search used standard blastp search parameters.

[0400] Sequences of hispidin synthase homologs from Neonothopanus nambi have been identified in Armillaria fuscipes, Armillaria mellea, Guyanagaster necrorhiza, Mycena citricolor, Neonothopanus gardneri, Omphalotus olearius, Panellus stipticus, Armillaria gallica, Armillaria ostoyae, Mycena chlorophos. Their nucleotide and amino acid sequences are shown in SEQ ID NOs 36-55. All identified enzymes were essentially similar to each other. The degree of amino acid sequence identity is shown in Table 5.

Table 5. Percentage of identity of amino acid sequences of full-length natural proteins of hispidin synthases SEQ ID NO:35 SEQ ID NO:53 SEQ ID NO:43 SEQ ID NO:45 SEQ ID NO:37 SEQ ID NO:41 SEQ ID NO:55 SEQ ID NO:47 SEQ ID NO:49 SEQ ID NO:51 SEQ ID NO:35 100 56 57 56 51 81 57 50 57 57 SEQ ID NO:53 56 100 80 52 45 55 88 46 88 88 SEQ ID NO:43 57 80 100 52 47 56 83 45 85 86 SEQ ID NO:45 56 52 52 100 53 54 52 54 53 53 SEQ ID NO:37 51 45 47 53 100 51 46 51 47 47 SEQ ID NO:41 81 55 56 54 51 100 55 50 56 56 SEQ ID NO:55 57 88 83 52 46 55 100 46 90 91 SEQ ID NO:47 50 46 45 54 51 50 46 100 46 46 SEQ ID NO:49 57 88 85 53 47 56 90 46 100 95 SEQ ID NO:51 57 88 86 53 47 56 91 46 95 100

[0401] Two highly homologous amino acid sequences of hispidin synthases, differing by a single amino acid substitution, have been isolated from Panellus stipticus. Their nucleotide and amino acid sequences are shown in SEQ ID NOs 36-39.

[0402] The identified enzymes were tested for their ability to convert caffeic acid to hispidin using the method described in Example 9.

[0403] Multiple alignment of the amino acid sequences of the identified proteins made it possible to identify several highly homologous fragments of the amino acid sequence characteristic of this group of enzymes. Consensus sequences for these fragments are shown in SEQ ID NOs: 70-77. These sequences are separated by extended amino acid sequences as shown in FIG. 2.

[0404] Neonothopanus nambi caffeoyl pyruvate hydrolase homolog sequences have been identified in Neonothopanus gardneri, Armillaria mellea, Armillaria fuscipes, Armillaria gallica, Armillaria ostoyae. The nucleotide and amino acid sequences of the identified homologs are shown in SEQ ID NOs: 66-75. The identified enzymes were tested for their ability to convert caffeoyl pyruvate to caffeic acid using the method described in Example 9.

[0405] All identified enzymes are substantially similar to each other and are 280-320 amino acids in length. The degree of amino acid sequence identity is shown in Table 6.

Table 6. Percentage of amino acid sequence identity of full-length natural proteins of caffeoylpyruvate hydrolases. SEQ ID NO:65 SEQ ID NO:73 SEQ ID NO:75 SEQ ID NO:67 SEQ ID NO:69 SEQ ID NO:71 SEQ ID NO:65 100 64 64 64 64 64 SEQ ID NO:73 64 100 92 62 96 95 SEQ ID NO:75 64 92 100 62 90 90 SEQ ID NO:67 64 62 62 100 60 61 SEQ ID NO:69 64 96 90 60 100 97 SEQ ID NO:71 64 95 90 61 97 100

[0406] Analysis using SMART (Simple Modular Architecture Research Tool) software, available on the Internet at http://smart.embl-heidelberg.de [Schultz et al., PNAS 1998; 95:5857-5864; Letunic I, Doerks T, Bork P Nucleic Acids Res 2014; doi:10.1093/nar/gku949] revealed that the identified enzymes include a fumarylacetase domain (EC 3.7.1.2) located closer to the C-terminus, about 200 amino acids long, but the conserved region begins at approximately amino acid 8 according to the amino acid numbering of caffeoylpyruvate hydrolase Neonotopanus nambi. Multiple alignment revealed consensus sequences (SEQ ID NOs 76-78) characteristic of this group of proteins, separated by amino acid inserts with lower identity. The positions of the consensus sequences are shown in (Fig. 3).

Example 11. Preparation of recombinant hispidin synthase and caffeoylpyruvate hydrolase and their use to obtain bioluminescence

[0407] At the 5' ends of the nucleic acids encoding hispidin synthase and caffeoylpyruvate hydrolase of Neonothopanus nambi, obtained as described in example 9, a sequence encoding polyhistidine (His-Tag) was operatively attached, and the resulting constructs were cloned into the pET vector -23 using restriction endonucleases NotI and SacI. Vectors were used to transform Escherichia coli strain BL21-DE3-codon+ cells, which was carried out by electroporation. Transformed cells were plated on Petri dishes with LB medium containing 1.5% agar, ampicillin 100 μg/ml, and incubated overnight at 37°C. Escherichia coli colonies were then transferred to 4 ml of liquid LB medium containing 100 μg/ml ampicillin, incubated overnight with shaking at 37°C. 1 ml of the overnight culture was transferred to 200 ml of Overnight Express Autoinduction medium (Novagen), to which ampicillin was previously added. The culture was incubated at 37°C for 3 hours until an optical density of 0.6 FU at 600 nm was reached, and then incubated at room temperature for 16 hours. Then the cells were pelleted by centrifugation at 4500 rpm for 20 minutes in an Eppendorf 5810R centrifuge, resuspended in 20 ml of buffer (50 mM Tris HCl pH 8.0, 150 mM NaCl), lysed by ultrasound in a Bioruptor device (Diagenode, Belgium) for 7 minutes at 0°C under conditions recommended by the manufacturer and centrifuged again. Protein from the lysate was obtained using affinity chromatography on Talon resin (Clontech, USA). The presence of the expected recombinant product was confirmed by electrophoresis by the presence of bands of the expected length.

[0408] Aliquots of the isolated recombinant proteins were used to test functionality.

[0409] To determine the functionality of hispidin synthase, 30 μl of a solution of the isolated recombinant protein was added to a test tube containing 100 μl of buffer (0.2 M Na-phosphate buffer, 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM) pH 8.0, all components - Sigma -Aldrich, USA), 0.5 μl of purified recombinant Neonothopanus nambi luciferase, obtained as described in Example 4, 1 mM NADPH (Sigma-Aldrich, USA), 15 μl of purified recombinant Neonothopanus nambi hispidin hydroxylase, obtained as described in Example 4, 10 mM ATP (ThermoFisher Scientific, USA), 1 mM CoA (Sigma-Aldrich, USA), 1 mM Malonyl-CoA (Sigma-Aldrich, USA). The tube was placed in a GloMax 20/20 luminometer (Promega, USA). The reaction mixtures exhibited bioluminescence when 20 μM caffeic acid (Sigma-Aldrich, USA) was added to the solution. The emitted light had an emission maximum in the region of 520-535 nm.

[0410] To determine the functionality of caffeoylpyruvate hydrolase, 10 μl of a solution of the isolated recombinant protein was added to a test tube containing 100 μl of buffer (0.2 M Na-phosphate buffer, 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM) pH 8.0), 0.5 μl of luciferase Neonothopanus nambi, 1 mM NADPH (Sigma-Aldrich, USA), 15 µl hispidin hydroxylase, 10 mM ATP (ThermoFisher Scientific, USA), 1 mM CoA (Sigma-Aldrich, USA), 1 mM Malonyl-CoA (Sigma-Aldrich , USA), 30 µl of purified recombinant hispidin synthase. The tube was placed in a GloMax 20/20 luminometer (Promega, USA). Bioluminescence of the reaction mixture was detected when 25 μM caffeoyl pyruvate was added to the solution, indicating the ability of the tested enzyme to decompose caffeoyl pyruvate to caffeic acid. The emitted light had an emission maximum in the region of 520-535 nm.

[0411] The resulting enzymes were used to produce luminescence (bioluminescence) in a reaction with luciferase and hispidin hydroxylase from Neonothopanus nambi, obtained as described in Example 4. 5 μl of a solution of each isolated recombinant protein was added to a test tube containing 100 μl of buffer (0.2 M Na -phosphate buffer, 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM) pH 8.0), 1 mM NADPH (Sigma-Aldrich, USA), 10 mM ATP (ThermoFisher Scientific, USA), 1 mM CoA (Sigma-Aldrich, USA ), 1 mM malonyl-CoA (Sigma-Aldrich, USA) and 0.2 μM one of the 3-arylacrylic acids: paracoumaric acid (Sigma-Aldrich, USA), cinnamic acid (Sigma-Aldrich, USA) or ferulic acid (Abcam, USA ). In another experiment, instead of substituted 3-arylacrylic acid, mushroom oxyluciferin analogues - (2E,5E)-2-hydroxy-6-(4-hydroxyphenyl)-4-oxohexa-2,5-diene, (2E,5E)- 2-hydroxy-4-oxo-6-phenylhexa-2,5-diene, or (2E,5E)-2-hydroxy-6-(4-hydroxy-3-methoxyphenyl)-4-oxohexa-2,5-diene acids - also at a concentration of 0.2 µM. The tubes were placed in a luminometer. The activity of the isolated recombinant proteins resulted in the emission of light in each of the described reactions.

Example 13. Preparation of hispidin from caffeic acid

[0412] An expression cassette containing a nucleic acid encoding the Neonothopanus nambi hispidin synthase (SEQ ID NOs 34, 35), under the control of the constitutive promoter J23100, and an expression cassette containing the 4'-phosphopantotheinyl transferase gene NpgA from Aspergillus nidulans (SEQ ID NOs 104, 105) under the control of the araBAD promoter, flanked by homology regions to the SS9 site, were obtained synthetically and cloned into a bacterial expression vector containing a zeocin resistance cassette. The resulting construct was used for transformation and integration into the genome of E. coli BW25113 using bacteriophage lambda protein-mediated recombination as described in Bassalo et al. [ACS Synth Biol. 2016 Jul 15;5(7):561-8], using selection for resistance to zeocin. Integration of the full-length construct was confirmed using PCR with primers specific to the SS9 homology regions, and then the correctness of the integrated construct was verified by sequencing the PCR product of genomic DNA using the Sanger method.

[0413] The resulting E. coli strain was used to produce hispidin. In the first step, bacteria were cultured in five 50-ml plastic tubes in LB medium for 10 hours with shaking at 200 rpm at 37°C. 250 ml of the resulting culture was added to 3.3 liters of fermentation medium in a Biostat B5 fermenter (Braun, Germany) so that the initial optical density of the culture at 600 nm was about 0.35. The fermentation medium contained 10 g/L peptone, 5 g/L caffeic acid, 5 g/L yeast extract, 10 g/L NaCl, 25 g/L glucose, 15 g/L (NH4)2SO4, 2 g/L KH ₂ PO ₄ , 2 g/l MgSO4 7×H ₂ O, 14.7× mg/l CaCl ₂ , 0.1 mg/l thiamine, 1.8 mg/l, and 0.1% solution of the following composition: EDTA 8 mg/l, CoCl ₂ 6×H ₂ O 2.5 mg/l, MnCl ₂ 4H ₂ O 15 mg/l, CuCl ₂ 2H ₂ O 1.5 mg/l, H ₃ BO3 3 mg/l, Na ₂ MoO4 2H ₂ O 2.5 mg/l l, Zn(CH ₃ COO)2 2H ₂ O 13 mg/l, iron(III) citrate 100 mg/l, thiamine hydrochloride 4.5 mg/l. Fermentation was carried out at 37×°C, with aeration 3×l/min and stirring at 200 rpm. After 25 hours of cultivation, arabinose was added to the culture to a final concentration of 0.1 × mM. The pH was controlled automatically by adding NH4OH, bringing the pH to 7.0. A solution containing glucose 500 g/l, caffeic acid 5 g/l, arabinose 2 g/l, tryptone 25 g/l, yeast extract 50 g/l, MgSO4 7H ₂ O 17.2 g/l, (NH4)SO4 7.5 g/L, ascorbic acid 18 g/L was added to the fermenter to maintain glucose levels whenever the pH increased to 7.1. After 56 hours of cultivation, the concentration of hispidin in the medium was 1.23×g/L. The fermenter medium, as well as hispidin purified from it using high-performance liquid chromatography, had activity in a bioluminescent reaction with hispidin hydroxylase and Neonothopanus nambi luciferase.

Example 13. Preparation of 3-hydroxyhispidine from caffeic acid

[0414] An expression cassette containing a nucleic acid encoding Neonothopanus nambi hispidin hydroxylase (SEQ ID NOs 1, 2) under the control of the J23100 promoter was produced synthetically and cloned into a bacterial expression vector containing a spectinomycin resistance gene. The resulting vector was transformed into E. coli cells expressing the Neonothopanus nambi hispidin synthase, the zeocin resistance gene and the NpgA gene, obtained as described in Example 12. The resulting bacteria were used to obtain 3-hydroxyhispidin by fermentation, according to the protocol described in Example 12. however, with the addition of spectinomycin at a concentration of 50 mg/ml to all culture media used. After 48 hours of cultivation, the concentration of 3-hydroxyhispidin in the medium was 2.3×g/L. The fermenter medium, as well as 3-hydroxyhispidin purified from it using high-performance liquid chromatography, was active in a bioluminescent reaction with Neonothopanus nambi luciferase.

Example 14. Preparation of hispidin from cell metabolites and tyrosine

[0415] To produce biosynthetic hispidin from tyrosine, an E. coli strain was obtained that efficiently produces tyrosine and caffeic acid. The E. coli strain was obtained as described in [Lin and Yan. Microbial Cell Fact. 2012 Apr 4;11:42]. The basis for creating the strain was the E. coli BW25113 line with an integrated mutant lacY permease gene (lacY A177C) at the attB site, ensuring uniform consumption of arabinose by bacterial cells. Expression cassettes containing the coding sequences of the Rhodobacter capsulatus tyrosine ammonium lyase genes (SEQ ID NOs: 106, 107), and the HpaB and HpaC components of the E. coli 4-hydroxyphenylacetate 3-monooxygenase reductase (SEQ ID NOs: 108-111), each under the control of the constitutive promoter J23100, were obtained synthetically and integrated into the genome of the E. coli strain, as described in Example 12. At the next stage, the plasmid obtained as described in Example 12 and containing the coding sequence of the Neonothopanus hispidin synthase was integrated into the E. coli genome nambi under the control of the constitutive J23100 promoter, the zeocin resistance cassette from the pGAP-Z vector, and the NpgA gene. Integration into the E. coli genome was carried out using recombination mediated by bacteriophage lambda proteins according to the method from [Bassalo et al., ACS Synth Biol. 2016; 5(7):561-568]. Integration of the full-length construct was confirmed using PCR with primers specific to the SS9 homology regions (5'-CGGAGCATTTTGCATG-3' and 5'-TGTAGGATCAAGCTCAG-3'), and then the correctness of the integrated construct was verified by sequencing the PCR product of genomic DNA using the Sanger method. The resulting bacterial strain was used to produce biosynthetic hispidin in a fermenter.

[0416] Cultivation of bacteria was carried out in a fermenter as described in Example 12, with the only difference being that caffeic acid was not added to the media for cultivating bacteria. Biosynthetic hispidin was isolated from the medium using high-performance liquid chromatography. The created strain was capable of producing 1.20 mg/L hispidin after 50 hours of fermentation. The purity of the resulting drug was 97.3%. The addition of tyrosine to the culture media at a concentration of 10 g/ml increased the yield of hispidin to 108.3 mg/ml.

Example 15. Creation of transgenic autonomously bioluminescent yeast Pichia pastoris

[0417] To create autonomously bioluminescent yeast Pichia pastoris, expression cassettes were synthesized containing, under the control of the GAP promoter and tAOX1 terminator, the coding sequences of Neonothopanus nambi luciferase (SEQ ID NOs: 79, 80), Neonothopanus nambi hispidin hydroxylase (SEQ ID NOs: 1, 2), Neonothopanus nambi hispidin synthase (SEQ ID NOs: 34, 35), Neonothopanus nambi caffeoylpyruvate hydrolase (SEQ ID NOs: 64, 65), Aspergillus nidulans NpgA protein (SEQ ID NOs: 104, 105), tyrosine ammonium -lyase from Rhodobacter capsulatus (SEQ ID NOs: 106, 107), and components HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase from E. coli (SEQ ID NOs: 108-111). Each expression cassette was flanked by BsmBI restriction enzyme recognition sequences. Regions of homology to the MET6 gene of Pichia pastoris (Uniprot F2QTU9), flanked by BsmBI restriction enzyme sites, were also synthetically obtained. Synthetic DNA was treated with BsmBI restriction enzymes and combined into one plasmid according to the Golden Gate cloning protocol described in [Iverson et al., ACS Synth Biol. 2016 Jan 15;5(1):99-103]. 10 fmol of each DNA fragment was mixed in a reaction containing 1× DNA ligase buffer (Promega, USA), 20 units of DNA ligase activity (Promega, USA), 10 units of restriction endonuclease activity in a total volume of 10 μl. The resulting reaction mixture was placed in a thermal cycler and incubated at temperatures of 16°C and 37°C according to the following protocol: 25 cycles of incubation at 37°C for 1.5 min and at 16°C for 3 min, then a single incubation at 50°C for 5 min, followed by a single incubation at 80°C for 10 min. 5 μl of the reaction mixture was transformed into chemically competent E. coli cells. The correct assembly of plasmid DNA was confirmed by Sanger sequencing, and a preparation of purified plasmid DNA was used to transform Pichia pastoris GS11 cells by electroporation. Electroporation was carried out using the lithium acetate-dithiothreitol method described in [Wu and Letchworth, Biotechniques, 2004, 36:152-4]. Electroporated cells were scattered onto Petri dishes with RDB medium containing 1 M sorbitol, 2% (w/v) glucose, 1.34% (w/v) yeast base nitrogen agar (YNB), 0.005% (w/v) amino acid mixture, 0.00004% (w/v) biotin and 2% (w/v) agar. Integration of the gene cassette into the genome was confirmed using PCR with primers annealing to homology regions. The resulting yeast strain, containing the correct insertion in the genome, was able to autonomously produce light, unlike the wild-type yeast strain (Fig. 7, 8).

Example 16. Creation of transgenic autonomously bioluminescent flowering plants

[0418] To create autonomously bioluminescent flowering plants based on the pBI121 vector (Clontech, USA), a binary vector for agrobacterial transformation was created containing the coding sequences of Neonothopanus nambi luciferase (SEQ ID NO: 112) and Neonothopanus nambi hispidin hydroxylase optimized for expression in plants (SEQ ID NO: 103), Neonothopanus nambi hispidin synthase (SEQ ID NO: 113), Neonothopanus nambi caffeoyl pyruvate hydrolase (SEQ ID NO: 114) and kanamycin resistance gene, each gene under the control of the 35S promoter from the color mosaic virus cabbage Sequences for assembly of expression cassettes were obtained synthetically; vector assembly was carried out according to the Golden Gate cloning protocol described in [Iverson et al., ACS Synth Biol. 2016 Jan 15;5(1):99-103].

[0419] Arabidopsis thaliana was transformed by co-cultivating plant tissue with the bacteria Agrobacterium tumefaciens strain AGL0 [Lazo et al., Biotechnology, 1991 Oct; 9(10):963-7], containing the created binary vector. Transformation was carried out using co-culture of Arabidopsis thaliana root segments (ecotype C24) as described [Valvekens et al., 1988, Proc. Nat. Acad. Sci. USA 85, 5536-5540]. The roots of Arabidopsis plants were cultivated on Gamborg B-5 agar medium with 20 g/l glucose, 0.5 g/l 2,4-dichlorophenoxyacetic acid and 0.05 kinetin for 3 days. Then the roots were cut into pieces 0.5 cm long and transferred to 10 ml of Gamborg B-5 liquid medium with 20 g/l glucose, 0.5 g/l 2,4-dichlorophenoxyacetic acid and 0.05 kinetin and added 1.0 ml of overnight culture medium of agrobacteria. Cocultivation of explants with agrobacteria was carried out for 2-3 minutes. After this, the explants were placed on sterile filters in Petri dishes with an agar medium of the same composition. After 48 hours of incubation in a thermostat at 25°C, the explants were transferred to fresh medium with cefotaxime 500 mg/l and kanamycin 50 mg/l. After three weeks, plant regeneration began on a selective medium containing kanamycin 50 mg/l. Transgenic plants were rooted and transferred to germination medium or soil. Bioluminescence was visualized using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). More than 90% of transgenic plants emitted light that was at least two orders of magnitude higher than the signal from wild-type plants.

[0420] Nicotiana benthamiana was transformed by co-cultivating plant tissue with the bacteria Agrobacterium tumefaciens strain AGL0 [Lazo et al., Biotechnology, 1991 Oct; 9(10):963-7], containing the created binary vector. Transformation was carried out using co-culture of Nicotiana benthamiana leaf segments. Then the leaves were cut into pieces 0.5 cm long and transferred to 10 ml of liquid Gamborg B-5 medium with 20 g/l glucose, 0.5 g/l 2,4-dichlorophenoxyacetic acid and 0.05 kinetin and added 1.0 ml of overnight culture medium of agrobacteria. Cocultivation of explants with agrobacteria was carried out for 2-3 minutes. After this, the explants were placed on sterile filters in Petri dishes with an agar medium of the same composition. After 48 hours of incubation in a thermostat at 25°C, the explants were transferred to fresh medium with cefotaxime 500 mg/l and kanamycin 50 mg/l. After three weeks, plant regeneration began on a selective medium containing kanamycin 50 mg/l. Transgenic plants were rooted and transferred to germination medium or soil. Bioluminescence was visualized using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). More than 90% of transgenic plants emitted light that was at least two orders of magnitude higher than the signal from wild-type plants. Photographs of autonomously glowing Nicotiana benthamiana plants are shown in Fig. 9.

[0421] To create the autonomously bioluminescent bentgrass Agrostis stolonifera L., the coding sequences of the fungal luciferin metabolic cascade genes, optimized for expression in plants and flanked by BsaI restriction endonuclease sites: Neonothopanus nambi luciferase (SEQ ID NO: 126), Neonothopanus nambi hispidin hydroxylase (SEQ ID NO: 117), Neonothopanus nambi hispidin synthase (SEQ ID NO: 127), Neonothopanus nambi caffeoylpyruvate hydrolase (SEQ ID NO: 128) and glyphosate herbicide resistance gene (bar gene ). Each sequence was under the control of the CmYLCV promoter [Stavolone et al., Plant Mol Biol. 2003 Nov;53(5):663-73]. Sequences were synthesized using standard methods. Vector assembly was carried out using the Golden Gate cloning protocol. Transformation was carried out using the method of agrobacterial transformation of embryogenic calus. An overnight culture of the bacteria Agrobacterium tumefaciens strain AGL0 was added to the liquid medium [Lazo et al., Biotechnology, 1991 Oct; 9(10):963-7], containing the created binary vector. After two days of cocultivation on Murashige-Skoog agar medium, the plants were transferred to fresh medium supplemented with 500 mg/L cefotaxime and 10 mg/L phosphinothricin. Plant regeneration began after three weeks. Transgenic plants were transplanted onto a medium containing half the Murashige-Skoog salts and 8 mg/L phosphinothricin for rooting. Rooted plants were planted in a greenhouse. About 25% of the resulting plants with correct and complete integration into the genome of the metabolic cascade had bioluminescence exceeding that of control wild-type plants.

[0422] Of particular interest are organisms that can emit light in certain tissues or at certain times of day. Such organisms use the resources required to emit light more efficiently. To create autonomously bioluminescent roses that emit light only in the petals, several varieties of roses with white petals were selected. Based on the pBI121 vector (Clontech, USA), two binary vectors for agrobacterial transformation were created containing a metabolic cascade of coding sequences optimized for expression in plants: Neonothopanus nambi luciferase, Neonothopanus nambi hispidin hydroxylase, Neonothopanus nambi hispidin synthase, Neonothopanus nambi caffeoylpyruvate hydrolase and neomycin resistance gene. All genes except the luciferase gene were placed under the control of the cauliflower mosaic virus 35S promoter. In one of the vectors, the luciferase gene was placed under the control of the rose chalcone synthase promoter, and in the other, under the control of the chrysanthemum UEP1 promoter. Synthetic nucleic acids necessary for vector assembly, flanked by BsaI restriction enzyme recognition sites, were used; vector assembly was carried out according to the Golden Gate cloning protocol. Transgenic rose plants Rosa hybrida L. cv. Tinike was obtained by co-cultivating embryogenic callus with the bacteria Agrobacterium tumefaciens strain AGL0 [Lazo et al., Biotechnology, 1991 Oct; 9(10):963-7], containing one of the binary vectors described above. Cultivation was carried out in a liquid medium containing macro- and micro-salts of Murashige-Skoog, with the addition of kinetin 1-2 mg/l, 2,4-dichlorophenoxyacetic acid - 3 mg/l and 6-benzylaminopurine 1 mg/l for 40 minutes The callus was transferred to an agar medium of the same composition. Two days later, the explants were transferred to fresh Murashige-Skoog medium supplemented with 500 mg/L cefotaxime and 50 mg/L kanamycin. The formation and regeneration of shoots occurred after 5-8 weeks. The shoots were transferred to a propagation or rooting medium. Rooted shoots were planted in a peat mixture in a greenhouse. Flowering was observed after 8 weeks. Plants with developed flowers were imaged using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). For each of the designs tested, all plants tested autonomously emitted light that was at least three orders of magnitude greater than the signal from wild-type plants. Light emitted only from petal tissues, confirming the tissue-specific functioning of the promoters.

[0423] To create autonomously bioluminescent plants, in which bioluminescence is regulated by circadian rhythms and activated in the dark, a previously obtained binary vector for agrobacterial transformation was used, containing the coding sequences of Neonothopanus nambi luciferase, Neonothopanus nambi hispidin hydroxylase, Neonothopanus nambi hispidin synthase, caffeoyl pyruvate -hydrolases of Neonothopanus nambi and the kanamycin resistance gene, each gene under the control of the 35S promoter from cauliflower mosaic virus. It replaced the promoter for the expression of Neonothopanus nambi luciferase with the promoter of the CAT3 gene from Arabidopsis thaliana. Transcription from the CAT3 gene promoter is regulated by circadian rhythms and is activated in the evening. The CAT3 promoter sequence is known in the art [Michael and McClung, Plant Physiol. 2002 Oct;130(2):627-38]. Arabidopsis thaliana was transformed by co-cultivating plant tissue with the bacteria Agrobacterium tumefaciens strain AGL0 [Lazo et al., Biotechnology, 1991 Oct; 9(10):963-7], containing the created binary vector. Transformation was carried out using co-culture of Arabidopsis thaliana root segments (ecotype C24) as described [Valvekens et al., 1988, Proc. Nat. Acad. Sci. USA 85, 5536-5540]. The roots of Arabidopsis plants were cultivated on Gamborg B-5 agar medium with 20 g/l glucose, 0.5 g/l 2,4-dichlorophenoxyacetic acid and 0.05 kinetin for 3 days. Then the roots were cut into pieces 0.5 cm long and transferred to 10 ml of Gamborg B-5 liquid medium with 20 g/l glucose, 0.5 g/l 2,4-dichlorophenoxyacetic acid and 0.05 kinetin and added 1.0 ml of overnight culture medium of agrobacteria. Cocultivation of explants with agrobacteria was carried out for 2-3 minutes. After this, the explants were placed on sterile filters in Petri dishes with an agar medium of the same composition. After 48 hours of incubation in a thermostat at 25°C, the explants were transferred to fresh medium with cefotaxime 500 mg/l and kanamycin 50 mg/l. After three weeks, plant regeneration began on a selective medium containing kanamycin 50 mg/l. Transgenic plants were rooted and transferred to a germination medium and grown under natural day-night conditions. Bioluminescence was visualized using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer) by placing plants in the instrument for 24 hours and recording bioluminescence intensity every half hour. Plants emitted light throughout the day, but the intensity of bioluminescence was significantly modulated by circadian rhythms: the integral intensity of luminescence during the dark period of the day exceeded the integral luminosity during the day by more than 1000 times for 85% of the analyzed plants.

Example 17. Creation of transgenic autonomously bioluminescent lower plants

[0424] The autonomously bioluminescent moss Physcomitrella patens was generated by co-transformation of protoplasts with plasmids in the manner described in Example 8. Expression cassettes were synthetically produced including coding sequences optimized for expression in plants for Neonothopanus nambi luciferase (SEQ ID NO: 112), hispidin hydroxylase Neonothopanus nambi (SEQ ID NO: 103), Neonothopanus nambi hispidin synthase (SEQ ID NO: 113), Neonothopanus nambi caffeoyl pyruvate hydrolase (SEQ ID NO: 114) and kanamycin resistance gene, each under the control of the rice actin 2 promoter. Expression cassettes were operatively stitched together in the pBI121 vector (Clontech, USA) so that the construct, including the complete metabolic cascade and the kanamycin resistance gene, was flanked by sequences matching the sequence of the target locus in the moss genome. Vector assembly was carried out according to the Golden Gate cloning protocol [Iverson et al., ACS Synth Biol. 2016 Jan 15;5(1):99-103]. The guide RNA gene, complementary to the target region in the moss genome, was also cloned into the vector. The plasmid with the listed genes was cotransformed with a plasmid for constitutive expression of Cas9 nuclease, according to the polyethylene glycol transformation protocol described in [Cove et al., Cold Spring Harb Protoc., 2009, 2]. The resulting transformed protoplasts were incubated without light for 24 hours in BG-11 medium and then transferred to Petri dishes with BG-11 medium and 8.5% agar. One month after growing on plates under continuous light irradiation, imaging was performed using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). 70% of the plants tested emitted light at least an order of magnitude higher than the signal from wild-type plants.

Example 18. Production of transgenic luminescent animals

[0425] Transgenic fish Danio rerio containing the Neonothopanus nambi hispidin hydroxylase gene were created according to the method described in [Hisano et al., Sci Rep., 2015, 5:8841]. The technique involves expression of a guide RNA and Cas9 nuclease to create a point break in a region homologous to the guide RNA sequence. To create transgenic animals, synthetic DNA fragments were ordered containing sequences of guide RNA from plasmid pX330, Addgene #42230 and Cas9 nuclease mRNA under the control of the bacteriophage T7 polymerase promoter. The resulting fragments were used for in vitro transcription using reagents from the MAXIscript T7 kit (Life Technologies, USA), and the synthesized RNA was purified using an RNA isolation kit (Evrogen, Russia).

[0426] The coding sequence of Neonothopanus nambi hispidin hydroxylase, flanked by 50 nucleotide sequences from the krtt1c19e gene of Danio rerio, described in [Hisano et al., Sci Rep., 2015, 5:8841] was obtained synthetically and cloned into the backbone of the plasmid pEGFP/ C1, containing the pUC origin of replication and the kanamycin resistance cassette. The resulting vector, Cas9 nuclease mRNA and guide RNA were dissolved in injection buffer (40 mM HEPES (pH 7.4), 240 mM KCl with the addition of 0.5% phenol red) and injected into 1-2-cell embryos of a previously obtained line of Danio rerio, stably expressing luciferase from Neonothopanus nambi, in a volume of about 1-2 nl. Of the 48 embryos, approximately 12 embryos survived the injection and showed normal development on the fourth day after fertilization.

[0427] To record the bioluminescent signal, a hispidin solution was injected intravenously into Danio rerio larvae according to the method described in [Cosentino et al., J Vis Exp.2010; (42): 2079]. Bioluminescence was recorded using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). After recording, genomic DNA was isolated from larvae to confirm integration of the hispidin hydroxylase gene into the genome. All larvae with correct integration of the Neonothopanus nambi hispidin hydroxylase gene into the genome demonstrated bioluminescence with an intensity at least two orders of magnitude higher than the intensity of the signal coming from wild-type fish after injection of a hispidin solution.

Example 19. Study of the effect of caffeoylpyruvate hydrolase on the luminescence of autonomously bioluminescent organisms

[0428] To study the effects of caffeilpyuruvat-hydrolasis on the glow of autonomously biooluminescent organisms, a binary vector was used for agricultural-bacterial transformation containing the encoding sequences of Neonothopanus nambi lucifers, neonothopanus nambi hydroxylasis, gypidin-syntase neonethopanus nam Bi, Coffeelap-Hydroles Neonothopanus Nambi and a gene of resistance to kanamycin , each gene - under the control of the 35S promoter from cauliflower mosaic virus, obtained as described in Example 16 and a control vector, characterized in that the caffeoylpyruvate hydrolase sequence has been removed from it. The vectors were used to transform Arabidopsis thaliana under identical conditions using the protocol described in Example 16. Bioluminescence was visualized using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). A comparison of the bioluminescence intensities of plants expressing all four genes of the bioluminescent system of Neonothopanus nambi with plants expressing only luciferase, hispidin hydroxylase and hispidin synthase revealed that plants additionally expressing caffeoylpyruvate hydrolase have an average of 8.3 times brighter bioluminescence. These data indicate that the expression of caffeoylpyruvate hydrolase increases the efficiency of the bioluminescent cascade, which leads to an increase in the intensity of light emitted by plants.

Example 20. Effect of external addition of caffeic acid on the bioluminescence of transgenic organisms

[0429] Autonomously bioluminescent transgenic Nicotiana benthamiana plants prepared as described in Example 16 were transferred to soil and cultivated for eight weeks. The plant stem was then cut and placed in water for two hours, after which the bioluminescence intensity was measured using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). The plants were then transferred to one of five aqueous solutions with caffeic acid concentrations of 0.4 g/L, 0.8 g/L, 1.6 g/L, 3.2 g/L, or 6.4 g/L, and control plants were placed in water. After another two hours of incubation in caffeic acid solution or water, bioluminescence was measured again. In all cases, the intensity of bioluminescence of plants incubated in a caffeic acid solution increased compared to the intensity before being placed in a caffeic acid solution, with the greatest changes observed for plants incubated in a solution with a concentration of 6.4 g/L. Control plants incubated in water did not show a significant change in bioluminescence intensity within four hours after the start of incubation.

Example 21. Use of genes of the bioluminescent system of fungi to analyze the activity of promoters and intracellular logical integration of external signals.

[0430] The coding sequences of Neonothopanus nambi hispidin synthase, hispidin hydroxylase and luciferase were used to monitor the simultaneous activation of multiple promoters. Synthetic expression cassettes containing the coding sequence of Neonothopanus nambi hispidin synthase (SEQ ID NOs: 34, 35) under the control of the arabinose-inducible E. coli araBAD promoter, the coding sequence of hispidin hydroxylase (SEQ ID NOs 1, 2) under the control of the IPTG-inducible T7 promoter /lacO, and the luciferase gene (SEQ ID NOs: 79, 80) under the control of the rhamnose-inducible promoter pRha, as well as the NpgA gene (SEQ ID NOs: 104, 105) under the control of the constitutive promoter J23100 (Registry of Standard Biological parts, Part:BBa_J23100 ). The resulting synthetic nucleic acids were cloned into the MoClo_Level2 vector [Weber et al., PLoS One. 2011 Feb 18;6(2):e16765] instead of the insert containing the LacZ gene using the restriction endonuclease BpiI. The resulting vector was transformed into competent cells of the E. coli strain BL21 (NEB, USA), containing a genomic copy of the bacteriophage T7 polymerase.

[0431] To determine the possibility of recording the simultaneous activation of several promoters, the cells obtained in the previous step were grown overnight in a flask in LB medium with a volume of 100 ml with the addition of ampicillin at a concentration of 100 mg/l. The next day, aliquots of the cell culture were placed for 120 minutes at a temperature of 24°C and shaking at 200 rpm in one of the following media:

1. LB medium with the addition of 1% arabinose,

2. LB medium with the addition of 0.2% rhamnose,

3. LB medium with the addition of 0.5% IPTG,

4. LB medium with the addition of 1% arabinose and 0.2% rhamnose,

5. LB medium supplemented with 1% arabinose and 0.5% IPTG,

6. LB medium supplemented with 0.2% rhamnose and 0.5% IPTG,

7. LB medium supplemented with 1% arabinose, 0.2% rhamnose and 0.5% IPTG,

8. LB medium (control).

[0432] After incubation, the cells were pelleted by centrifugation, the medium was replaced with phosphate-buffered saline with pH 7.4 (Sigma-Aldrich, USA) with the addition of caffeic acid (Sigma-Aldrich, USA) at a concentration of 1 g/l, the cells were resuspended by pipetting. Cell bioluminescence was analyzed after half an hour using a GloMax 20/20 luminometer (Promega, USA). The experiment was repeated in triplicate. Of the eight analyzed samples, the intensity of bioluminescence was significantly different from the bioluminescence of control bacteria incubated in LB medium (medium No. 8), only for bacteria incubated in medium No. 7 (LB medium with the addition of 1% arabinose, 0.2% rhamnose and 0.5% IPTG) . Thus, the luminescence of bacteria made it possible to judge that the bacteria were placed in an environment that provided simultaneous activation of three different promoters. In the experiment, bacterial cells integrated information about the presence of substances inducing the activity of promoters in the external environment and signaled by luminescence only in the case when all three substances were present in the environment at the same time, intracellularly performing the logical “AND” operation.

[0433] Synthetic expression cassettes containing (1) a hispidin hydroxylase coding sequence (SEQ ID NOs: 1, 2) under the control of the Odf2 promoter according to [Pletz et al., Biochim Biophys Acta. 2013 Jun;1833(6):1338-46]; (2) the coding sequence of hispidin synthase (SEQ ID NOs: 34, 35) under the control of the promoter of the cyclin-dependent kinase CDK7; (3) luciferase (SEQ ID NOs: 79, 80) under the control of the CCNH gene promoter was cloned into the pmKate2-keratin vector (Evrogen, Russia) instead of the sequences of the cytomegalovirus promoter and mKate2-keratin insert. Also, the coding sequence of the NpgA gene (SEQ ID NOs: 104, 105) was cloned into the pmKate2-keratin vector instead of the mKate2-keratin insertion sequence. All resulting vectors were co-transfected into HEK293T cells using the FuGENE HD transfection reagent (Promega, USA) according to the manufacturer’s protocol. 24 hours after transfection, caffeic acid was added to the medium at a concentration of 5 mg/ml, and cell luminescence was detected using a Leica TCS SP8 microscope. Light emission allowed us to determine the simultaneous activation of the Odf2, CCNH and CDK7 promoters, and the intensity of the emitted light was related to the cell cycle stage.

[0434] The data obtained indicate that the genes of the bioluminescent system of fungi can be used to monitor the simultaneous activity of several promoters, to detect the presence of various substances and their combinations in the environment, as well as for the logical integration of external signals inside the cell.

Example 22. Detection of hispidin in plant extracts

[0435] The coding sequences for Neonothopanus nambi hispidin hydroxylase and luciferase, obtained as described in Example 1, were cloned into the pET23 vector under the control of the T7 promoter. Purified plasmid DNA preparations were used for transcription and translation of proteins in vitro using the PURExpress In Vitro Protein Synthesis Kit (NEB, USA). The resulting reaction mixture was used to analyze the presence and concentration of hispidin and its functional analogues in the lysates of about 19 different plants (Chrysanthemum sp., Ananas comosus, Petunia atkinsiana, Picea abies, Urtica dioica, Solanum lycopersicum, Nicotiana benthamiana, Nicotiana tobacum, Arabidopsis thaliana, Rosa glauca, Rosa rubiginosa, Equisetum arvense, Equisetum telmateia, Polygala sabulosa, Rosa rugosa, Clematis tashiroi, Kalanchoe sp., Triticum aestivum, Dianthus caryophyllus), adding 2 μl of plant lysate to 100 μl of the reaction mixture and recording the intensity of emitted light using a GloMax luminometer (Promega, USA). It was determined that the maximum concentration of hispidin and its functional analogs is contained in lysates of horsetails Equisetum arvense and Equisetum telmateia. Also, hispidin or its functional analogues were found in the lysates of Polygala sabulosa, Rosa rugosa and Clematis tashiroi.

Example 23: Identification of PKS capable of catalyzing the synthesis of hispidin and their use for the production of hispidin in in vitro and in vivo systems.

[0436] Fungal luciferin precursors, such as hispidin, belong to the group of polyketide derivatives. It is known from the prior art that the enzymes involved in the synthesis of polyketides in plants belong to the protein superfamily of polyketide synthases, and unlike fungal polyketide synthases, plant polyketide synthases are relatively compact proteins that use CoA esters of acids, including 3-arylacrylic acids, as substrates .No polyketide synthase was known from the prior art that could catalyze the reaction of converting the CoA ester of caffeic acid into hispidin, but hispidin is found in many plant organisms.

[0437] Using bioinformatics analysis, 11 polyketide synthases potentially capable of catalyzing hispidin synthesis were selected from the following sources:

Aquilaria sinensis (2 enzymes),

Hydrangea macrophylla,

Arabidopsis thaliana,

Physcomitrella patens,

Polygonum cuspidatum,

Rheum palmatum,

Rheum tataricum,

Wachendorfia thyrsiflora,

Piper methysticum (two enzymes).

[0438] Selected nucleotide sequences were optimized for expression in cells of the yeast Pichia pastoris and the plant Nicotiana benthamiana. The resulting nucleic acids were synthesized, cloned into the pGAPZ vector, and used to test the ability of the expressed proteins to synthesize hispidin.

[0439] To do this, the pGAPZ plasmid containing the Arabidopsis thaliana coumarate-CoA ligase 1 gene (nucleotide and amino acid sequences for it are shown in SEQ ID NOs: 140, 141), also obtained using oligonucleotide synthesis. The plasmid was linearized at the AvrII restriction site and used for transformation into Pichia pastoris GS115 cells.

[0440] The resulting yeast cells constitutively expressing Neonothopanus nambi luciferase and hispidin hydroxylase and Arabidopsis thaliana coumarate-CoA ligase 1 were transfected with linearized plasmids containing PKS coding sequences and plated on Petri dishes with RDB medium containing 1 M sorbitol, 2 % (w/v) glucose, 1.34% (w/v) yeast base nitrogen agar (YNB), 0.005% (w/v) amino acid mixture, 0.00004% (w/v) biotin and 2% (w/v) agar . To identify enzymes with hispidin synthase activity, the resulting colonies were sprayed with a solution of caffeic acid, detecting the presence of hispidin synthase in the cells by the appearance of bioluminescence. The light emitted by the colonies was detected using IVIS Spectrum CT (PerkinElmer, USA). A yeast line constitutively expressing luciferase, hispidin hydroxylase, and coumarate-CoA ligase 1, as well as wild-type yeast cells, were used as negative controls. Among the analyzed genes, 11 enzymes had hispidin synthase activity, the sequence of which is shown in SEQ ID NOs: 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138. The amino acid sequences they encode are shown in SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139 respectively. The enzymes exhibiting the greatest activity were PKS1 and PKS2 from Aquilaria sinensis (SEQ ID NOs:119, 121), PKS from Arabidopsis thaliana (SEQ ID NO:123) and PKS from Hydrangea macrophylla (SEQ ID NO:125).

[0441] Nucleic acid encoding PKS from Hydrangea macrophylla (SEQ ID NOs: 124, 125) was used to produce the recombinant protein according to the procedure described in Example 4. The presence of the expected recombinant product was confirmed by electrophoresis by the presence of bands of the expected length. Aliquots of the isolated recombinant protein were used to test functionality: 30 μl of a solution of isolated recombinant protein was added to a tube containing 100 μl of buffer (0.2 M Na-phosphate buffer, 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM) pH 8.0, all components - Sigma -Aldrich, USA), 0.5 μl of purified recombinant Neonothopanus nambi luciferase, obtained as described in Example 4, 1 mM NADPH (Sigma-Aldrich, USA), 15 μl of purified recombinant Neonothopanus nambi hispidin hydroxylase, obtained as described in Example 4, 10 mM ATP (ThermoFisher Scientific, USA), 1 mM CoA (Sigma-Aldrich, USA), 1 mM Malonyl-CoA (Sigma-Aldrich, USA). The tube was placed in a GloMax 20/20 luminometer (Promega, USA). The reaction mixtures exhibited bioluminescence when 20 μM caffeyl-CoA was added to the solution. The emitted light had an emission maximum in the region of 520-535 nm.

[0442] Nucleic acid encoding PKS2 from Aquilaria sinensis (SEQ ID NO:120, 121) was used to obtain a hispidin producer strain. To do this, we synthesized an expression cassette containing the nucleic acid SEQ ID NO: 120 under the control of the constitutive promoter J23100, and an expression cassette containing the nucleic acid SEQ ID NO: 140 encoding 4-coumarate-CoA ligase 1 from Arabidopsis thaliana, under the control of the araBAD promoter ; both expression cassettes were flanked by regions of homology to the SS9 site. The expression cassettes were cloned into a bacterial expression vector containing a zeocin resistance cassette and used for transformation and integration into the genome of E. coli BW25113 by bacteriophage lambda protein-mediated recombination as described in Bassalo et al. [ACS Synth Biol. 2016 Jul 15;5(7):561-8], using selection for resistance to zeocin. Integration of the full-length construct was confirmed using PCR with primers specific to the SS9 homology regions, and then the correctness of the integrated construct was verified by sequencing the PCR product of genomic DNA using the Sanger method.

[0443] The resulting E. coli strain was used to produce hispidin. In the first step, bacteria were cultured in five 50-ml plastic tubes in LB medium for 10 hours with shaking at 200 rpm at 37°C. 250 ml of the resulting culture was added to 3.3 liters of fermentation medium in a Biostat B5 fermenter (Braun, Germany) so that the initial optical density of the culture at 600 nm was about 0.35. _The fermentation medium contained 10 g/L peptone, 5 g/L caffeic acid, 5 g/L yeast extract, 10 g/L NaCl, 25 g/L glucose, 15 g/L ( _NH4 ) _2SO4 , 2 g/L l KH ₂ PO ₄ , 2 g/l MgSO ₄ 7 × H ₂ O, 14.7 × mg/l CaCl ₂ , 0.1 mg/l thiamine, 1.8 mg/l, and 0.1% solution of the following composition: EDTA 8 mg/l , CoCl ₂ 6×H ₂ O 2.5 mg/l, MnCl ₂ 4H ₂ O 15 mg/l, CuCl ₂ 2H ₂ O 1.5 mg/l, H ₃ BO ₃ 3 mg/l, Na ₂ MoO4 2H ₂ O 2.5 mg/l, Zn(CH ₃ COO) ₂ 2H ₂ O 13 mg/l, iron(III) citrate 100 mg/l, thiamine hydrochloride 4.5 mg/l. Fermentation was carried out at 37°C, with aeration 3×l/min and stirring at 200 rpm. After 25 hours of cultivation, arabinose was added to the culture to a final concentration of 0.1 × mM. The pH was controlled automatically by adding NH4OH, bringing the pH to 7.0. A solution containing glucose 500 g/l, caffeic acid 5 g/l, arabinose 2 g/l, tryptone 25 g/l, yeast extract 50 g/l, MgSO4 7H ₂ O 17.2 g/l, (NH ₄ )SO ₄ 7.5 g/L, ascorbic acid 18 g/L were added to the fermenter to maintain glucose levels each time the pH increased to 7.1. After 56 hours of cultivation, the concentration of hispidin in the medium was 3.48×g/L. The fermenter medium, as well as hispidin purified from it using high-performance liquid chromatography, had activity in a bioluminescent reaction with hispidin hydroxylase and Neonothopanus nambi luciferase, obtained as described in Example 4.

[0444] To create autonomously bioluminescent yeast Pichia pastoris, expression cassettes containing, under the control of the GAP promoter and tAOX1 terminator, the coding sequences of Neonothopanus nambi luciferase (SEQ ID NOs: 79, 80), Neonothopanus nambi hispidin hydroxylase (SEQ ID NOs: 1, 2), Neonothopanus nambi hispidin synthase (SEQ ID NOs: 34, 35), Neonothopanus nambi caffeoylpyruvate hydrolase (SEQ ID NOs: 64, 65), Rhodobacter capsulatus tyrosine ammonium lyase (SEQ ID NOs: 106, 107), and components HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase of E. coli (SEQ ID NOs: 108-111), described as described in Example 15, and similar expression cassettes containing 4-coumarate-CoA coding sequences were synthesized -ligase 1 from Arabidopsis thaliana (SEQ ID NOs: 140, 141) and three PKS: from Aquilaria sinensis (SEQ ID NOs: 120, 121), PKS from Arabidopsis thaliana (SEQ ID NOs: 122, 123) and PKS from Hydrangea macrophylla (SEQ ID NO:124, 125). Each expression cassette was flanked by BsmBI restriction enzyme recognition sequences. Regions of homology to the MET6 gene of Pichia pastoris (Uniprot F2QTU9), flanked by BsmBI restriction enzyme sites, were also synthetically obtained. Synthetic DNA was treated with BsmBI restriction enzymes and combined into one plasmid according to the Golden Gate cloning protocol described in [Iverson et al., ACS Synth Biol. 2016 Jan 15;5(1):99-103]. Three plasmids were produced, differing in the PKS included in their composition. The resulting plasmids were used to obtain transgenic yeast Pichia pastoris according to the method described in Example 15. Integration of the gene cassette into the genome was confirmed using PCR from primers that burned to homology sites. All three resulting yeast strains containing the correct insertion in the genome were able to autonomously produce light, unlike the wild-type yeast strain.

[0445] To create autonomously bioluminescent flowering plants based on the pBI121 vector (Clontech, USA), a set of binary vectors for agrobacterial transformation was created containing the coding sequences of Neonothopanus nambi luciferase (SEQ ID NO: 112), Neonothopanus hispidin hydroxylase optimized for expression in plants nambi (SEQ ID NO: 103), Neonothopanus nambi caffeoylpyruvate hydrolase (SEQ ID NO: 114), kanamycin resistance gene, and PKS (SEQ ID NOs: 122, 123), each gene under the control of the 35S promoter from the color mosaic virus cabbage Sequences for assembly of expression cassettes were obtained synthetically; vector assembly was carried out according to the Golden Gate cloning protocol described in [Iverson et al., ACS Synth Biol. 2016 Jan 15;5(1):99-103]. Nicotiana tabacum was transformed by co-cultivating plant tissue with the bacteria Agrobacterium tumefaciens strain AGL0 [Lazo et al., Biotechnology, 1991 Oct; 9(10):963-7], containing the created binary vector. Transformation was carried out using co-culture of Nicotiana tabacum leaf segments. Then the leaves were cut into pieces 0.5 cm long and transferred to 10 ml of liquid Gamborg B-5 medium with 20 g/l glucose, 0.5 g/l 2,4-dichlorophenoxyacetic acid and 0.05 kinetin and added 1.0 ml of overnight culture medium of agrobacteria. Cocultivation of explants with agrobacteria was carried out for 2-3 minutes. After this, the explants were placed on sterile filters in Petri dishes with an agar medium of the same composition. After 48 hours of incubation in a thermostat at 25°C, the explants were transferred to fresh medium with cefotaxime 500 mg/l and kanamycin 50 mg/l. After three weeks, plant regeneration began on a selective medium containing kanamycin 50 mg/l. Transgenic plants were rooted and transferred to germination medium or soil. Bioluminescence was visualized using an IVIS Spectrum In Vivo Imaging System (Perkin Elmer). The transgenic plants emitted light that was at least three orders of magnitude greater than the signal from wild-type plants.

Example 24 Nucleic Acid Combinations

[0446] Combination 1:

[0447] Composition: (a) Nucleic acid encoding hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26 , 28; and (b) A nucleic acid encoding a luciferase, the amino acid sequence of which is selected from the group 80, 82, 84, 86, 88, 90, 92, 94, 96, 98.

[0448] The combination can be used to produce bioluminescence in in vitro or in vivo expression systems in the presence of a substance selected from the group of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-ones having the structural formula

,

in which the substituent at the sixth position is a 2-arylvinyl or 2-heteroarylvinyl substituent (R-CH=CH-), including 2-(3,4-dihydroxystyryl), 2-(4-hydroxystyryl), 2-(4- (diethylamino)styryl), 2-(2-(1H-indol-3-yl)vinyl), 2-(2-(1,2,3,5,6,7-hexahydropyrido[3,2,1-ij ]quinolin-9-yl)vinyl), 2-(6-hydroxynaphthalene-2-yl)vinyl.

[0449] Also, this combination can be used to study the dependence of two promoters in heterologous expression systems.

[0450] Also, this combination can be used to detect hispidin and its analogues in biological samples.

[0451] This combination can also be used to label cells using bioluminescence. Appearing in the presence of hispidin and its functional analogues.

[0452] Combination 2:

[0453] Composition: (a) Nucleic acid encoding hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26 , 28, and (b) A nucleic acid encoding hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55.

[0454] The combination can be used to produce fungal luciferin in in vitro or in vivo expression systems from a substance selected from a substituted acrylic acid with the structural formula

[0455] ,

where R is aryl or heteroaryl (for example, from caffeic acid).

[0456] Combination 3:

[0457] Includes all components specified in Combination 2, as well as a nucleic acid encoding a luciferase, the amino acid sequence of which is selected from the group 80, 82, 84, 86, 88, 90, 92, 94, 96, 98.

[0458] The combination can be used to produce bioluminescence in in vitro or in vivo expression systems in the presence of a substance selected from the substituted acrylic acid with the structural formula

[0459] ,

where R is aryl or heteroaryl.

[0460] The combination can be used to produce bioluminescent cells and transgenic organisms. Also, this combination can be used to study the dependence of three promoters in heterologous expression systems.

[0461] Combination 4.

[0462] Includes all components specified in Combination 3, also a nucleic acid encoding caffeoyl pyruvate hydrolase, the amino acid sequence of which is selected from the group SEQ ID NOs: 65, 67, 69, 71, 73, 75.

[0463] The combination can be used to produce bioluminescent cells and transgenic organisms.

[0464] Combination 5.

[0465] Composition: (a) Nucleic acid encoding hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55; and (b) A nucleic acid encoding a 4'-phosphopantotheinyl transferase gene, the amino acid sequence of which is shown in SEQ ID NO: 105

[0466] The combination can be used to produce hispidin from caffeic acid in in vitro and in vivo expression systems.

[0467] Combination 6

[0468] Composition: (a) Nucleic acid encoding hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55; (b) A nucleic acid encoding a 4'-phosphopantotheinyl transferase gene, the amino acid sequence of which is shown in SEQ ID NO: 105; and (c) nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid with the structural formula

[0469] ,

where R is aryl or heteroaryl from cell metabolites (for example, nucleic acids encoding tyrosine ammonium lyase and components HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase).

[0470] The combination can be used to produce hispidin from tyrosine in in vitro and in vivo expression systems.

[0471] Combinations 2-4 may also include the coding sequence of the NpgA 4'-phosphopantotheinyl transferase gene (SEQ ID NOs: 104, 105) or other enzyme exhibiting the same activity.

[0472] Combination 7

[0473] Composition: (a) Nucleic acid encoding hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26 , 28, and (b) A nucleic acid encoding a PKS, the amino acid sequence of which is selected from the group of SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139.

[0474] The combination can be used to produce 3-hydroxyhispidine in in vitro or in vivo expression systems from caffeoyl-CoA.

[0475] Combination 8

[001] Includes all components specified in Combination 7, as well as a nucleic acid encoding a luciferase, the amino acid sequence of which is selected from the group 80, 82, 84, 86, 88, 90, 92, 94, 96, 98. The combination can be used to obtain bioluminescence in in vitro or in vivo expression systems in the presence of caffeoyl-CoA.

[0476] Combination 9

[0477] Includes all components specified in Combination 8, also a nucleic acid encoding caffeoyl pyruvate hydrolase, the amino acid sequence of which is selected from the group SEQ ID NOs: 65, 67, 69, 71, 73, 75.

[0478] The combination can be used to produce bioluminescent cells and transgenic organisms.

[0479] Combination 10

[0480] Composition: (a) A nucleic acid encoding a PKS, the amino acid sequence of which is selected from the group of SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139; and (b) a nucleic acid encoding 4-coumarate CoA ligase 1 from Arabidopsis thaliana, the amino acid sequence of which is shown in SEQ ID NO: 141.

[0481] The combination can be used to produce hispidin from caffeic acid in in vitro and in vivo expression systems.

[0482] Combination 11

[0483] Includes all components specified in Combination 10, also nucleic acids encoding caffeic acid biosynthetic enzymes (e.g., nucleic acids encoding tyrosine ammonium lyase and components HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase).

[0484] The combination can be used to produce hispidin from tyrosine in in vitro and in vivo expression systems.

[0485] Example 25: Recombinant Protein Combinations

[0486] Combination 1

[0487] Composition: (a) hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28; and (b) hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55.

[0488] The combination can be used to obtain mushroom luciferin from a substance selected from 3-arylacrylic acid with the structural formula ,

where R is aryl or heteroaryl (for example from caffeic acid).

[0489] Combination 2

[0490] Includes the components indicated in combination 1, as well as luciferase, the amino acid sequence of which is selected from the group 80, 82, 84, 86, 88, 90, 92, 94, 96, 98.

[0491] The combination can be used to detect the presence of 3-arylacrylic acid with the structural formula

[0492] ,

where R is aryl or heteroaryl (for example, from caffeic acid).

[0493] Combination 3

[0494] Composition: (a) hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28; and (b) PKS, the amino acid sequence of which is selected from the group SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139; and (c) 4-coumarate-CoA ligase 1 from Arabidopsis thaliana, the amino acid sequence of which is shown in SEQ ID NO: 141. The combination can be used to produce fungal luciferin from caffeic acid.

Example 25: Sets

[0495] In the examples below, nucleic acids can be included in expression cassettes or vectors and operatively linked to regulatory elements for their expression in a host cell. Alternatively, the nucleic acid may contain flanking sequences for insertion into the target vector. Nucleic acids can be incorporated into promoterless vectors for convenient cloning of target regulatory elements.

[0496] Reagent Set #1 includes a purified hispidin synthase preparation of the present invention and can be used to obtain hispidin from caffeic acid. The kit can also be used to obtain another 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

from the corresponding 3-arylacrylic acid with the structural formula ,

where R is aryl or heteroaryl.

[0497] The reagent kit may also include a reaction buffer. For example, 0.2 M sodium phosphate buffer (pH 8.0) with the addition of 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM), 1 mM NADPH, 10 mM ATP, 1 mM CoA, 1 mM Malonyl-CoA or components for preparing the reaction buffer .

[0498] The reagent kit may also include deionized water.

[0499] The reagent kit may also include instructions for use of the kit.

[0500] Reagent Set #2 includes a purified hispidin synthase preparation of the present invention and a purified hispidin hydroxylase preparation of the present invention and can be used to obtain fungal luciferin from a substance selected from 3-aryl acrylic acid with the structural formula ,

where R is aryl or heteroaryl (for example, from caffeic acid).

[0501] The reagent kit may also include a reaction buffer: 0.2 M sodium phosphate buffer (pH 8.0) with the addition of 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM), 1 mM NADPH, 10 mM ATP, 1 mM CoA, 1 mM Malonyl-CoA or components for preparing the reaction buffer.

[0502] The reagent kit may also include deionized water.

[0503] The reagent kit may also include instructions for use of the kit.

[0504] Reagent Set #3 includes a purified hispidin hydroxylase preparation of the present invention and can be used to obtain fungal luciferin from 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one having the structural formula ,

where R is aryl or heteroaryl. For example, the kit can be used to obtain 3-hydroxyhispidin from hispidin.

[0505] The reagent kit may also include a reaction buffer. For example, 0.2 M sodium phosphate buffer (pH 8.0) with the addition of 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM), 1 mM NADPH.

[0506] The reagent kit may also include deionized water.

[0507] The reagent kit may also include instructions for use of the kit.

[0508] Reagent Kits #4 and #5 differ from Kits #2 and #3 in that they contain purified luciferase whose substrate is 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having a structural formula ,

where R is aryl or heteroaryl.

[0509] The kits can be used to detect 3-arylacrylic acid with the structural formula ,

where R is aryl or heteroaryl (for example, from caffeic acid) and/or 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

,

where R is aryl or heteroaryl, (for example, hispidine) in biological samples, for example, in plant extracts, fungal extracts and microorganisms.

[0510] Reagent kits may also include a reaction buffer (as described in kits 2 and 3) for performing the reaction or components for preparing a reaction buffer.

[0511] The reagent kit may also include deionized water.

[0512] The reagent kit may also include instructions for use of the kit.

[0513] The reagent kit may also include caffeic acid. For example, an aqueous solution of caffeic acid or a precipitate for dilution in water.

[0514] The reagent kit may also include hispidin.

[0515] Methods of using kits

[0516] To determine the presence of caffeic acid in the test sample, add 5 μl of the enzyme mixture to 95 μl of reaction buffer thawed on ice to the cuvette, mix gently, add 5 μl of the analyzed sample, mix gently again and place in the luminometer. Integrate the bioluminescent signal for two minutes under temperature conditions not exceeding 30°C. Under the same conditions, carry out control measurements with the addition of 5 μl of caffeic acid solution or 5 μl of water instead of an aliquot of the analyzed sample. The presence of caffeic acid in a sample in detectable quantities can be indicated if the light emitted by the analyzed sample exceeds the background signal recorded from the water sample.

[0517] Sensitivity: the kit allows you to determine the presence of caffeic acid in a medium in a concentration exceeding 1 nM.

[0518] Storage conditions: All components of the kit should be stored at a temperature not exceeding -20°C.

[0519] To determine the presence of hispidin in the test sample, add 5 μl of the enzyme mixture to 95 μl of reaction buffer thawed on ice to the cuvette, mix gently, add 5 μl of the analyzed sample, mix gently again and place in the luminometer. Integrate the bioluminescent signal for two minutes under temperature conditions not exceeding 30°C. Under the same conditions, carry out control measurements by adding 5 μl of hispidin solution or 5 μl of water instead of an aliquot of the analyzed sample. The presence of hispidin in a sample in detectable quantities can be indicated if the light emitted by the analyzed sample exceeds the background signal recorded from the sample with water.

[0520] Sensitivity: the kit allows you to determine the presence of hispidin in a medium at a concentration exceeding 100 pM.

[0521] Storage conditions: All components of the kit should be stored at a temperature not exceeding -20°C.

[0522] Reagent kit #6 includes a nucleic acid encoding the hispidin hydroxylase of the present invention. For example, hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28.

[0523] The reagent kit may also contain instructions for use of the nucleic acid.

[0524] The reagent kit may also contain deionized water or a buffer to dissolve the lyophilized nucleic acid and/or dilute the nucleic acid solution.

[0525] The reagent kit may also contain primers complementary to regions of said nucleic acid to amplify the nucleic acid or a fragment thereof.

[0526] The reagent kit can be used to produce recombinant hispidin hydroxylase of the present invention or to express hispidin hydroxylase in cells and/or cell lines and/or organisms. Once a nucleic acid is expressed in cells, cell lines and/or organisms, these cells, cell lines and/or organisms acquire the ability to catalyze the conversion reaction of exogenous or endogenous 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having a structural formula

,

to 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula ,

where R is aryl or heteroaryl. For example, they acquire the ability to catalyze the conversion of hispidin to 3-hydroxyhispidin.

[0527] Reagent kit #7 includes a nucleic acid encoding the hispidin synthase of the present invention. For example, hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55.

[0528] The reagent kit may also contain instructions for use of the nucleic acid.

[0529] The reagent kit may also contain deionized water or a buffer to dissolve the lyophilized nucleic acid and/or dilute the nucleic acid solution.

[0530] The reagent kit may also contain primers complementary to regions of said nucleic acid to amplify the nucleic acid or a fragment thereof.

[0531] The reagent kit may also contain a nucleic acid encoding a 4'-phosphopantotheinyl transferase, for example, a 4'-phosphopantotheinyl transferase having the amino acid sequence shown in SEQ ID NO 105.

[0532] The reagent kit can be used to produce recombinant hispidin synthase of the present invention or to express hispidin hydroxylase in cells and/or cell lines and/or organisms.

[0533] Upon expression of a nucleic acid in cells, cell lines and/or organisms, these cells, cell lines and/or organisms acquire the ability to catalyze the conversion reaction of 3-arylacrylic acid with the structural formula

[0534] ,

where R is aryl or heteroaryl, in 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula

.

[0535] For example, they acquire the ability to catalyze the reaction of caffeic acid to hispidin and/or cinnamic acid to (E)-4-hydroxy-6-styryl-2H-pyran-2-one and/or p-coumaric acid to bisnoryangonin and/or (E)-3-(6-hydroxynaphthalene-2-yl)propenoic acid in (E)-4-hydroxy-6-(2-(6-hydroxynaphthalene-2-yl)vinyl)-2H-pyran-2-one and/or (E)-3-(1H-indol-3-yl)propenoic acid in (E)-6-(2-(1H-indol-3-yl)vinyl)-4-hydroxy-2H-pyran- 2-on.

[0536] The reagent kit may also contain nucleic acids encoding tyrosine ammonium lyase and the HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase components. In this composition, the kit can be used to obtain hispidin from tyrosine in in vitro and in vivo expression systems.

[0537] Reagent set #8 includes a nucleic acid encoding the hispidin synthase of the present invention and a nucleic acid encoding the hispidin hydroxylase of the present invention. For example, hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NOs: 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55 and hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28.

[0538] The reagent kit may also contain instructions for use of the nucleic acids.

[0539] The reagent kit may also contain deionized water or a buffer to dissolve the lyophilized nucleic acid and/or dilute the nucleic acid solution.

[0540] The reagent kit may also contain primers complementary to regions of the nucleic acids included in the kit to amplify those nucleic acids or fragments thereof.

[0541] The reagent kit may also contain a nucleic acid encoding a 4'-phosphopantotheinyl transferase, for example, a 4'-phosphopantotheinyl transferase having the amino acid sequence shown in SEQ ID NO 105.

[0542] The reagent kit may also contain nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid from cellular metabolites, for example, encoding tyrosine ammonium lyase and components HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase.

[0543] The kit can be used for any of the applications described for kits 6 and 7. The kit can also be used for the expression of hispidin hydroxylase and hispidin synthase in cells and/or cell lines and/or organisms. Upon expression of the nucleic acid in cells, cell lines and/or organisms, these cells, cell lines and/or organisms acquire the ability to produce 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula ,

where R is aryl or heteroaryl, from the corresponding 3-arylacrylic acid with the structural formula

.

[0544] The kit can also be used to express in cells and/or cell lines and/or organisms hispidin hydroxylase, hispidin synthase along with tyrosine ammonium lyase and the HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase components. Once the nucleic acid is expressed in cells, cell lines and/or organisms, the cells, cell lines and/or organisms acquire the ability to produce hispidin from tyrosine and cell metabolites.

[0545] Reagent Set #9 includes a nucleic acid encoding the hispidin hydroxylase of the present invention. For example, a hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28 and a nucleic acid encoding a luciferase capable oxidize with the release of light at least one of the fungal luciferins. For example, a luciferase whose amino acid sequence is selected from SEQ ID NOs: 80, 82, 84, 86, 88, 90, 92, 94, 96, 98 can be used.

[0546] The reagent kit may also contain instructions for use of the nucleic acids.

[0547] The reagent kit may also contain deionized water or a buffer to dissolve the lyophilized nucleic acid and/or dilute the nucleic acid solution.

[0548] The reagent kit may also contain primers complementary to regions of the nucleic acids included in the kit to amplify those nucleic acids or fragments thereof.

[0549] The kit can be used for labeling cells and/or cell lines and/or organisms, wherein said cells, cell lines and/or organisms acquire, as a result of the expression of said nucleic acids, the ability to bioluminesce in the presence of exogenous or endogenous fungal preluciferin. For example, they acquire the ability to bioluminescence in the presence of hispidin.

[0550] The kit can also be used to study co-activation of target gene promoters.

[0551] The kit may also include a nucleic acid encoding the hispidin synthase of the present invention, for example, a hispidin synthase, the amino acid sequence of which is selected from the group SEQ ID NO: 35, 37, 39, 41, 43, 45, 47, 49, 51 , 53, 55. In this case, the kit can be used to obtain cells, cell lines and transgenic organisms capable of bioluminescence in the presence of exogenous or endogenous 3-arylacrylic acid with the structural formula

[0552]

where R is aryl or heteroaryl. For example, in the presence of 3-arylacrylic acid selected from the group: caffeic acid, or cinnamic acid, or p-coumaric acid, or coumaric acid, or umbellic acid, or sinapic acid, or ferulic acid. In particular, the kit can be used to obtain autonomously bioluminescent transgenic organisms, for example, plants or fungi.

[0553] The kit may also include a nucleic acid encoding a 4'-phosphopantotheinyl transferase, for example, a 4'-phosphopantotheinyl transferase having the amino acid sequence shown in SEQ ID NO 105 or the like.

[0554] The kit may also contain nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid from cell metabolites.

[0555] The kit may also contain a nucleic acid encoding the caffeoyl pyruvate hydrolase of the present invention, for example, caffeoyl pyruvate hydrolase, the amino acid sequence of which is selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75.

[0556] The kit can also be used for any of the applications described for kits #6-#8.

[0557] The kit can also be used to create cell lines to determine the presence of caffeic acid in a test sample.

[0558] Reagent set #10 includes cells of Agrobacterium tumefaciens strain AGL0 carrying a plasmid containing the coding sequences of hispidin synthase, hispidin hydroxylase, luciferase, phosphopantotheinyl transferase NpgA and an antibiotic resistance gene (for example, kanamycin) under the control of a suitable promoter, for example, the 35S promoter of cauliflower mosaic virus.

[0559] The reagent kit may also include primers to determine whether the expression cassette has been correctly integrated into the cells of dicotyledonous flowering plants.

[0560] The reagent kit can be used to produce autonomously bioluminescent dicotyledonous plants.

[0561] The reagent kit may also include instructions for use of the kit.

[0562] Method of application: Transform a dicotyledonous plant with agrobacterium cells from the kit according to the protocol optimally suited for this type of plant. Plant selection is carried out on a medium with an antibiotic (for example, kanamycin). Correct full-length integration of the expression cassette is performed using PCR with primers from the kit.

[0563] Storage conditions: competent Agrobacterium cells should be stored at a temperature not exceeding -70°C; caffeic acid solution can be stored at temperatures not exceeding -20°C.

[0564] Reagent Kit #11

[0565] The kit includes a purified PKS preparation and a purified hispidin hydroxylase preparation of the present invention and can be used to obtain fungal luciferin from caffeoyl-CoA. The reagent kit may also include a reaction buffer: 0.2 M sodium phosphate buffer (pH 8.0) with the addition of 0.5 M Na ₂ SO4, 0.1% dodecyl maltoside (DDM), 1 mM NADPH, 10 mM ATP, 1 mM Malonyl-CoA or components for preparing the reaction buffer. The reagent kit may also include deionized water. The reagent kit may also include instructions for using the kit.

[0566] Reagent Kit #12

[0567] The kit includes a nucleic acid encoding a PKS and a nucleic acid encoding a hispidin hydroxylase of the present invention. For example, PKS, the amino acid sequence of which is selected from the group SEQ ID NOs: 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139 and hispidin hydroxylase, the amino acid sequence of which is selected from the group SEQ ID NOs: 2, 4, 6, 8 10, 12, 14, 16, 18, 20, 22, 24, 26, 28.

[0568] The reagent kit may also contain instructions for use of the nucleic acids. The reagent kit may also contain deionized water or a buffer for dissolving the lyophilized nucleic acid and/or diluting the nucleic acid solution. The reagent kit may also contain primers complementary to regions of the nucleic acids included in the kit for amplification of these nucleic acids or fragments thereof.

[0569] The reagent kit may also contain a nucleic acid encoding a coumarate CoA ligase, for example, a coumarate CoA ligase having the amino acid sequence shown in SEQ ID NO 141.

[0570] The reagent kit may also contain nucleic acids encoding enzymes for the biosynthesis of caffeic acid from cellular metabolites, for example, encoding tyrosine ammonium lyase and the components HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase. The reagent kit may also contain a nucleic acid encoding the caffeoyl pyruvate hydrolase of the present invention.

[0571] The reagent kit can be used to express hispidin hydroxylase and PKS in cells and/or cell lines and/or organisms. Once the nucleic acid is expressed in cells, cell lines and/or organisms, these cells, cell lines and/or organisms acquire the ability to produce 3-hydroxyhispidin from caffeic acid. The kit can also be used to express in cells and/or cell lines and/or organisms hispidin hydroxylase, PKS together with coumarate CoA ligase, caffeoyl pyruvate hydrolase and/or a combination of tyrosine ammonium lyase and HpaB and HpaC 4- components Hydroxyphenylacetate 3-monooxygenase reductase. Once the nucleic acids are expressed in cells, cell lines and/or organisms, the cells, cell lines and/or organisms acquire the ability to produce 3-hydroxyhispidin from tyrosine and cell metabolites.

[0572] The kit may also contain a nucleic acid encoding a luciferase capable of oxidizing 3-hydroxyhispidin to release light. In this case, the kit can be used for labeling cells and/or cell lines and/or organisms, wherein said cells, cell lines and/or organisms acquire, as a result of the expression of said nucleic acids, the ability to bioluminesce in the presence of exogenous or endogenous hispidin and/or caffeyl-CoA and/or caffeic acid. For example, cells, cell lines and/or organisms acquire the ability to undergo autonomous bioluminescence.

--->

LIST OF SEQUENCES

<110> Limited Liability Company "Planta"

<120> Caffeylpyruvate hydrolases and their applications

<130> 424509

<160> 141

<170> Patent In version 3.5

<210> 1

<211> 1266

<212> DNA

<213> Neonothopanus nambi

<400> 1

atggcatcgt ttgagaattc tctaagcgtt ttgattgtcg gggccggact tggtgggctt 60

gctgctgcca tcgcgctgcg tcgccaaggg catgtcgtga aaatatacga ctcctctagc 120

ttcaaagccg aacttggtgc gggactcgct gtgccgccta acaccttgcg cagtctacag 180

caacttggtt gcaataccga gaacctcaat ggtgtggata atctttgctt cactgcgatg 240

gggtatgacg ggagtgtagg gatgatgaac aacatgactg actatcgaga ggcatacggt 300

acttcttgga tcatggtcca ccgcgttgac ttgcataacg agctgatgcg cgtagcactt 360

gatccaggtg ggctcggacc tcctgcgaca ctccatctta atcatcgtgt cacattctgc 420

gatgtcgacg cttgcaccgt gacattcacc aacgggacca ctcaatcagc tgatctcatc 480

gttggtgcag acggtatacg ctctaccatt cggcggtttg tcttagaaga agacgtgact 540

gtgcctgcgt caggaatcgt cgggtttcga tggcttgtac aagctgacgc gctggaccca 600

tatcctgaac tcgactggat tgttaaaaag cctcctctag gcgcgcgact gatctccact 660

cctcagaatc cacagtctgg tgttggcttg gctgacaggc gcactatcat catctacgca 720

tgtcgtggcg gcaccatggt caatgtcctt gcagtgcatg atgacgaacg tgaccagaac 780

accgcagatt ggagtgtacc ggcttccaaa gacgatctat ttcgtgtttt ccacgattac 840

catccacgct ttcggcggct tttagagctt gcgcaggata ttaatctctg gcaaatgcgt 900

gttgtacctg ttttgaaaaa atgggttaac aagcgggttt gcttgttagg agatgctgcg 960

cacgcttctt taccgacgtt gggtcaaggt tttggtatgg gtctggaaga tgccgtagca 1020

cttggtacac tccttccaaa gggtaccact gcatctcaga tcgagactcg acttgcggtg 1080

tacgaacagc tacgtaagga tcgtgcggaa tttgttgcgg ctgaatcata tgaagagcaa 1140

tatgttcctg aaatgcgggg actttatctg aggtcaaagg aactgcgtga tagagtcatg 1200

ggttatgata tcaaagtgga gagcgagaag gttctcgaga cgctcctaag aagttctaat 1260

tctgcc 1266

<210> 2

<211> 422

<212>PRT

<213> Neonothopanus nambi

<400> 2

Met Ala Ser Phe Glu Asn Ser Leu Ser Val Leu Ile Val Gly Ala Gly

1 5 10 15

Leu Gly Gly Leu Ala Ala Ala Ile Ala Leu Arg Arg Gln Gly His Val

20 25 30

Val Lys Ile Tyr Asp Ser Ser Ser Phe Lys Ala Glu Leu Gly Ala Gly

35 40 45

Leu Ala Val Pro Pro Asn Thr Leu Arg Ser Leu Gln Gln Leu Gly Cys

50 55 60

Asn Thr Glu Asn Leu Asn Gly Val Asp Asn Leu Cys Phe Thr Ala Met

65 70 75 80

Gly Tyr Asp Gly Ser Val Gly Met Met Asn Asn Met Thr Asp Tyr Arg

85 90 95

Glu Ala Tyr Gly Thr Ser Trp Ile Met Val His Arg Val Asp Leu His

100 105 110

Asn Glu Leu Met Arg Val Ala Leu Asp Pro Gly Gly Leu Gly Pro Pro

115 120 125

Ala Thr Leu His Leu Asn His Arg Val Thr Phe Cys Asp Val Asp Ala

130 135 140

Cys Thr Val Thr Phe Thr Asn Gly Thr Thr Gln Ser Ala Asp Leu Ile

145 150 155 160

Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Arg Phe Val Leu Glu

165 170 175

Glu Asp Val Thr Val Pro Ala Ser Gly Ile Val Gly Phe Arg Trp Leu

180 185 190

Val Gln Ala Asp Ala Leu Asp Pro Tyr Pro Glu Leu Asp Trp Ile Val

195 200 205

Lys Lys Pro Pro Leu Gly Ala Arg Leu Ile Ser Thr Pro Gln Asn Pro

210 215 220

Gln Ser Gly Val Gly Leu Ala Asp Arg Arg Thr Ile Ile Ile Tyr Ala

225 230 235 240

Cys Arg Gly Gly Thr Met Val Asn Val Leu Ala Val His Asp Asp Glu

245 250 255

Arg Asp Gln Asn Thr Ala Asp Trp Ser Val Pro Ala Ser Lys Asp Asp

260 265 270

Leu Phe Arg Val Phe His Asp Tyr His Pro Arg Phe Arg Arg Leu Leu

275 280 285

Glu Leu Ala Gln Asp Ile Asn Leu Trp Gln Met Arg Val Val Pro Val

290 295 300

Leu Lys Lys Trp Val Asn Lys Arg Val Cys Leu Leu Gly Asp Ala Ala

305 310 315 320

His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly Leu Glu

325 330 335

Asp Ala Val Ala Leu Gly Thr Leu Leu Pro Lys Gly Thr Thr Ala Ser

340 345 350

Gln Ile Glu Thr Arg Leu Ala Val Tyr Glu Gln Leu Arg Lys Asp Arg

355 360 365

Ala Glu Phe Val Ala Ala Glu Ser Tyr Glu Glu Gln Tyr Val Pro Glu

370 375 380

Met Arg Gly Leu Tyr Leu Arg Ser Lys Glu Leu Arg Asp Arg Val Met

385 390 395 400

Gly Tyr Asp Ile Lys Val Glu Ser Glu Lys Val Leu Glu Thr Leu Leu

405 410 415

Arg Ser Ser Asn Ser Ala

420

<210> 3

<211> 1620

<212> DNA

<213> Omphalotus olearius

<400> 3

atgacgccct ccgagagtcc tttgaatatc tcgattgttg gtgctgggct cggggggctt 60

gctgcagcta ttgcgctgcg tcgtcaaggt catatcatca gaatcttcga ctcgtcaagt 120

tttaaaacgg aactgggtgc tggacttgct gttccaccca atacattacg cagtcttcag 180

gaacttggct gtgatattca gaacttcaat gccgtggaca atctttgttt caccgcgatg 240

ggctacgacg ggagtgtagg gatgatgaac aatatgactg actatcgtga ggcgtatggt 300

gttccctggg tcatggtcca ccgcgttgac ctacataatg aactgcgacg tgtggcactc 360

gatccagatg gccttggacc tcctgcagca ttgcacctca atcatcgtgt gacatcctgc 420

gatgtcgatt cctgcaccgt cacattcgct aacggaaccg ctcatacagc ggatctcatc 480

gttggcgcgg atggtatacg ctcttccatc cgacccttcg tgttgggaga agacgtaatc 540

gtacctgcaa caggaatcgc aggatttcga tggctcatac aagccgaccg gctagatgcg 600

tatcctgaac tcgactggat tgtcaagaac cctcctctcg gcgcgcgatt gatttctgct 660

ccggctcgga aggaacgttc tgtaatcagc gaagcccggc ctgatcgacg tacgattata 720

ttatatgcgt gtcgtggtgg tactattgtc aatgtccttg cggtgcacga cgacgaacgt 780

gatcaggaca ccgtagaatg gagcgtgcca gctaccaaag acgacctatt tcgcgttttc 840

aacgattatc acccaagatt tcggcgactt ctggacctgg cggaggatgt taatctctgg 900

cagatgcgtg ttgtgcctgt tttgagacga tgggttaata aacgggtttg cttgctggga 960

gatgcagcac atgcttcttt accgacattg ggtcaaggtt ttggtatggg tctcgaggat 1020

gcggtggcac ttggtacgct tcttccgagc gggactactg tgtcacagat tgaaatccga 1080

ctttgggtgt atgaaaaact gcgcaaggag cgtgctgaat ttgtttcggc tgaatcgtat 1140

gaagaacact gctctgtgga ttgctataaa tctcataaag cccagtcgac atccaataca 1200

gtgacagaag cagacgacat cttggaagaa ccaaagcctc tgaagccttt atcgtctctg 1260

aaatggccgt acgttcccga agaaccttcg tatcctgatc ccctccaaag aaacgacccc 1320

aaacccctac aactcagaca ctacgaagca atagctacat ctcctgcagt acgggaggtc 1380

ctatcgagtc atccgaatct ccctgcttta ttgacgtcta tcgacaaact gagaggtttt 1440

gatcgcgaac gagctttaga aaaggcgttg gaggttactg cgcctgcgct tgttgatgat 1500

tcaagggctg tagcgctgga ggacgatgta ctcgcaatga gagcattggt agaagcgatt 1560

gaaggtgctg ttaggggcaa taaagaagac gcattaggtc tggattggac tggtagtact 1620

<210> 4

<211> 540

<212>PRT

<213> Omphalotus olearius

<400> 4

Met Thr Pro Ser Glu Ser Pro Leu Asn Ile Ser Ile Val Gly Ala Gly

1 5 10 15

Leu Gly Gly Leu Ala Ala Ala Ile Ala Leu Arg Arg Gln Gly His Ile

20 25 30

Ile Arg Ile Phe Asp Ser Ser Ser Phe Lys Thr Glu Leu Gly Ala Gly

35 40 45

Leu Ala Val Pro Pro Asn Thr Leu Arg Ser Leu Gln Glu Leu Gly Cys

50 55 60

Asp Ile Gln Asn Phe Asn Ala Val Asp Asn Leu Cys Phe Thr Ala Met

65 70 75 80

Gly Tyr Asp Gly Ser Val Gly Met Met Asn Asn Met Thr Asp Tyr Arg

85 90 95

Glu Ala Tyr Gly Val Pro Trp Val Met Val His Arg Val Asp Leu His

100 105 110

Asn Glu Leu Arg Arg Val Ala Leu Asp Pro Asp Gly Leu Gly Pro Pro

115 120 125

Ala Ala Leu His Leu Asn His Arg Val Thr Ser Cys Asp Val Asp Ser

130 135 140

Cys Thr Val Thr Phe Ala Asn Gly Thr Ala His Thr Ala Asp Leu Ile

145 150 155 160

Val Gly Ala Asp Gly Ile Arg Ser Ser Ile Arg Pro Phe Val Leu Gly

165 170 175

Glu Asp Val Ile Val Pro Ala Thr Gly Ile Ala Gly Phe Arg Trp Leu

180 185 190

Ile Gln Ala Asp Arg Leu Asp Ala Tyr Pro Glu Leu Asp Trp Ile Val

195 200 205

Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Ser Ala Pro Ala Arg Lys

210 215 220

Glu Arg Ser Val Ile Ser Glu Ala Arg Pro Asp Arg Arg Thr Ile Ile

225 230 235 240

Leu Tyr Ala Cys Arg Gly Gly Thr Ile Val Asn Val Leu Ala Val His

245 250 255

Asp Asp Glu Arg Asp Gln Asp Thr Val Glu Trp Ser Val Pro Ala Thr

260 265 270

Lys Asp Asp Leu Phe Arg Val Phe Asn Asp Tyr His Pro Arg Phe Arg

275 280 285

Arg Leu Leu Asp Leu Ala Glu Asp Val Asn Leu Trp Gln Met Arg Val

290 295 300

Val Pro Val Leu Arg Arg Trp Val Asn Lys Arg Val Cys Leu Leu Gly

305 310 315 320

Asp Ala Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met

325 330 335

Gly Leu Glu Asp Ala Val Ala Leu Gly Thr Leu Leu Pro Ser Gly Thr

340 345 350

Thr Val Ser Gln Ile Glu Ile Arg Leu Trp Val Tyr Glu Lys Leu Arg

355 360 365

Lys Glu Arg Ala Glu Phe Val Ser Ala Glu Ser Tyr Glu Glu His Cys

370 375 380

Ser Val Asp Cys Tyr Lys Ser His Lys Ala Gln Ser Thr Ser Asn Thr

385 390 395 400

Val Thr Glu Ala Asp Asp Ile Leu Glu Glu Pro Lys Pro Leu Lys Pro

405 410 415

Leu Ser Ser Leu Lys Trp Pro Tyr Val Pro Glu Glu Pro Ser Tyr Pro

420 425 430

Asp Pro Leu Gln Arg Asn Asp Pro Lys Pro Leu Gln Leu Arg His Tyr

435 440 445

Glu Ala Ile Ala Thr Ser Pro Ala Val Arg Glu Val Leu Ser Ser His

450 455 460

Pro Asn Leu Pro Ala Leu Leu Thr Ser Ile Asp Lys Leu Arg Gly Phe

465 470 475 480

Asp Arg Glu Arg Ala Leu Glu Lys Ala Leu Glu Val Thr Ala Pro Ala

485 490 495

Leu Val Asp Asp Ser Arg Ala Val Ala Leu Glu Asp Asp Val Leu Ala

500 505 510

Met Arg Ala Leu Val Glu Ala Ile Glu Gly Ala Val Arg Gly Asn Lys

515 520 525

Glu Asp Ala Leu Gly Leu Asp Trp Thr Gly Ser Thr

530 535 540

<210> 5

<211> 1329

<212> DNA

<213> Guyanagaster necrorhiza

<400> 5

atgcaacaaa tcgacgaagt gtgcccattg aaagtgatcg ttgtaggtgc tggacttggg 60

ggcctttctg ctgccattgc ccttcgtagg caaggccatt gtgtccatat actggaatcg 120

tcaagtttca agagcgaact tggcgcaggt ctcgcagtac cgcccaatac tgtacgctca 180

cttcgaggcc taggctgtaa catcgacaat ctcaagcctg tggataatct ttgtttcact 240

gccatggcgc atgacggaag tcctggtatg atgaataaca tgacggacta tggccaggcg 300

tatggagatc cttgggtaat ggcgcatcgt gttgaccttc acaatgagct catgcgagtg 360

gcccttgaac ccgaaaaaac gggacctcct gcccagcttc gtctggacag ccaggtggca 420

tcttgcaatg tagatgcctg taccgtttct cttgtcgacg gaacaattta ttccgctgat 480

cttatcgttg gtgcagacgg aattaggtct accatacgct cctatgtttt ggacgcagaa 540

atagacatac ctcctaccgg tatcgctgga taccgttggc tcacacctgc agaagctttg 600

gagccatatc ccgaactcga ctggatcatc aagaacccac ccctaggagc acgtttaatc 660

acagctcctg tacgccgaaa cgacagcatt gagcagtcgg gtcctgctcc tatttctgag 720

aaggctgaca agcgtacgat catcatctac gcgtgccgga atggtactat gcttaacgtt 780

ctcggtgtac acgatgaccc tcgcaaccag aacgaagttg gatggaacgt gccagttacc 840

caagaaagtt tgctggattt ttttaaagac tatcatcccc gattcaagcg tctgcttgag 900

ctggctgaca atgttcatct gtggcaaatg cgcgtcgtcc cgcggcttga gacttggatc 960

aataaacgcg tgtgtttgtt gggcgattct gcgcatgcat cattaccaac actcggtcaa 1020

ggcttcggga tgggacttga ggatgccgta gcccttgcaa ccctccttcc gatgggaacc 1080

aaagtgtctg acatcgagaa ccgcctcgtc gcctacgaaa gcttgcgtaa ggggcgcgct 1140

gagtatgtgg ccatggaatc gttcgaacaa cagaatatcc cggaaaagcg aggcttgtat 1200

ctcaggtctt ctatgatgcg tgacgaaatc atgggttatg atgtcaaagc cgaggctgag 1260

aaggttttta aagaattaat gacctcgact gataaggtta cataccgtcc ccatgtggac 1320

tgtctatgg 1329

<210> 6

<211> 443

<212>PRT

<213> Guyanagaster necrorhiza

<400> 6

Met Gln Gln Ile Asp Glu Val Cys Pro Leu Lys Val Ile Val Val Gly

1 5 10 15

Ala Gly Leu Gly Gly Leu Ser Ala Ala Ile Ala Leu Arg Arg Gln Gly

20 25 30

His Cys Val His Ile Leu Glu Ser Ser Ser Phe Lys Ser Glu Leu Gly

35 40 45

Ala Gly Leu Ala Val Pro Pro Asn Thr Val Arg Ser Leu Arg Gly Leu

50 55 60

Gly Cys Asn Ile Asp Asn Leu Lys Pro Val Asp Asn Leu Cys Phe Thr

65 70 75 80

Ala Met Ala His Asp Gly Ser Pro Gly Met Met Asn Asn Met Thr Asp

85 90 95

Tyr Gly Gln Ala Tyr Gly Asp Pro Trp Val Met Ala His Arg Val Asp

100 105 110

Leu His Asn Glu Leu Met Arg Val Ala Leu Glu Pro Glu Lys Thr Gly

115 120 125

Pro Pro Ala Gln Leu Arg Leu Asp Ser Gln Val Ala Ser Cys Asn Val

130 135 140

Asp Ala Cys Thr Val Ser Leu Val Asp Gly Thr Ile Tyr Ser Ala Asp

145 150 155 160

Leu Ile Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Ser Tyr Val

165 170 175

Leu Asp Ala Glu Ile Asp Ile Pro Pro Thr Gly Ile Ala Gly Tyr Arg

180 185 190

Trp Leu Thr Pro Ala Glu Ala Leu Glu Pro Tyr Pro Glu Leu Asp Trp

195 200 205

Ile Ile Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Thr Ala Pro Val

210 215 220

Arg Arg Asn Asp Ser Ile Glu Gln Ser Gly Pro Ala Pro Ile Ser Glu

225 230 235 240

Lys Ala Asp Lys Arg Thr Ile Ile Ile Tyr Ala Cys Arg Asn Gly Thr

245 250 255

Met Leu Asn Val Leu Gly Val His Asp Asp Pro Arg Asn Gln Asn Glu

260 265 270

Val Gly Trp Asn Val Pro Val Thr Gln Glu Ser Leu Leu Asp Phe Phe

275 280 285

Lys Asp Tyr His Pro Arg Phe Lys Arg Leu Leu Glu Leu Ala Asp Asn

290 295 300

Val His Leu Trp Gln Met Arg Val Val Pro Arg Leu Glu Thr Trp Ile

305 310 315 320

Asn Lys Arg Val Cys Leu Leu Gly Asp Ser Ala His Ala Ser Leu Pro

325 330 335

Thr Leu Gly Gln Gly Phe Gly Met Gly Leu Glu Asp Ala Val Ala Leu

340 345 350

Ala Thr Leu Leu Pro Met Gly Thr Lys Val Ser Asp Ile Glu Asn Arg

355 360 365

Leu Val Ala Tyr Glu Ser Leu Arg Lys Gly Arg Ala Glu Tyr Val Ala

370 375 380

Met Glu Ser Phe Glu Gln Gln Asn Ile Pro Glu Lys Arg Gly Leu Tyr

385 390 395 400

Leu Arg Ser Ser Met Met Arg Asp Glu Ile Met Gly Tyr Asp Val Lys

405 410 415

Ala Glu Ala Glu Lys Val Phe Lys Glu Leu Met Thr Ser Thr Asp Lys

420 425 430

Val Thr Tyr Arg Pro His Val Asp Cys Leu Trp

435 440

<210> 7

<211> 1284

<212> DNA

<213> Panellus stipticus

<400> 7

atgtcacaag gcactgaaga ctcgccctca ctcttcgcaa tagttgtcgg tgctggtctg 60

gtcggctttg ccgctgccgt cgcgctgcgc cgtcaaggtc accgtgtcac gatctacgaa 120

tcttcaagct ttaagacaga acttggggcg ggcctcgcga tcccttcaaa cacattgcga 180

tgcttggttg gccttggatg tactgtcgcc aacatggatc ccgtcaataa tctttgtttt 240

acatcgatgg catacgatgg taccgcgggg atgaaaagcg acagctccga ctacgaggcg 300

cagtatggca ctccctggat catggcccac cgcgtcgatc tgcacaagga gcttcgtcgc 360

ctggcagtgg atcccgaggg caccggtccc cccgcagaac tgcaccttag ccaccgggtt 420

gtctcctgcg acgtcgaatc cggctccgtc acgctcctgg acggctccgt tcagtcagca 480

gacttgataa ttggggctga tggaattcgt tcaaccgttc gcaaatttgt cgtaggcgaa 540

gaaatagcaa tccccccatc tcgtacggcg ggcttccgct ggctcacaca agcaagcgct 600

cttgacccct accccgaact ggattggatc gtgaaaacgc cgccgttggg tgctcgggta 660

atctccgctc cgattcaacc tccagaagtc acccacatcg accaccgtac gatcgtcatc 720

tatgcctgtc gcggaggtcg tctcgtaaat gttctaggaa tacatgagga tctgcgcgac 780

caggattctg tcgactggaa cgtccccata acgcgagaag cgctgctcca ctttttcgga 840

gactaccacc cacggttcaa gcggctcttg gagctcgcgg aggacgtaca cgtctggcag 900

atgcgcgtgg tccccccact gccgacatgg gtgaacggcc gcgtatgcat catgggcgac 960

gcagcgcatg catcgcttcc cacactggga caagggtttg gcatgggcct cgaagacgcg 1020

gtcgcgctcg ggacgctgct ttctagttcg acgccgtcaa gcgacattcc aagccgtctc 1080

gtcgcgtacg agaagcttcg caaagcgcgc gctgagtatg tttccaaaga gtcgtacgaa 1140

cagcagcatg tccgagaaaa gagagggctg tatctccggt cccgagaaat gcgggatgtg 1200

atcatgggat acgacgtgaa gaaggaggca gaacggatct tgagcgagat cagcattgca 1260

caagaacagt gtgctgttca tgat 1284

<210> 8

<211> 428

<212>PRT

<213> Panellus stipticus

<400> 8

Met Ser Gln Gly Thr Glu Asp Ser Pro Ser Leu Phe Ala Ile Val Val

1 5 10 15

Gly Ala Gly Leu Val Gly Phe Ala Ala Ala Val Ala Leu Arg Arg Gln

20 25 30

Gly His Arg Val Thr Ile Tyr Glu Ser Ser Ser Phe Lys Thr Glu Leu

35 40 45

Gly Ala Gly Leu Ala Ile Pro Ser Asn Thr Leu Arg Cys Leu Val Gly

50 55 60

Leu Gly Cys Thr Val Ala Asn Met Asp Pro Val Asn Asn Leu Cys Phe

65 70 75 80

Thr Ser Met Ala Tyr Asp Gly Thr Ala Gly Met Lys Ser Asp Ser Ser

85 90 95

Asp Tyr Glu Ala Gln Tyr Gly Thr Pro Trp Ile Met Ala His Arg Val

100 105 110

Asp Leu His Lys Glu Leu Arg Arg Leu Ala Val Asp Pro Glu Gly Thr

115 120 125

Gly Pro Pro Ala Glu Leu His Leu Ser His Arg Val Val Ser Cys Asp

130 135 140

Val Glu Ser Gly Ser Val Thr Leu Leu Asp Gly Ser Val Gln Ser Ala

145 150 155 160

Asp Leu Ile Ile Gly Ala Asp Gly Ile Arg Ser Thr Val Arg Lys Phe

165 170 175

Val Val Gly Glu Glu Ile Ala Ile Pro Pro Ser Arg Thr Ala Gly Phe

180 185 190

Arg Trp Leu Thr Gln Ala Ser Ala Leu Asp Pro Tyr Pro Glu Leu Asp

195 200 205

Trp Ile Val Lys Thr Pro Pro Leu Gly Ala Arg Val Ile Ser Ala Pro

210 215 220

Ile Gln Pro Pro Glu Val Thr His Ile Asp His Arg Thr Ile Val Ile

225 230 235 240

Tyr Ala Cys Arg Gly Gly Arg Leu Val Asn Val Leu Gly Ile His Glu

245 250 255

Asp Leu Arg Asp Gln Asp Ser Val Asp Trp Asn Val Pro Ile Thr Arg

260 265 270

Glu Ala Leu Leu His Phe Phe Gly Asp Tyr His Pro Arg Phe Lys Arg

275 280 285

Leu Leu Glu Leu Ala Glu Asp Val His Val Trp Gln Met Arg Val Val

290 295 300

Pro Pro Leu Pro Thr Trp Val Asn Gly Arg Val Cys Ile Met Gly Asp

305 310 315 320

Ala Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly

325 330 335

Leu Glu Asp Ala Val Ala Leu Gly Thr Leu Leu Ser Ser Ser Thr Pro

340 345 350

Ser Ser Asp Ile Pro Ser Arg Leu Val Ala Tyr Glu Lys Leu Arg Lys

355 360 365

Ala Arg Ala Glu Tyr Val Ser Lys Glu Ser Tyr Glu Gln Gln His Val

370 375 380

Arg Glu Lys Arg Gly Leu Tyr Leu Arg Ser Arg Glu Met Arg Asp Val

385 390 395 400

Ile Met Gly Tyr Asp Val Lys Lys Glu Ala Glu Arg Ile Leu Ser Glu

405 410 415

Ile Ser Ile Ala Gln Glu Gln Cys Ala Val His Asp

420 425

<210> 9

<211> 1284

<212> DNA

<213> Panellus stipticus

<400> 9

atgtcacaag gcactgaaga ctcgccctca ctcttcgcaa tagttgtcgg tgctggtctg 60

gtcggctttg ccgctgccgt cgcgctgcgc cgtcaaggtc accgtgtcac gatctacgaa 120

tcttcaagct ttaagacaga acttggggcg ggcctcgcga tcccttcaaa cacattgcga 180

tgcttggttg gccttggatg tactgtcgcc aacatggatc ccgtcaataa tctttgtttt 240

acatcgatgg catacgatgg taccgcgggg atgaaaagcg acagctccga ctacgaggcg 300

cagtatggca ctccctggat catggcccac cgcgtcgatc tgcacaagga gcttcgtcgc 360

ctggcagtgg atcccgaggg caccggtccc cccgcagaac tgcaccttag ccaccgggtt 420

gtctcctgcg acgtcgaatc cggctccgtc acgctcctgg acggctccgt tcagtcagca 480

gacttgataa ttggggctga tggaattcgt tcaaccgttc gcaaatttgt cgtaggcgaa 540

gaaatagcaa tccccccatc tcgtacggcg ggcttccgct ggctcacaca agcaagcgct 600

cttgacccct accccgaact ggattggatc gtgaaaacgc cgccgttggg tgctcgggta 660

atctccgctc cgattcaacc tccagaagtc acccacatcg accaccgtac gatcgtcatc 720

tatgcctgtc gcggaggtcg tctcgtaaat gttctaggaa tacatgagga tctgcgcgac 780

caggattctg tcgactggaa cgtccccata acgcgagaag cgctgctcca ctttttcgga 840

gactaccacc cacggttcaa gcggctcttg gagctcgcgg aggacgtaca cgtctggcag 900

atgcgcgtgg tccccccact gccgacatgg gtgaacggcc gcgtatgcat catgggcgac 960

gcagcgcatg catcgcttcc cacactggga caagggtttg gcatgggcct cgaagacgcg 1020

gtcgcgctcg ggacgctgct tcctagttcg acgccgtcaa gcgacattcc aagccgtctc 1080

gtcgcgtacg agaagcttcg caaagcgcgc gctgagtatg tttccaaaga gtcgtacgaa 1140

cagcagcatg tccgagaaaa gagagggctg tatctccggt cccgagaaat gcgggatgtg 1200

atcatgggat acgacgtgaa gaaggaggca gaacggatct tgagcgagat cagcattgca 1260

caagaacagt gtgctgttca tgat 1284

<210> 10

<211> 428

<212>PRT

<213> Panellus stipticus

<400> 10

Met Ser Gln Gly Thr Glu Asp Ser Pro Ser Leu Phe Ala Ile Val Val

1 5 10 15

Gly Ala Gly Leu Val Gly Phe Ala Ala Ala Val Ala Leu Arg Arg Gln

20 25 30

Gly His Arg Val Thr Ile Tyr Glu Ser Ser Ser Phe Lys Thr Glu Leu

35 40 45

Gly Ala Gly Leu Ala Ile Pro Ser Asn Thr Leu Arg Cys Leu Val Gly

50 55 60

Leu Gly Cys Thr Val Ala Asn Met Asp Pro Val Asn Asn Leu Cys Phe

65 70 75 80

Thr Ser Met Ala Tyr Asp Gly Thr Ala Gly Met Lys Ser Asp Ser Ser

85 90 95

Asp Tyr Glu Ala Gln Tyr Gly Thr Pro Trp Ile Met Ala His Arg Val

100 105 110

Asp Leu His Lys Glu Leu Arg Arg Leu Ala Val Asp Pro Glu Gly Thr

115 120 125

Gly Pro Pro Ala Glu Leu His Leu Ser His Arg Val Val Ser Cys Asp

130 135 140

Val Glu Ser Gly Ser Val Thr Leu Leu Asp Gly Ser Val Gln Ser Ala

145 150 155 160

Asp Leu Ile Ile Gly Ala Asp Gly Ile Arg Ser Thr Val Arg Lys Phe

165 170 175

Val Val Gly Glu Glu Ile Ala Ile Pro Pro Ser Arg Thr Ala Gly Phe

180 185 190

Arg Trp Leu Thr Gln Ala Ser Ala Leu Asp Pro Tyr Pro Glu Leu Asp

195 200 205

Trp Ile Val Lys Thr Pro Pro Leu Gly Ala Arg Val Ile Ser Ala Pro

210 215 220

Ile Gln Pro Pro Glu Val Thr His Ile Asp His Arg Thr Ile Val Ile

225 230 235 240

Tyr Ala Cys Arg Gly Gly Arg Leu Val Asn Val Leu Gly Ile His Glu

245 250 255

Asp Leu Arg Asp Gln Asp Ser Val Asp Trp Asn Val Pro Ile Thr Arg

260 265 270

Glu Ala Leu Leu His Phe Phe Gly Asp Tyr His Pro Arg Phe Lys Arg

275 280 285

Leu Leu Glu Leu Ala Glu Asp Val His Val Trp Gln Met Arg Val Val

290 295 300

Pro Pro Leu Pro Thr Trp Val Asn Gly Arg Val Cys Ile Met Gly Asp

305 310 315 320

Ala Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly

325 330 335

Leu Glu Asp Ala Val Ala Leu Gly Thr Leu Leu Pro Ser Ser Thr Pro

340 345 350

Ser Ser Asp Ile Pro Ser Arg Leu Val Ala Tyr Glu Lys Leu Arg Lys

355 360 365

Ala Arg Ala Glu Tyr Val Ser Lys Glu Ser Tyr Glu Gln Gln His Val

370 375 380

Arg Glu Lys Arg Gly Leu Tyr Leu Arg Ser Arg Glu Met Arg Asp Val

385 390 395 400

Ile Met Gly Tyr Asp Val Lys Lys Glu Ala Glu Arg Ile Leu Ser Glu

405 410 415

Ile Ser Ile Ala Gln Glu Gln Cys Ala Val His Asp

420 425

<210> 11

<211> 1284

<212> DNA

<213> Panellus stipticus

<400> 11

atgtcacaag gcactgaaga ctcgccctca ctcttcgcaa tagttgtcgg tgctggtctg 60

gtcggctttg ccgctgccgt cgcgctgcgc cgtcaaggtc accgtgtcac tatctacgaa 120

tcttcaagct ttaagacaga acttggggcg ggcctcgcga tcccttcaaa cacattgcga 180

tgcttggttg gccttggatg tactgtcgcc aatctggatc ccgtcaataa tctttgtttt 240

acatcgatgg cttacgatgg taccgcgggg atgaaaagcg acagctccga ctacgaggcg 300

cagtatggca ctccctggat catggcccac cgcgtcgatc tgcacaagga gcttcgtcgc 360

ctggcagtgg atcccgaggg caccggtccc cccgcagaac tgcaccttag ccaccgggtt 420

gtctcctgcg acgtcgaatc cggctccgtc acgctcctgg acggctccat tcagtcagca 480

gacttgataa ttggggctga tggaattcgt tcaaccgttc gcaaatttgt cgtaggcgaa 540

gaaatagcaa tccccccatc tcgtacggcg ggcttccgtt ggctcacaca agcaagcgct 600

cttgacccct accccgaact ggattggatc gtgaaaacgc cgccgttggg tgctcgggta 660

atctccgctc cgattcaacc tccagaagtc acccacatcg accaccgtac gatcgtcatc 720

tatgcctgtc gcggaggtcg tctcgtaaat gttctaggaa tacatgagga tctgcgcgac 780

caggattctg tcgactggaa cgtccccata acgcgagaag cgctgctcca ctttttcgga 840

gactaccacc cacggttcaa gcggctcttg gagctcgcgg aggacgtaca cgtctggcag 900

atgcgcgtgg tccccccact gccgacatgg gtgaacggcc gcgtatgcat catgggcgac 960

gcagcgcatg catcgcttcc cacactggga caagggtttg gcatgggcct cgaagacgcg 1020

gtcgcgctcg ggacgctgct tcctagttcg acgccgtcag gcgacattcc aagccgtctc 1080

gtcgcgtacg agaagcttcg caaagcgcgc gctgagtatg tttccaaaga gtcgtacgaa 1140

cagcagcatg tccgagaaaa gagagggctg tatctccggt cccgagaaat gcgggatgtg 1200

atcatgggat acgacgtgaa gaaggaggca gaacggatct ttagcgagat cagcattgca 1260

caagaacagc gtgctgttca tgat 1284

<210> 12

<211> 428

<212>PRT

<213> Panellus stipticus

<400> 12

Met Ser Gln Gly Thr Glu Asp Ser Pro Ser Leu Phe Ala Ile Val Val

1 5 10 15

Gly Ala Gly Leu Val Gly Phe Ala Ala Ala Val Ala Leu Arg Arg Gln

20 25 30

Gly His Arg Val Thr Ile Tyr Glu Ser Ser Ser Phe Lys Thr Glu Leu

35 40 45

Gly Ala Gly Leu Ala Ile Pro Ser Asn Thr Leu Arg Cys Leu Val Gly

50 55 60

Leu Gly Cys Thr Val Ala Asn Leu Asp Pro Val Asn Asn Leu Cys Phe

65 70 75 80

Thr Ser Met Ala Tyr Asp Gly Thr Ala Gly Met Lys Ser Asp Ser Ser

85 90 95

Asp Tyr Glu Ala Gln Tyr Gly Thr Pro Trp Ile Met Ala His Arg Val

100 105 110

Asp Leu His Lys Glu Leu Arg Arg Leu Ala Val Asp Pro Glu Gly Thr

115 120 125

Gly Pro Pro Ala Glu Leu His Leu Ser His Arg Val Val Ser Cys Asp

130 135 140

Val Glu Ser Gly Ser Val Thr Leu Leu Asp Gly Ser Ile Gln Ser Ala

145 150 155 160

Asp Leu Ile Ile Gly Ala Asp Gly Ile Arg Ser Thr Val Arg Lys Phe

165 170 175

Val Val Gly Glu Glu Ile Ala Ile Pro Pro Ser Arg Thr Ala Gly Phe

180 185 190

Arg Trp Leu Thr Gln Ala Ser Ala Leu Asp Pro Tyr Pro Glu Leu Asp

195 200 205

Trp Ile Val Lys Thr Pro Pro Leu Gly Ala Arg Val Ile Ser Ala Pro

210 215 220

Ile Gln Pro Pro Glu Val Thr His Ile Asp His Arg Thr Ile Val Ile

225 230 235 240

Tyr Ala Cys Arg Gly Gly Arg Leu Val Asn Val Leu Gly Ile His Glu

245 250 255

Asp Leu Arg Asp Gln Asp Ser Val Asp Trp Asn Val Pro Ile Thr Arg

260 265 270

Glu Ala Leu Leu His Phe Phe Gly Asp Tyr His Pro Arg Phe Lys Arg

275 280 285

Leu Leu Glu Leu Ala Glu Asp Val His Val Trp Gln Met Arg Val Val

290 295 300

Pro Pro Leu Pro Thr Trp Val Asn Gly Arg Val Cys Ile Met Gly Asp

305 310 315 320

Ala Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly

325 330 335

Leu Glu Asp Ala Val Ala Leu Gly Thr Leu Leu Pro Ser Ser Thr Pro

340 345 350

Ser Gly Asp Ile Pro Ser Arg Leu Val Ala Tyr Glu Lys Leu Arg Lys

355 360 365

Ala Arg Ala Glu Tyr Val Ser Lys Glu Ser Tyr Glu Gln Gln His Val

370 375 380

Arg Glu Lys Arg Gly Leu Tyr Leu Arg Ser Arg Glu Met Arg Asp Val

385 390 395 400

Ile Met Gly Tyr Asp Val Lys Lys Glu Ala Glu Arg Ile Phe Ser Glu

405 410 415

Ile Ser Ile Ala Gln Glu Gln Arg Ala Val His Asp

420 425

<210> 13

<211> 1248

<212> DNA

<213> Neonothopanus gardneri

<400> 13

atggcactat ctgagagtcc tttgaacgtc ttgatagtag gagcggggct cggggggctt 60

gctgctgcca tagcactacg tcgtcaaggg catatcgtga aaatattcga ttcctccagt 120

ttcaaaaccg aacttggtgc aggacttgct gtcccgccta ataccctgcg tagtctgcag 180

gaactcgggt gcagtgtcga gaacctcaat gctgtggata atctttgctt cactgcgatg 240

gggtatgacg ggagtgtagg aatgatgaac aatatgaccg actatcgaga ggcgtacggt 300

catccttggg tcatggttca ccgtgtcgac ttgcataatg agctgaagcg cgtggcgctt 360

gatccagacg gcctcggacc tcctgcaact ttgcatctca accatcgtgt cacattctgc 420

gacatcgact cttgcactgt cacgttcgct aatgggactt ctaaatcggc agatcttatc 480

gtaggcgcag acggtatacg ctctaccatt cgcaagttca ttcttggaga agacgtcgtt 540

atacccgcgt caggaatagc agggtttcga tggcttgtgc aagctgacgc gctggatccg 600

tatcctgaac tcgactggat cgtgaagaac cctcctctag gagcccgact gatttccgct 660

cctaggaatc aacagtctac tgataggcgc actatcatca tctatgcgtg tcgtagcggc 720

accatggtca acgtactcgc agtacatgat gatgatcgtg accagaacgc cgtagattgg 780

agtgcaccag cttccaaaga tgatttattc cacatcttcc acgactacca cccacgattc 840

cagcggcttc tggagctggc gcaagatatc aatctctggc aaatgcgtgt tgttcctgtt 900

ctgaaacaat gggttaacaa acgtgtttgc ttgttaggag atgcggcaca cgcttcttta 960

cctacattag ggcagggatt tggtatgggt ctagaagatg ccgtagcact gggtacgctt 1020

cttccaaagg gggccacagt atctcagatc gagagccgac tcgcggtgta tgaaattctg 1080

cgcaaggagc gtgctgaatt tgtttcggct gagtcatatg aagagcagta cgttccagaa 1140

aaacgcgggc tttacttaag atcgaaggaa ttgcgcgaca gtgtcatggg ttacgatatt 1200

aaaatggaga gcgagaaggt tctcgtggca ttacttagcg gttcttca 1248

<210> 14

<211> 416

<212>PRT

<213> Neonothopanus gardneri

<400> 14

Met Ala Leu Ser Glu Ser Pro Leu Asn Val Leu Ile Val Gly Ala Gly

1 5 10 15

Leu Gly Gly Leu Ala Ala Ala Ile Ala Leu Arg Arg Gln Gly His Ile

20 25 30

Val Lys Ile Phe Asp Ser Ser Ser Phe Lys Thr Glu Leu Gly Ala Gly

35 40 45

Leu Ala Val Pro Pro Asn Thr Leu Arg Ser Leu Gln Glu Leu Gly Cys

50 55 60

Ser Val Glu Asn Leu Asn Ala Val Asp Asn Leu Cys Phe Thr Ala Met

65 70 75 80

Gly Tyr Asp Gly Ser Val Gly Met Met Asn Asn Met Thr Asp Tyr Arg

85 90 95

Glu Ala Tyr Gly His Pro Trp Val Met Val His Arg Val Asp Leu His

100 105 110

Asn Glu Leu Lys Arg Val Ala Leu Asp Pro Asp Gly Leu Gly Pro Pro

115 120 125

Ala Thr Leu His Leu Asn His Arg Val Thr Phe Cys Asp Ile Asp Ser

130 135 140

Cys Thr Val Thr Phe Ala Asn Gly Thr Ser Lys Ser Ala Asp Leu Ile

145 150 155 160

Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Lys Phe Ile Leu Gly

165 170 175

Glu Asp Val Val Ile Pro Ala Ser Gly Ile Ala Gly Phe Arg Trp Leu

180 185 190

Val Gln Ala Asp Ala Leu Asp Pro Tyr Pro Glu Leu Asp Trp Ile Val

195 200 205

Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Ser Ala Pro Arg Asn Gln

210 215 220

Gln Ser Thr Asp Arg Arg Thr Ile Ile Ile Tyr Ala Cys Arg Ser Gly

225 230 235 240

Thr Met Val Asn Val Leu Ala Val His Asp Asp Asp Arg Asp Gln Asn

245 250 255

Ala Val Asp Trp Ser Ala Pro Ala Ser Lys Asp Asp Leu Phe His Ile

260 265 270

Phe His Asp Tyr His Pro Arg Phe Gln Arg Leu Leu Glu Leu Ala Gln

275 280 285

Asp Ile Asn Leu Trp Gln Met Arg Val Val Pro Val Leu Lys Gln Trp

290 295 300

Val Asn Lys Arg Val Cys Leu Leu Gly Asp Ala Ala His Ala Ser Leu

305 310 315 320

Pro Thr Leu Gly Gln Gly Phe Gly Met Gly Leu Glu Asp Ala Val Ala

325 330 335

Leu Gly Thr Leu Leu Pro Lys Gly Ala Thr Val Ser Gln Ile Glu Ser

340 345 350

Arg Leu Ala Val Tyr Glu Ile Leu Arg Lys Glu Arg Ala Glu Phe Val

355 360 365

Ser Ala Glu Ser Tyr Glu Glu Gln Tyr Val Pro Glu Lys Arg Gly Leu

370 375 380

Tyr Leu Arg Ser Lys Glu Leu Arg Asp Ser Val Met Gly Tyr Asp Ile

385 390 395 400

Lys Met Glu Ser Glu Lys Val Leu Val Ala Leu Leu Ser Gly Ser Ser

405 410 415

<210> 15

<211> 1257

<212> DNA

<213> Mycena citricolor

<400> 15

atgaacaccc ccaataacgc tctcgatgtt attgttgtcg gtgctggcct ggttggtttc 60

gcggccgctg ctgctctacg ccgacaaggt catcgcgtga ctatctacga gacttccagc 120

ttcaagaacg agctaggagc tggccttgct attccaccca acactgtccg tggcctaatt 180

ggcttgggat gtgtgattga gaacttggac ccggtggaga atctatgcgt ttccgtcgct 240

tttgacggaa gtgctggtat gcgcagtgac cagaccaact acgaagcaag ctacggcctt 300

ccctggatca tggtgcatcg cgtcgatttg cacaatgagc ttcgtcgggt tgctctcagt 360

gccgagggga acattggtcc cccagccgag ctacgcctgg accaccgagt cagctcgtgt 420

gatgtcgaga aatgcactgt gacgctgagc aatggcgata cccaccacgc ggatctgatc 480

attggagcag acgggatcca ttctacaatc cgatccttcg tcgtgggcga ggaaatcgtc 540

attccgccct ccaagacagc cggtttccgc tggctcacag agagtactgc gttggagccc 600

tatccggaat tggactggat tgtgaagatc ccaccacttg gcgcccggct gatctctgcg 660

ccaatgaacc ctgcgccacc gcaggtcgac caccggacga tcatcatcta cgcctgtcgt 720

ggcagtacac tgataaatgt actcggagtc catgaggatc tccgcgatca agatacagtt 780

ccctggaatg cacccgtaac ccaatcggag ctgctccagt tctttggcga ttaccatccg 840

cgattcaaac gattgttgga gcttgcaaat gatgttcatg tgtggcagat gcgagtagtg 900

ccccgcttgg agacctgggt caatcgtcgg gtttgcatta tgggcgatgc tgcgcatgca 960

tcactcccca cgctgggtca aggtttcgga atggggctcg aggatgcagt cgctcttgga 1020

acactccttc cgcttgggac aactcccgaa gagatcccgg accgtctcac cctctggcag 1080

gatctcgtca aacctcgggc tgagtttgtc gcgactgaat cctacgaaca gcagcatatt 1140

cctgcgaaac ggggactcta tcttcgctcg caggagatgc gcgactgggt catgggatac 1200

gatgtccagg ctgaggcaca gaaggtcttg gcgggagctg tgaatagatc caaggga 1257

<210> 16

<211> 419

<212>PRT

<213> Mycena citricolor

<400> 16

Met Asn Thr Pro Asn Asn Ala Leu Asp Val Ile Val Val Gly Ala Gly

1 5 10 15

Leu Val Gly Phe Ala Ala Ala Ala Ala Leu Arg Arg Gln Gly His Arg

20 25 30

Val Thr Ile Tyr Glu Thr Ser Ser Phe Lys Asn Glu Leu Gly Ala Gly

35 40 45

Leu Ala Ile Pro Pro Asn Thr Val Arg Gly Leu Ile Gly Leu Gly Cys

50 55 60

Val Ile Glu Asn Leu Asp Pro Val Glu Asn Leu Cys Val Ser Val Ala

65 70 75 80

Phe Asp Gly Ser Ala Gly Met Arg Ser Asp Gln Thr Asn Tyr Glu Ala

85 90 95

Ser Tyr Gly Leu Pro Trp Ile Met Val His Arg Val Asp Leu His Asn

100 105 110

Glu Leu Arg Arg Val Ala Leu Ser Ala Glu Gly Asn Ile Gly Pro Pro

115 120 125

Ala Glu Leu Arg Leu Asp His Arg Val Ser Ser Cys Asp Val Glu Lys

130 135 140

Cys Thr Val Thr Leu Ser Asn Gly Asp Thr His His Ala Asp Leu Ile

145 150 155 160

Ile Gly Ala Asp Gly Ile His Ser Thr Ile Arg Ser Phe Val Val Gly

165 170 175

Glu Glu Ile Val Ile Pro Pro Ser Lys Thr Ala Gly Phe Arg Trp Leu

180 185 190

Thr Glu Ser Thr Ala Leu Glu Pro Tyr Pro Glu Leu Asp Trp Ile Val

195 200 205

Lys Ile Pro Pro Leu Gly Ala Arg Leu Ile Ser Ala Pro Met Asn Pro

210 215 220

Ala Pro Pro Gln Val Asp His Arg Thr Ile Ile Ile Tyr Ala Cys Arg

225 230 235 240

Gly Ser Thr Leu Ile Asn Val Leu Gly Val His Glu Asp Leu Arg Asp

245 250 255

Gln Asp Thr Val Pro Trp Asn Ala Pro Val Thr Gln Ser Glu Leu Leu

260 265 270

Gln Phe Phe Gly Asp Tyr His Pro Arg Phe Lys Arg Leu Leu Glu Leu

275 280 285

Ala Asn Asp Val His Val Trp Gln Met Arg Val Val Pro Arg Leu Glu

290 295 300

Thr Trp Val Asn Arg Arg Val Cys Ile Met Gly Asp Ala Ala His Ala

305 310 315 320

Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly Leu Glu Asp Ala

325 330 335

Val Ala Leu Gly Thr Leu Leu Pro Leu Gly Thr Thr Pro Glu Glu Ile

340 345 350

Pro Asp Arg Leu Thr Leu Trp Gln Asp Leu Val Lys Pro Arg Ala Glu

355 360 365

Phe Val Ala Thr Glu Ser Tyr Glu Gln Gln His Ile Pro Ala Lys Arg

370 375 380

Gly Leu Tyr Leu Arg Ser Gln Glu Met Arg Asp Trp Val Met Gly Tyr

385 390 395 400

Asp Val Gln Ala Glu Ala Gln Lys Val Leu Ala Gly Ala Val Asn Arg

405 410 415

Ser Lys Gly

<210> 17

<211> 1260

<212> DNA

<213> Mycena citricolor

<400> 17

atgaacaccc ccaataacgc tctcgatgtt attgttgtcg gtgctggcct ggttggtttc 60

gcggccgctg ctgctctacg ccgacaaggt catcgcgtga ctatctatga gacttccagc 120

ttcaagaacg agttaggac tggcctggct attccaccca acactgtccg tggtctaatt 180

ggcttgggat gtgtgattga gaacttggac ccggtggaga atctatgctt cactgccgtc 240

gcgtttgacg gaagtgctgg tatgcgcagc gaccaaacca actacgaagc aagttacggc 300

cttccctgga tcatggtgca tcgtgtcgat ttgcacaacg agcttcgtcg ggttgctctc 360

agcgccgagg ggaacactgg tcccccagcg gagctacgcc tggaccaccg agtcagctcg 420

tgtgatgtcg agaaatgcac tgtgacgctg agcaatgggg atacccacca cgcagatctg 480

atcattggag cagacgggat ccattctaca atccgatcct tcgttgtggg cgaggaaatc 540

gtcattccgc cctccaagac agccggtttc cgctggctca cagagagtac tgcgttggag 600

ccctatccgg aattggactg gattgtgaag atcccaccac ttggcgcccg gctgatctct 660

gcgccaatga accctgcgcc accgcaggtc gaccaccgga cgatcatcat ctacgcctgt 720

cgtggcagta cactgataaa tgtactcgga gtccatgagg atctccgcga tcaagataca 780

gtcccctgga atgcacccgt aacccaatcg gagctgctcc agttctttgg cgattaccat 840

ccgcggttca aacgattgtt ggagcttgca aatgatgttc atgtgtggca gatgcgagta 900

gtgccccgct tggagacctg ggtcaatcgt cgggtttgca ttatgggcga tgctgcgcat 960

gcatcactcc ccacgctggg tcaaggtttc ggaatggggc tcgaggatgc agtcgctctt 1020

ggaacactcc ttccgcttgg gacaactccc gaagagatcc cggaccgtct caccctctgg 1080

caggatctcg tcaaacctcg ggctgagttt gtcgcgactg aatcctacga acagcagcat 1140

attcctgcga aacggggact ctatcttcgc tcgcaggaga tgcgcgactg ggtcatggga 1200

tacgatgtcc aggctgaggc acagaaggtc ttggcgggag ctgtgaatag atccaaggga 1260

<210> 18

<211> 420

<212>PRT

<213> Mycena citricolor

<400> 18

Met Asn Thr Pro Asn Asn Ala Leu Asp Val Ile Val Val Gly Ala Gly

1 5 10 15

Leu Val Gly Phe Ala Ala Ala Ala Ala Leu Arg Arg Gln Gly His Arg

20 25 30

Val Thr Ile Tyr Glu Thr Ser Ser Phe Lys Asn Glu Leu Gly Ala Gly

35 40 45

Leu Ala Ile Pro Pro Asn Thr Val Arg Gly Leu Ile Gly Leu Gly Cys

50 55 60

Val Ile Glu Asn Leu Asp Pro Val Glu Asn Leu Cys Phe Thr Ala Val

65 70 75 80

Ala Phe Asp Gly Ser Ala Gly Met Arg Ser Asp Gln Thr Asn Tyr Glu

85 90 95

Ala Ser Tyr Gly Leu Pro Trp Ile Met Val His Arg Val Asp Leu His

100 105 110

Asn Glu Leu Arg Arg Val Ala Leu Ser Ala Glu Gly Asn Thr Gly Pro

115 120 125

Pro Ala Glu Leu Arg Leu Asp His Arg Val Ser Ser Cys Asp Val Glu

130 135 140

Lys Cys Thr Val Thr Leu Ser Asn Gly Asp Thr His His Ala Asp Leu

145 150 155 160

Ile Ile Gly Ala Asp Gly Ile His Ser Thr Ile Arg Ser Phe Val Val

165 170 175

Gly Glu Glu Ile Val Ile Pro Pro Ser Lys Thr Ala Gly Phe Arg Trp

180 185 190

Leu Thr Glu Ser Thr Ala Leu Glu Pro Tyr Pro Glu Leu Asp Trp Ile

195 200 205

Val Lys Ile Pro Pro Leu Gly Ala Arg Leu Ile Ser Ala Pro Met Asn

210 215 220

Pro Ala Pro Pro Gln Val Asp His Arg Thr Ile Ile Ile Tyr Ala Cys

225 230 235 240

Arg Gly Ser Thr Leu Ile Asn Val Leu Gly Val His Glu Asp Leu Arg

245 250 255

Asp Gln Asp Thr Val Pro Trp Asn Ala Pro Val Thr Gln Ser Glu Leu

260 265 270

Leu Gln Phe Phe Gly Asp Tyr His Pro Arg Phe Lys Arg Leu Leu Glu

275 280 285

Leu Ala Asn Asp Val His Val Trp Gln Met Arg Val Val Pro Arg Leu

290 295 300

Glu Thr Trp Val Asn Arg Arg Val Cys Ile Met Gly Asp Ala Ala His

305 310 315 320

Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly Leu Glu Asp

325 330 335

Ala Val Ala Leu Gly Thr Leu Leu Pro Leu Gly Thr Thr Pro Glu Glu

340 345 350

Ile Pro Asp Arg Leu Thr Leu Trp Gln Asp Leu Val Lys Pro Arg Ala

355 360 365

Glu Phe Val Ala Thr Glu Ser Tyr Glu Gln Gln His Ile Pro Ala Lys

370 375 380

Arg Gly Leu Tyr Leu Arg Ser Gln Glu Met Arg Asp Trp Val Met Gly

385 390 395 400

Tyr Asp Val Gln Ala Glu Ala Gln Lys Val Leu Ala Gly Ala Val Asn

405 410 415

Arg Ser Lys Gly

420

<210> 19

<211> 1287

<212> DNA

<213> Armillaria mellea

<400> 19

atgcaacaaa tcgacgaagc gcgcccattg aaagtgatag tagtgggtgc tggactttgt 60

gggctttccg ccgccattgc acttcgtagg caagggcatc atgttcatat acttgaatct 120

tcaagtttta agagcgagct tggcgcaggt ctcgccgtcc cacccaatac tgtacgctct 180

cttcgaggcc taggttgtaa catcgacaat ctcaagcccg tggataattt gtgtttctct 240

gccatggcgc atgacggaag cccaggcatg atgaataaca tgacagacta tcacaaggcg 300

tacggtgatc cttgggtaat ggcacatcgt gtcgacctcc ataacgagct cttgcgagtg 360

gctttcgacc ccgaaggaac agggcctcct gctcaacttc gtttgggcgt ccaggtagtg 420

acttgcgata tggaagcttg tacaatttcc cttgtcgatg gaacagtctg ttccgccgat 480

cttatcgtag gagctgacgg tattaagtcg accatacgct cctgtgttct aggcaaagaa 540

atagacatac ctcctaccgg tatcgccgga taccgctggc tcataccggc agaagctttg 600

gagccctatc ccgagctcga ctggattatc aagaacccac ccctaggagc acgtttaatc 660

acggatcccg tacgccgaac tgaacaaacg gatgacggta agaaggctga caagcgcacg 720

atcataatct atgcgtgccg cagtggcacg atgatcaacg ttcttggtgt gcacgatgac 780

ctgcgcaacc agaatgaagt cggatggaac gtaccagtca cacgggaaaa cttgctggag 840

tttttcgggg actaccaccc acggtttaag cgtttactcc agctagccga tagtattcat 900

ttgtggcaaa tgcgtgttgt cccacggctt gacacatgga ttaatagatg cgtgtgtttg 960

ctgggcgatt ctgcacatgc gtcattacca actctcgggc aaggcttcgg aatgggtctt 1020

gaggatgccg tagctctcgc agccctcctt ccgatgggaa ccaatgcgtc tgacgttgag 1080

aaccgcctta tcgcctacga aagcttgcgt aaggagcgtg cagagtatgt agccacggaa 1140

tcattagaac agcaggatat tgcaggaaag cgaggcttgt atctcaggtc tcctatgatg 1200

cgcgataaaa taatgggtta tgatattaaa gcggaagctg agaaggtttt aatcgaatta 1260

aaaaattcga cagctcagca ggtcact 1287

<210> 20

<211> 429

<212>PRT

<213> Armillaria mellea

<400> 20

Met Gln Gln Ile Asp Glu Ala Arg Pro Leu Lys Val Ile Val Val Gly

1 5 10 15

Ala Gly Leu Cys Gly Leu Ser Ala Ala Ile Ala Leu Arg Arg Gln Gly

20 25 30

His His Val His Ile Leu Glu Ser Ser Ser Phe Lys Ser Glu Leu Gly

35 40 45

Ala Gly Leu Ala Val Pro Pro Asn Thr Val Arg Ser Leu Arg Gly Leu

50 55 60

Gly Cys Asn Ile Asp Asn Leu Lys Pro Val Asp Asn Leu Cys Phe Ser

65 70 75 80

Ala Met Ala His Asp Gly Ser Pro Gly Met Met Asn Asn Met Thr Asp

85 90 95

Tyr His Lys Ala Tyr Gly Asp Pro Trp Val Met Ala His Arg Val Asp

100 105 110

Leu His Asn Glu Leu Leu Arg Val Ala Phe Asp Pro Glu Gly Thr Gly

115 120 125

Pro Pro Ala Gln Leu Arg Leu Gly Val Gln Val Val Thr Cys Asp Met

130 135 140

Glu Ala Cys Thr Ile Ser Leu Val Asp Gly Thr Val Cys Ser Ala Asp

145 150 155 160

Leu Ile Val Gly Ala Asp Gly Ile Lys Ser Thr Ile Arg Ser Cys Val

165 170 175

Leu Gly Lys Glu Ile Asp Ile Pro Pro Thr Gly Ile Ala Gly Tyr Arg

180 185 190

Trp Leu Ile Pro Ala Glu Ala Leu Glu Pro Tyr Pro Glu Leu Asp Trp

195 200 205

Ile Ile Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Thr Asp Pro Val

210 215 220

Arg Arg Thr Glu Gln Thr Asp Asp Gly Lys Lys Ala Asp Lys Arg Thr

225 230 235 240

Ile Ile Ile Tyr Ala Cys Arg Ser Gly Thr Met Ile Asn Val Leu Gly

245 250 255

Val His Asp Asp Leu Arg Asn Gln Asn Glu Val Gly Trp Asn Val Pro

260 265 270

Val Thr Arg Glu Asn Leu Leu Glu Phe Phe Gly Asp Tyr His Pro Arg

275 280 285

Phe Lys Arg Leu Leu Gln Leu Ala Asp Ser Ile His Leu Trp Gln Met

290 295 300

Arg Val Val Pro Arg Leu Asp Thr Trp Ile Asn Arg Cys Val Cys Leu

305 310 315 320

Leu Gly Asp Ser Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe

325 330 335

Gly Met Gly Leu Glu Asp Ala Val Ala Leu Ala Ala Leu Leu Pro Met

340 345 350

Gly Thr Asn Ala Ser Asp Val Glu Asn Arg Leu Ile Ala Tyr Glu Ser

355 360 365

Leu Arg Lys Glu Arg Ala Glu Tyr Val Ala Thr Glu Ser Leu Glu Gln

370 375 380

Gln Asp Ile Ala Gly Lys Arg Gly Leu Tyr Leu Arg Ser Pro Met Met

385 390 395 400

Arg Asp Lys Ile Met Gly Tyr Asp Ile Lys Ala Glu Ala Glu Lys Val

405 410 415

Leu Ile Glu Leu Lys Asn Ser Thr Ala Gln Gln Val Thr

420 425

<210> 21

<211> 1278

<212> DNA

<213> Armillaria fuscipes

<400> 21

atgcaacaaa tcgacgaagc gcgtccatg aaggtgctag tcgtgggtgc tggactctgt 60

gggctttccg ccgccatcgc acttcgtaga caagggcatc acgtccatat acttgaatct 120

tcaagtttca agagcgaact tggggcaggt ctcgccgtgc cacctaatac tgtgcgctct 180

cttcgaggtc tcggctgtga catagacaat ctcaagcccg tggataatct ttgtttcact 240

gctatgtcgc atgacggaag cccaggcatg atgaataaca tgacggacta tcgcaaggcg 300

tatggtgatc cttgggtgat ggcacatcgt gtcgaccttc ataacgagct tatgcgagtg 360

gctctcgacc ctgatgggac aggtcctcct gcccaacttc gtttgggcgt ccaggttgtg 420

tcttgcgatg ttgaagcttg tactgtttcc cttgtcgatg gagaggtctg ttccgccgat 480

cttatcgttg gagctgatgg tatcaggtca accatacgct cctatgttct aggcaaagaa 540

atagatatac ctcctaccgg cattgccgga taccgctggc ttacaccatc agaggctttg 600

gagccttttc ccgaacttga ttggattatc aagaacccac ctctaggagc acgtctaatc 660

accgctccca tacgccggaa cgaacaaatg aatgacggtg agatggctga caagcgtacg 720

atcatcatct acgcgtgccg caacggcaca atgattaacg ttctcggtgt gcacgatgac 780

ccgcgcaacc agaatgaagt cggatggaac gtgccagtaa cccaagaaaa attgctcgaa 840

tttttcggag actaccaccc acggttcaaa agtttacttc agctatctga tagtattcat 900

ttgtggcaaa tgcgtgttgt tccacggctt gacacatgga tcaatcaacg tgtgtgtttg 960

ctgggcgatt ctgcacatgc gtcattacca acgctcgggc aaggcttcgg gatgggtctt 1020

gaggacgcca tagctcttgc aaccctcctt ccgatgggcg ccaaagtgtc ggacattgag 1080

aatcgcctta tcgcctacga aagcctgcgt aaggagcgtg cagagtttgt agccacggaa 1140

tcattagaac agcaagatat tcccgaaaag cgaggcttgt atctcagatc ccctatgatg 1200

cgcgataaaa taatgggtta cgatatcaaa gccgaagctg aaaaggtttt aatggagtta 1260

ttgagctcga aagctcaa 1278

<210> 22

<211> 426

<212>PRT

<213> Armillaria fuscipes

<400> 22

Met Gln Gln Ile Asp Glu Ala Arg Pro Leu Lys Val Leu Val Val Gly

1 5 10 15

Ala Gly Leu Cys Gly Leu Ser Ala Ala Ile Ala Leu Arg Arg Gln Gly

20 25 30

His His Val His Ile Leu Glu Ser Ser Ser Phe Lys Ser Glu Leu Gly

35 40 45

Ala Gly Leu Ala Val Pro Pro Asn Thr Val Arg Ser Leu Arg Gly Leu

50 55 60

Gly Cys Asp Ile Asp Asn Leu Lys Pro Val Asp Asn Leu Cys Phe Thr

65 70 75 80

Ala Met Ser His Asp Gly Ser Pro Gly Met Met Asn Asn Met Thr Asp

85 90 95

Tyr Arg Lys Ala Tyr Gly Asp Pro Trp Val Met Ala His Arg Val Asp

100 105 110

Leu His Asn Glu Leu Met Arg Val Ala Leu Asp Pro Asp Gly Thr Gly

115 120 125

Pro Pro Ala Gln Leu Arg Leu Gly Val Gln Val Val Ser Cys Asp Val

130 135 140

Glu Ala Cys Thr Val Ser Leu Val Asp Gly Glu Val Cys Ser Ala Asp

145 150 155 160

Leu Ile Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Ser Tyr Val

165 170 175

Leu Gly Lys Glu Ile Asp Ile Pro Pro Thr Gly Ile Ala Gly Tyr Arg

180 185 190

Trp Leu Thr Pro Ser Glu Ala Leu Glu Pro Phe Pro Glu Leu Asp Trp

195 200 205

Ile Ile Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Thr Ala Pro Ile

210 215 220

Arg Arg Asn Glu Gln Met Asn Asp Gly Glu Met Ala Asp Lys Arg Thr

225 230 235 240

Ile Ile Ile Tyr Ala Cys Arg Asn Gly Thr Met Ile Asn Val Leu Gly

245 250 255

Val His Asp Asp Pro Arg Asn Gln Asn Glu Val Gly Trp Asn Val Pro

260 265 270

Val Thr Gln Glu Lys Leu Leu Glu Phe Phe Gly Asp Tyr His Pro Arg

275 280 285

Phe Lys Ser Leu Leu Gln Leu Ser Asp Ser Ile His Leu Trp Gln Met

290 295 300

Arg Val Val Pro Arg Leu Asp Thr Trp Ile Asn Gln Arg Val Cys Leu

305 310 315 320

Leu Gly Asp Ser Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe

325 330 335

Gly Met Gly Leu Glu Asp Ala Ile Ala Leu Ala Thr Leu Leu Pro Met

340 345 350

Gly Ala Lys Val Ser Asp Ile Glu Asn Arg Leu Ile Ala Tyr Glu Ser

355 360 365

Leu Arg Lys Glu Arg Ala Glu Phe Val Ala Thr Glu Ser Leu Glu Gln

370 375 380

Gln Asp Ile Pro Glu Lys Arg Gly Leu Tyr Leu Arg Ser Pro Met Met

385 390 395 400

Arg Asp Lys Ile Met Gly Tyr Asp Ile Lys Ala Glu Ala Glu Lys Val

405 410 415

Leu Met Glu Leu Leu Ser Ser Lys Ala Gln

420 425

<210> 23

<211> 1266

<212> DNA

<213> Armillaria gallica

<400> 23

atgcaacaaa tcgacgaagc gcgcgcattg aaagtgatag tcgtgggtgc tggactttgc 60

gggctttccg ccgccattgt acttcgtagg caagggcatc acgtccatat acttgaatct 120

tcaagtttca agagcgaact tggtgcaggt cttgccgtgc cgcccaacac tgtacgctct 180

cttcgaggtc taggttgtaa catcgacaat ctcaagcccg tggataatct ttgtttcact 240

gccatggctc atgacggaag cccaggcatg atgaataaca tgacagacta tcagaaggcg 300

tacggcgatc cttgggtaat ggcgcatcgt gtcgacctcc ataacgagct catgcgagtg 360

gctctcgacc ctgaaggaac aggccctgct gcccagcttc gtttgggcgt ccaggtggtg 420

tcttgtgatg tggaagcttg taccatttct cttgtcgatg gatcaatctg taccgccgat 480

cttatcgtcg gagctgatgg tattaggtca accatacggt cctatgtttt gggcaaagaa 540

atagacatac ctcctaccgg tattgctgga taccgctggc tcacaccggc agaagctttg 600

gacccatatc ccgaactcga ctggattatc aagaacccac ccctgggagc acgtttaatc 660

acagctcccg ttcgccgaaa cgataaggcg gatgacggtg agaaagctga caagcgcacg 720

atcataatct acgcgtgccg cagtggcact atgatcaacg ttctcggtgt gcacgatgac 780

ccgcgcaacc agaatgaagt cggatggaac gtaccagtta cccaggaaaa tttgttggag 840

tttttcgaag actaccaccc acggtttaag cgtttacttc agctgaccga taacattcat 900

ttgtggcaaa tgcgtgttgt cccgcggctt gacacatgga ttaataaacg cgtgtgtttg 960

ctgggcgatt ctgcacatgc gtcattacca acactcgggc aaggcttcgg gatgggtctt 1020

gaggacgccg tagctctcgc aaccctcctt ccgatgggaa ccaaattgtc tgacattgaa 1080

aaccgtcttg tcgcctacga aagcttgcgt aaggagcgtg ctgagtatgt agccacggaa 1140

tcattagaac agcaggatat tccgggaaag cgaggcttgt atctcaggtc tcctgtgatg 1200

cgcgataaaa taatgggtta tgatatcaaa gccgaagctg agaaggtttt aatggaattg 1260

ataacc 1266

<210> 24

<211> 422

<212>PRT

<213> Armillaria gallica

<400> 24

Met Gln Gln Ile Asp Glu Ala Arg Ala Leu Lys Val Ile Val Val Gly

1 5 10 15

Ala Gly Leu Cys Gly Leu Ser Ala Ala Ile Val Leu Arg Arg Gln Gly

20 25 30

His His Val His Ile Leu Glu Ser Ser Ser Phe Lys Ser Glu Leu Gly

35 40 45

Ala Gly Leu Ala Val Pro Pro Asn Thr Val Arg Ser Leu Arg Gly Leu

50 55 60

Gly Cys Asn Ile Asp Asn Leu Lys Pro Val Asp Asn Leu Cys Phe Thr

65 70 75 80

Ala Met Ala His Asp Gly Ser Pro Gly Met Met Asn Asn Met Thr Asp

85 90 95

Tyr Gln Lys Ala Tyr Gly Asp Pro Trp Val Met Ala His Arg Val Asp

100 105 110

Leu His Asn Glu Leu Met Arg Val Ala Leu Asp Pro Glu Gly Thr Gly

115 120 125

Pro Ala Ala Gln Leu Arg Leu Gly Val Gln Val Val Ser Cys Asp Val

130 135 140

Glu Ala Cys Thr Ile Ser Leu Val Asp Gly Ser Ile Cys Thr Ala Asp

145 150 155 160

Leu Ile Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Ser Tyr Val

165 170 175

Leu Gly Lys Glu Ile Asp Ile Pro Pro Thr Gly Ile Ala Gly Tyr Arg

180 185 190

Trp Leu Thr Pro Ala Glu Ala Leu Asp Pro Tyr Pro Glu Leu Asp Trp

195 200 205

Ile Ile Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Thr Ala Pro Val

210 215 220

Arg Arg Asn Asp Lys Ala Asp Asp Gly Glu Lys Ala Asp Lys Arg Thr

225 230 235 240

Ile Ile Ile Tyr Ala Cys Arg Ser Gly Thr Met Ile Asn Val Leu Gly

245 250 255

Val His Asp Asp Pro Arg Asn Gln Asn Glu Val Gly Trp Asn Val Pro

260 265 270

Val Thr Gln Glu Asn Leu Leu Glu Phe Phe Glu Asp Tyr His Pro Arg

275 280 285

Phe Lys Arg Leu Leu Gln Leu Thr Asp Asn Ile His Leu Trp Gln Met

290 295 300

Arg Val Val Pro Arg Leu Asp Thr Trp Ile Asn Lys Arg Val Cys Leu

305 310 315 320

Leu Gly Asp Ser Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe

325 330 335

Gly Met Gly Leu Glu Asp Ala Val Ala Leu Ala Thr Leu Leu Pro Met

340 345 350

Gly Thr Lys Leu Ser Asp Ile Glu Asn Arg Leu Val Ala Tyr Glu Ser

355 360 365

Leu Arg Lys Glu Arg Ala Glu Tyr Val Ala Thr Glu Ser Leu Glu Gln

370 375 380

Gln Asp Ile Pro Gly Lys Arg Gly Leu Tyr Leu Arg Ser Pro Val Met

385 390 395 400

Arg Asp Lys Ile Met Gly Tyr Asp Ile Lys Ala Glu Ala Glu Lys Val

405 410 415

Leu Met Glu Leu Ile Thr

420

<210> 25

<211> 1278

<212> DNA

<213> Armillaria ostoyae

<400> 25

atgcaacaaa tcgacgaagc gcacccattg acagtgatag tcgtgggtgc tggactttgc 60

gggctttccg ccgccattgc acttcgtagg caagggcatc acgtccatat acttgaatct 120

tcaagtttca aaagcgaact tggcgcaggt ctcgccgtgc cgcccaacac tgtacgctct 180

cttcgaggtc taggttgtaa catcgacaat ctcaagcccg tggataatct ttgtttcact 240

gccatggcgc atgacggaag cccaggcatg atgaataaca tgacagacta tcacaaggcg 300

tacggcgagc cttgggtaat ggcgcatcgt gtcgacctcc ataacgagct catgcgagtg 360

gctctcgacc ctgaaggaac aggtcctcct gctcagcttc gtttgggtgt ccaggtggtg 420

tcttgcgatg tggaagcttg taccatttct cttgtcgatg gatcaatctg ttccgccgat 480

cttatcgtcg gagctgatgg tattaggtca accatacgct cctatgtttt gggcaaagaa 540

atagacatac ctcctaccgg tattgctgga taccgttggc tcacaccggc agaagctttg 600

gagccatatc ccgaactcga ctggattatc aagaacccac ccctgggagc acgtttaatc 660

acggctcccg tacgccgaaa cgataagacg gatgacggtg agaagactga caagcgcacg 720

atcataatct acgcgtgccg caatggcact atgatcaacg ttctcggtgt gcacgatgac 780

ccgcgcaacc agaatgaagt cggatggaac gtaccagtta cccaggaaaa tttgttggag 840

tttttcgaag actaccaccc acggtttaag cgtttacttc agttggccga tagtattcat 900

ttgtggcaaa tgcgtgttgt cccacggctt gacacatgga ttaataaacg cgtgtgtttg 960

ctgggcgatt ctgcacatgc gtcattacca acactcgggc aaggcttcgg gatgggtctt 1020

gaggacgccg tagctctcgc agccctcctt ccgatgggaa ccaaagtgtc tgacgttgag 1080

agtcgtctta tcgcctacga aagcttgcgt aaggagcgtg ctgagtatgt agccacggaa 1140

tcattagaac agcagaatat tccgggaaag cgaggcttgt atctcaggtc tcctatgatg 1200

cgcgataaaa taatgggtta tgatatcaaa gccgaagctg agaaggtttt aatggaatta 1260

ataacctcga cagctcag 1278

<210> 26

<211> 426

<212>PRT

<213> Armillaria ostoyae

<400> 26

Met Gln Gln Ile Asp Glu Ala His Pro Leu Thr Val Ile Val Val Gly

1 5 10 15

Ala Gly Leu Cys Gly Leu Ser Ala Ala Ile Ala Leu Arg Arg Gln Gly

20 25 30

His His Val His Ile Leu Glu Ser Ser Ser Phe Lys Ser Glu Leu Gly

35 40 45

Ala Gly Leu Ala Val Pro Pro Asn Thr Val Arg Ser Leu Arg Gly Leu

50 55 60

Gly Cys Asn Ile Asp Asn Leu Lys Pro Val Asp Asn Leu Cys Phe Thr

65 70 75 80

Ala Met Ala His Asp Gly Ser Pro Gly Met Met Asn Asn Met Thr Asp

85 90 95

Tyr His Lys Ala Tyr Gly Glu Pro Trp Val Met Ala His Arg Val Asp

100 105 110

Leu His Asn Glu Leu Met Arg Val Ala Leu Asp Pro Glu Gly Thr Gly

115 120 125

Pro Pro Ala Gln Leu Arg Leu Gly Val Gln Val Val Ser Cys Asp Val

130 135 140

Glu Ala Cys Thr Ile Ser Leu Val Asp Gly Ser Ile Cys Ser Ala Asp

145 150 155 160

Leu Ile Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Ser Tyr Val

165 170 175

Leu Gly Lys Glu Ile Asp Ile Pro Pro Thr Gly Ile Ala Gly Tyr Arg

180 185 190

Trp Leu Thr Pro Ala Glu Ala Leu Glu Pro Tyr Pro Glu Leu Asp Trp

195 200 205

Ile Ile Lys Asn Pro Pro Leu Gly Ala Arg Leu Ile Thr Ala Pro Val

210 215 220

Arg Arg Asn Asp Lys Thr Asp Asp Gly Glu Lys Thr Asp Lys Arg Thr

225 230 235 240

Ile Ile Ile Tyr Ala Cys Arg Asn Gly Thr Met Ile Asn Val Leu Gly

245 250 255

Val His Asp Asp Pro Arg Asn Gln Asn Glu Val Gly Trp Asn Val Pro

260 265 270

Val Thr Gln Glu Asn Leu Leu Glu Phe Phe Glu Asp Tyr His Pro Arg

275 280 285

Phe Lys Arg Leu Leu Gln Leu Ala Asp Ser Ile His Leu Trp Gln Met

290 295 300

Arg Val Val Pro Arg Leu Asp Thr Trp Ile Asn Lys Arg Val Cys Leu

305 310 315 320

Leu Gly Asp Ser Ala His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe

325 330 335

Gly Met Gly Leu Glu Asp Ala Val Ala Leu Ala Ala Leu Leu Pro Met

340 345 350

Gly Thr Lys Val Ser Asp Val Glu Ser Arg Leu Ile Ala Tyr Glu Ser

355 360 365

Leu Arg Lys Glu Arg Ala Glu Tyr Val Ala Thr Glu Ser Leu Glu Gln

370 375 380

Gln Asn Ile Pro Gly Lys Arg Gly Leu Tyr Leu Arg Ser Pro Met Met

385 390 395 400

Arg Asp Lys Ile Met Gly Tyr Asp Ile Lys Ala Glu Ala Glu Lys Val

405 410 415

Leu Met Glu Leu Ile Thr Ser Thr Ala Gln

420 425

<210> 27

<211> 1266

<212> DNA

<213> Mycena chlorophos

<400> 27

atgccctcca ccgccgaatc tccgcagccg ctcaagatcg tcatcgtcgg tgctgggctc 60

gttggtctcg ctgctgccat tgcgcttcgt cgcgagggtc atcatgtaga gatatacgaa 120

tcgtcgacgt tcaagaccga actcggcgcc ggtctcgcga taccgtgcaa taccctccgt 180

agcctcattg agctggggtg cattgtggct aacttgaacc cggtggagaa cctctgtttc 240

acgtctatgg cgcacgacgg aagcgagtca ggcatgcgaa gcgaccacac cgactacgag 300

gcgcgttatg ggaccccttg ggtcatggcg catcgcgtcg acatacacgc agagctgctc 360

cgaatggcca ccacctccga tattcccggc ccaccggcga cactgcatct cggccaacgc 420

gtccttgcct gcaatgtgtc cgactgctcc attgcactgg ccaccggcaa aacgatctca 480

gcggatctcg tcgttggtgc cgatgggatt cgctcgacca ttcgagctgc tgttcttggc 540

gaagatatcc acattccggc atcgggcact gccggcttcc gatggctcgt agattccgcc 600

gcgctggatc cctatcccga gctggactgg attgtgaaag cccgtccgct tggcgctcgc 660

gttatttctg ccccgatggg cctcgcactc gaagatcatc gtaccattgt gatctatgcg 720

tgtcgcggcg gtaacttgat caacgttctt gcggtccacg aagacaagcg cgaccaggag 780

gctgtccctt ggaatgtccc tttgacgcgc gaagccctct tggacttctt tagcgactac 840

cacccgcgtt tccgccgtct cttcgagctc gcgccggtcg acggaattca cgtctggcag 900

atgcgggtcg taccaccttt ggccaactgg atccgtgacc gcgtttgcat tctcggcgac 960

gcggcgcatg cgtctcttcc tactatgggc cagggctttg gccaaggtct cgaagacgcc 1020

gttgcgctag cgactttgct cccgctagga acgcgtagaa cggatatccc cgctcgtcta 1080

gtggcgtatg aggggatgcg caagcctcgg accgagtgga ttgcacgcga atcgtttgag 1140

cagcaggccg tcgcggaaaa gcgcggcatt tacttgcgct ctatcgaaat gcgcgatgcg 1200

gttatggggt ataatgttcg cgaggaggct aagcgcgtct tgtccgagct cactaaatct 1260

gattgt 1266

<210> 28

<211> 422

<212>PRT

<213> Mycena chlorophos

<400> 28

Met Pro Ser Thr Ala Glu Ser Pro Gln Pro Leu Lys Ile Val Ile Val

1 5 10 15

Gly Ala Gly Leu Val Gly Leu Ala Ala Ala Ile Ala Leu Arg Arg Glu

20 25 30

Gly His His Val Glu Ile Tyr Glu Ser Ser Thr Phe Lys Thr Glu Leu

35 40 45

Gly Ala Gly Leu Ala Ile Pro Cys Asn Thr Leu Arg Ser Leu Ile Glu

50 55 60

Leu Gly Cys Ile Val Ala Asn Leu Asn Pro Val Glu Asn Leu Cys Phe

65 70 75 80

Thr Ser Met Ala His Asp Gly Ser Glu Ser Gly Met Arg Ser Asp His

85 90 95

Thr Asp Tyr Glu Ala Arg Tyr Gly Thr Pro Trp Val Met Ala His Arg

100 105 110

Val Asp Ile His Ala Glu Leu Leu Arg Met Ala Thr Thr Ser Asp Ile

115 120 125

Pro Gly Pro Pro Ala Thr Leu His Leu Gly Gln Arg Val Leu Ala Cys

130 135 140

Asn Val Ser Asp Cys Ser Ile Ala Leu Ala Thr Gly Lys Thr Ile Ser

145 150 155 160

Ala Asp Leu Val Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Ala

165 170 175

Ala Val Leu Gly Glu Asp Ile His Ile Pro Ala Ser Gly Thr Ala Gly

180 185 190

Phe Arg Trp Leu Val Asp Ser Ala Ala Leu Asp Pro Tyr Pro Glu Leu

195 200 205

Asp Trp Ile Val Lys Ala Arg Pro Leu Gly Ala Arg Val Ile Ser Ala

210 215 220

Pro Met Gly Leu Ala Leu Glu Asp His Arg Thr Ile Val Ile Tyr Ala

225 230 235 240

Cys Arg Gly Gly Asn Leu Ile Asn Val Leu Ala Val His Glu Asp Lys

245 250 255

Arg Asp Gln Glu Ala Val Pro Trp Asn Val Pro Leu Thr Arg Glu Ala

260 265 270

Leu Leu Asp Phe Phe Ser Asp Tyr His Pro Arg Phe Arg Arg Leu Phe

275 280 285

Glu Leu Ala Pro Val Asp Gly Ile His Val Trp Gln Met Arg Val Val

290 295 300

Pro Pro Leu Ala Asn Trp Ile Arg Asp Arg Val Cys Ile Leu Gly Asp

305 310 315 320

Ala Ala His Ala Ser Leu Pro Thr Met Gly Gln Gly Phe Gly Gln Gly

325 330 335

Leu Glu Asp Ala Val Ala Leu Ala Thr Leu Leu Pro Leu Gly Thr Arg

340 345 350

Arg Thr Asp Ile Pro Ala Arg Leu Val Ala Tyr Glu Gly Met Arg Lys

355 360 365

Pro Arg Thr Glu Trp Ile Ala Arg Glu Ser Phe Glu Gln Gln Ala Val

370 375 380

Ala Glu Lys Arg Gly Ile Tyr Leu Arg Ser Ile Glu Met Arg Asp Ala

385 390 395 400

Val Met Gly Tyr Asn Val Arg Glu Glu Ala Lys Arg Val Leu Ser Glu

405 410 415

Leu Thr Lys Ser Asp Cys

420

<210> 29

<211> 72

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 1 for

hispidin hydroxylases

<220>

<221> MISC_FEATURE

<222> (1)..(72)

<223> X - any amino acid

<400> 29

Val Gly Ala Gly Leu Xaa Gly Xaa Xaa Ala Ala Xaa Xaa Leu Arg Arg

1 5 10 15

Xaa Gly His Xaa Xaa Xaa Xaa Xaa Xaa Xaa Ser Xaa Phe Lys Xaa Glu

20 25 30

Xaa Gly Ala Gly Xaa Ala Xaa Pro Xaa Asn Thr Xaa Xaa Xaa Leu Xaa

35 40 45

Xaa Leu Gly Cys Xaa Xaa Xaa Asn Xaa Xaa Xaa Val Xaa Asn Leu Cys

50 55 60

Xaa Xaa Xaa Xaa Xaa Xaa Asp Gly

65 70

<210> 30

<211> 33

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 2 for

hispidin hydroxylases

<220>

<221> MISC_FEATURE

<222> (1)..(33)

<223> X - any amino acid

<400> 30

Gly Met Xaa Xaa Xaa Xaa Xaa Xaa Tyr Xaa Xaa Xaa Tyr Gly Xaa Xaa

1 5 10 15

Trp Xaa Met Xaa His Arg Val Asp Xaa His Xaa Glu Leu Xaa Arg Xaa

20 25 30

Ala

<210> 31

<211> 96

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 3 for

hispidin hydroxylases

<220>

<221> MISC_FEATURE

<222> (1)..(96)

<223> X - any amino acid

<400> 31

Gly Pro Xaa Ala Xaa Leu Xaa Leu Xaa Xaa Xaa Val Xaa Xaa Cys Xaa

1 5 10 15

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Gly Xaa Xaa Xaa Xaa Ala

20 25 30

Asp Leu Xaa Xaa Gly Ala Asp Gly Ile Xaa Ser Xaa Xaa Arg Xaa Xaa

35 40 45

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Pro Xaa Xaa Xaa Xaa Xaa Gly Xaa

50 55 60

Arg Trp Xaa Xaa Xaa Xaa Xaa Xaa Leu Xaa Xaa Xaa Pro Glu Leu Asp

65 70 75 80

Trp Xaa Xaa Lys Xaa Xaa Pro Leu Gly Ala Arg Xaa Xaa Xaa Xaa Pro

85 90 95

<210> 32

<211> 57

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 4 for

hispidin hydroxylases

<220>

<221> MISC_FEATURE

<222> (1)..(57)

<223> X - any amino acid

<400> 32

Arg Thr Ile Xaa Xaa Tyr Ala Cys Arg Xaa Xaa Xaa Xaa Xaa Asn Val

1 5 10 15

Leu Xaa Xaa His Xaa Asp Xaa Arg Xaa Gln Xaa Xaa Xaa Xaa Trp Xaa

20 25 30

Xaa Pro Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Asp Tyr His

35 40 45

Pro Arg Phe Xaa Xaa Leu Xaa Xaa Leu

50 55

<210> 33

<211> 83

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 5 for

hispidin hydroxylases

<220>

<221> MISC_FEATURE

<222> (1)..(82)

<223> X - any amino acid

<400> 33

Trp Gln Xaa Arg Val Xaa Pro Xaa Leu Xaa Xaa Trp Xaa Xaa Xaa Xaa

1 5 10 15

Xaa Cys Xaa Xaa Gly Asp Xaa Ala His Ala Ser Leu Pro Thr Xaa Gly

20 25 30

Gln Gly Phe Gly Xaa Gly Leu Glu Asp Ala Xaa Ala Leu Xaa Xaa Leu

35 40 45

Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Arg Leu Xaa Xaa

50 55 60

Xaa Xaa Xaa Xaa Xaa Lys Xaa Arg Xaa Glu Xaa Xaa Xaa Xaa Glu Ser

65 70 75 80

Xaa Glu Gln

<210> 34

<211> 4832

<212> DNA

<213> Neonothopanus nambi

<400> 34

aaaaccatcc ttatattctc gctttgatgc tggctgtatg gaaacttgga ggcaccttcg 60

ctcctattga tgtccattct cctgccgaat tggtagctgg catgctgaac atagtctctc 120

cttcttgctt ggttattccg agctcagatg taactaatca aactcttgcg tgcgatctta 180

atatccccgt cgttgcattt cacccacatc aatccactat tcctgagctg aacaagaagt 240

acctcaccga ttctcaaatt tctccggatc ttcctttttc agatccaaac cggcctgctc 300

tgtacctctt cacttcgtcc gccacttctc gaagtaatct caaatgcgtg cctctcactc 360

acacctttat cttacgcaac agcctctcga agcgtgcatg gtgcaagcgt atgcgtccag 420

agacagactt tgacggcata cgcgttcttg gatgggcccc gtggtctcac gtcctagcac 480

acatgcaaga catcggacca ctcaccttac ttaatgccgg atgctacgtt tttgcgacta 540

ctccatccac gtaccctacg gaattgaagg acgacaggga cctgatatct tgcgcggcaa 600

atgctatcat gtacaagggc gtcaagtcat ttgcttgtct tccctttgta ctcggagggc 660

tgaaggcatt atgcgagtct gagccatccg tgaaggcgca tctacaggtc gaggagagag 720

ctcaactcct gaagtctctg caacacatgg aaattcttga gtgtggaggt gccatgctcg 780

aagcaagtgt tgcgtcttgg gctattgaga actgcattcc catttcgatc ggtattggta 840

tgacggagac tggtggagcg ctctttgcag gccccgttca ggccatcaaa accgggtttt 900

cttcagagga taaattcatt gaagatgcta cttacttgct cgttaaggat gatcatgaga 960

gtcatgctga ggaggatatt aacgaggtg aactagttgt gaaaagtaaa atgctcccac 1020

gaggctacct tggctatagt gatccttcct tctcagtcga cgatgctggc tgggttacat 1080

ttagaacagg agacagatac agcgttacac ctgacggaaa gttttcctgg ctgggccgga 1140

acactgattt cattcagatg accagtggtg agacgctgga tccccgacca attgagagct 1200

cgctctgcga aagttctctt atttctagag catgcgttat cggagataaa tttctcaacg 1260

ggcctgctgc tgctgtttgt gcgatcattg agcttgagcc cacagcggtg gaaaaaggac 1320

aagctcactc gcgtgagata gcaagagttt tcgcacctat taatcgagac ctaccgcctc 1380

ctcttaggat tgcatggtcg cacgttttgg ttctccagcc ctcggagaag ataccgatga 1440

cgaagaaggg taccatcttc cgcaagaaaa ttgagcaggt gtttggctct gcgttgggtg 1500

gcagctctgg agataactct caagccactg cggatgctgg cgttgttcga cgagacgagt 1560

tatcgaacac tgtcaagcac ataattagcc gtgttttagg agtttccgat gacgaattac 1620

tttggacgct atcatttgcg gagttaggaa tgacgtcagc actagccact cgcatcgcca 1680

acgagttgaa cgaagtttta gttggagtta atctccctat caacgcttgc tatatacatg 1740

tcgaccttcc ttctctaagc aatgccgtct atgcgaaact tgcacacctc aagttaccag 1800

atcgtactcc cgaacccagg caagcccctg tcgaaaactc tggtgggaag gagatcgttg 1860

tcgttggcca ggcctttcgt cttcctggct caataaacga tgtcgcctct cttcgagacg 1920

cattcctggc gagacaagca tcatccatta tcactgaaat accatccgat cgctgggacc 1980

acgccagctt ctatcccaag gatatacgtt tcaacaaggc tggccttgtg gatatagcca 2040

attatgatca tagctttttc ggactgacgg caaccgaagc gctctatctg tcgccaacta 2100

tgcgtctagc attagaagtt tcgtttgaag cgctagagaa tgctaatatc ccggtgtcac 2160

aactcaaggg ttcgcaaaca gcggtttatg ttgctactac agatgacgga tttgagaccc 2220

ttttgaatgc cgaggccggc tatgatgctt atacaagatt ctatggcact ggtcgagcag 2280

caagtacagc gagcgggcgc ataagctgtc ttcttgatgt ccatggaccc tctattactg 2340

ttgatacggc atgcagtgga ggggctgttt gtattgacca agcaatcgac tatctacaat 2400

catcgagtgc agcagacacc gctatcatat gtgctagtaa cacgcactgc tggccaggct 2460

cgttcaggtt tctttccgca caagggatgg tatccccagg aggacgatgc gcgacattta 2520

caactgatgc tgatggctac gtgccctctg agggcgcggt cgccttcata ttgaaaaccc 2580

gagaagcagc tatgcgtgac aaggacacta tcctcgcgac aatcaaagcg acacagatat 2640

cgcacaatgg ccgatctcaa ggtcttgtgg caccgaatgt caactcgcaa gctgaccttc 2700

atcgctcgtt gcttcaaaaa gctggcctta gcccggctga tatccgtttc attgaagctc 2760

atgggacagg aacgtcactg ggagacctct cagaaattca agctataaat gatgcttata 2820

cctcctctca gccgcgcacg accggcccac tcatagtcag cgcttccaaa acggtcattg 2880

gtcataccga accagctggc cccttggtcg gtatgctgtc ggtcttgaac tctttcaaag 2940

aaggcgccgt ccctggtctc gcccatctta ccgcagacaa tttgaatccc tcgctggact 3000

gttcttctgt gccacttctc attccctatc aacctgttca cctggctgca cccaagcctc 3060

accgagctgc tgtaaggtca tacggctttt caggtaccct gggcggcatc gttctagagg 3120

ctcctgacga agaaagatta gaagaagagc tgccaaatga caagcccatg ttgttcgtcg 3180

tcagcgcaaa gacacataca gcactaatcg aatacctggg gcggtatctc gagttcctct 3240

tgcaggcgaa cccccaagat ttttgtgaca tttgttatac aagctgcgtt gggcgggagc 3300

actatagata tcgctatgct tgtgtagcaa atgatatgga ggacctcata ggccaactcc 3360

agaaacgttt gggcagcaag gtgccgccaa agccgtcata caaacgcggt gctttggcct 3420

ttgccttttc tggtcagggt acacaattcc gagggatggc gacagagctt gcaaaagcgt 3480

actccggctt ccgaaagatc gtgtcggatc tcgcaaagag agctagcgag ttgtcaggtc 3540

atgccattga ccgttttctt cttgcatatg acataggcgc tgaaaatgta gctcctgata 3600

gtgaggcaga ccagatttgc atctttgtgt atcagtgttc tgtccttcgc tggctgcaga 3660

ctatggggat tagacccagt gcagtgatag gccatagcct cggggagatc tcagcttctg 3720

tggcggcagg agcactttct cttgactccg ctttggatct tgtcatctca cgagctcgcc 3780

ttttgcgctc ttcggcaagt gctcctgcag gaatggcagc tatgtctgcc tcgcaagacg 3840

aggttgtgga gttgattggg aaactagacc tcgacaaggc taattcgctc agcgtttcgg 3900

tcataaatgg tccccaaaat actgtcgtgt ccggctcttc agcggctatt gaaagcatag 3960

tggctttagc gaaagggaga aagatcaaag cgtctgccct gaatatcaat caagcttttc 4020

atagtccata cgtcgacagt gccgtccctg gtctccgtgc ttggtcagaa aagcatatct 4080

cctcagctcg gccattgcaa attccgctgt attcaacgtt gttgggagca caaatctctg 4140

agggagagat gttgaatcca gatcactggg tcgaccatgc acggaagcct gtacagttcg 4200

cacaagcagc cacaaccatg aaagaatcct tcaccggagt catcatagat atcggccctc 4260

aagtagtggc ttggtcactt ctgctctcga acgggctcac gtccgtgact gcgctcgctg 4320

cgaaaagagg gagaagtcaa caggtggctt tcttaagcgc cttggcggat ttgtatcaag 4380

attacggtgt tgttcctgat tttgtcgggc tttatgctca gcaggaagat gcttcgaggt 4440

tgaagaagac ggatatcttg acgtatccgt tccagcgggg cgaagagact ctttctagtg 4500

gttctagcac tccgacattg gaaaacacgg atttggattc cggtaaggaa ttacttatgg 4560

gaccgactcg ggggttgtta cgcgcggacg acttgcgtga cagtatcgtt tcttctgtga 4620

aggatgttct ggaactcaag tcaaatgaag acctcgattt gtctgaaagt ctgaatgcgc 4680

ttggtatgga ctcgatcatg ttcgctcagt tacggaagcg tattggggaa ggactcggat 4740

tgaatgttcc gatggttttt ctgtcggacg cgttttctat tggtgagatg gttagtaatc 4800

ttgtggaaca ggcggaggcg tctgaggaca at 4832

<210> 35

<211> 1678

<212>PRT

<213> Neonothopanus nambi

<400> 35

Met Asn Ser Ser Lys Asn Pro Pro Ser Thr Leu Leu Asp Val Phe Leu

1 5 10 15

Asp Thr Ala Arg Asn Leu Asp Thr Ala Leu Arg Asn Val Leu Glu Cys

20 25 30

Gly Glu His Arg Trp Ser Tyr Arg Glu Leu Asp Thr Val Ser Ser Ala

35 40 45

Leu Ala Gln His Leu Arg Tyr Thr Val Gly Leu Ser Pro Thr Val Ala

50 55 60

Val Ile Ser Glu Asn His Pro Tyr Ile Leu Ala Leu Met Leu Ala Val

65 70 75 80

Trp Lys Leu Gly Gly Thr Phe Ala Pro Ile Asp Val His Ser Pro Ala

85 90 95

Glu Leu Val Ala Gly Met Leu Asn Ile Val Ser Pro Ser Cys Leu Val

100 105 110

Ile Pro Ser Ser Asp Val Thr Asn Gln Thr Leu Ala Cys Asp Leu Asn

115 120 125

Ile Pro Val Val Ala Phe His Pro His Gln Ser Thr Ile Pro Glu Leu

130 135 140

Asn Lys Lys Tyr Leu Thr Asp Ser Gln Ile Ser Pro Asp Leu Pro Phe

145 150 155 160

Ser Asp Pro Asn Arg Pro Ala Leu Tyr Leu Phe Thr Ser Ser Ala Thr

165 170 175

Ser Arg Ser Asn Leu Lys Cys Val Pro Leu Thr His Thr Phe Ile Leu

180 185 190

Arg Asn Ser Leu Ser Lys Arg Ala Trp Cys Lys Arg Met Arg Pro Glu

195 200 205

Thr Asp Phe Asp Gly Ile Arg Val Leu Gly Trp Ala Pro Trp Ser His

210 215 220

Val Leu Ala His Met Gln Asp Ile Gly Pro Leu Thr Leu Leu Asn Ala

225 230 235 240

Gly Cys Tyr Val Phe Ala Thr Thr Pro Ser Thr Tyr Pro Thr Glu Leu

245 250 255

Lys Asp Asp Arg Asp Leu Ile Ser Cys Ala Ala Asn Ala Ile Met Tyr

260 265 270

Lys Gly Val Lys Ser Phe Ala Cys Leu Pro Phe Val Leu Gly Gly Leu

275 280 285

Lys Ala Leu Cys Glu Ser Glu Pro Ser Val Lys Ala His Leu Gln Val

290 295 300

Glu Glu Arg Ala Gln Leu Leu Lys Ser Leu Gln His Met Glu Ile Leu

305 310 315 320

Glu Cys Gly Gly Ala Met Leu Glu Ala Ser Val Ala Ser Trp Ala Ile

325 330 335

Glu Asn Cys Ile Pro Ile Ser Ile Gly Ile Gly Met Thr Glu Thr Gly

340 345 350

Gly Ala Leu Phe Ala Gly Pro Val Gln Ala Ile Lys Thr Gly Phe Ser

355 360 365

Ser Glu Asp Lys Phe Ile Glu Asp Ala Thr Tyr Leu Leu Val Lys Asp

370 375 380

Asp His Glu Ser His Ala Glu Glu Asp Ile Asn Glu Gly Glu Leu Val

385 390 395 400

Val Lys Ser Lys Met Leu Pro Arg Gly Tyr Leu Gly Tyr Ser Asp Pro

405 410 415

Ser Phe Ser Val Asp Asp Ala Gly Trp Val Thr Phe Arg Thr Gly Asp

420 425 430

Arg Tyr Ser Val Thr Pro Asp Gly Lys Phe Ser Trp Leu Gly Arg Asn

435 440 445

Thr Asp Phe Ile Gln Met Thr Ser Gly Glu Thr Leu Asp Pro Arg Pro

450 455 460

Ile Glu Ser Ser Leu Cys Glu Ser Ser Leu Ile Ser Arg Ala Cys Val

465 470 475 480

Ile Gly Asp Lys Phe Leu Asn Gly Pro Ala Ala Ala Val Cys Ala Ile

485 490 495

Ile Glu Leu Glu Pro Thr Ala Val Glu Lys Gly Gln Ala His Ser Arg

500 505 510

Glu Ile Ala Arg Val Phe Ala Pro Ile Asn Arg Asp Leu Pro Pro Pro

515 520 525

Leu Arg Ile Ala Trp Ser His Val Leu Val Leu Gln Pro Ser Glu Lys

530 535 540

Ile Pro Met Thr Lys Lys Gly Thr Ile Phe Arg Lys Lys Ile Glu Gln

545 550 555 560

Val Phe Gly Ser Ala Leu Gly Gly Ser Ser Gly Asp Asn Ser Gln Ala

565 570 575

Thr Ala Asp Ala Gly Val Val Arg Arg Asp Glu Leu Ser Asn Thr Val

580 585 590

Lys His Ile Ile Ser Arg Val Leu Gly Val Ser Asp Asp Glu Leu Leu

595 600 605

Trp Thr Leu Ser Phe Ala Glu Leu Gly Met Thr Ser Ala Leu Ala Thr

610 615 620

Arg Ile Ala Asn Glu Leu Asn Glu Val Leu Val Gly Val Asn Leu Pro

625 630 635 640

Ile Asn Ala Cys Tyr Ile His Val Asp Leu Pro Ser Leu Ser Asn Ala

645 650 655

Val Tyr Ala Lys Leu Ala His Leu Lys Leu Pro Asp Arg Thr Pro Glu

660 665 670

Pro Arg Gln Ala Pro Val Glu Asn Ser Gly Gly Lys Glu Ile Val Val

675 680 685

Val Gly Gln Ala Phe Arg Leu Pro Gly Ser Ile Asn Asp Val Ala Ser

690 695 700

Leu Arg Asp Ala Phe Leu Ala Arg Gln Ala Ser Ser Ile Ile Thr Glu

705 710 715 720

Ile Pro Ser Asp Arg Trp Asp His Ala Ser Phe Tyr Pro Lys Asp Ile

725 730 735

Arg Phe Asn Lys Ala Gly Leu Val Asp Ile Ala Asn Tyr Asp His Ser

740 745 750

Phe Phe Gly Leu Thr Ala Thr Glu Ala Leu Tyr Leu Ser Pro Thr Met

755 760 765

Arg Leu Ala Leu Glu Val Ser Phe Glu Ala Leu Glu Asn Ala Asn Ile

770 775 780

Pro Val Ser Gln Leu Lys Gly Ser Gln Thr Ala Val Tyr Val Ala Thr

785 790 795 800

Thr Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Glu Ala Gly Tyr Asp

805 810 815

Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Ala Ser Thr Ala Ser

820 825 830

Gly Arg Ile Ser Cys Leu Leu Asp Val His Gly Pro Ser Ile Thr Val

835 840 845

Asp Thr Ala Cys Ser Gly Gly Ala Val Cys Ile Asp Gln Ala Ile Asp

850 855 860

Tyr Leu Gln Ser Ser Ser Ala Ala Asp Thr Ala Ile Ile Cys Ala Ser

865 870 875 880

Asn Thr His Cys Trp Pro Gly Ser Phe Arg Phe Leu Ser Ala Gln Gly

885 890 895

Met Val Ser Pro Gly Gly Arg Cys Ala Thr Phe Thr Thr Asp Ala Asp

900 905 910

Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys Thr Arg

915 920 925

Glu Ala Ala Met Arg Asp Lys Asp Thr Ile Leu Ala Thr Ile Lys Ala

930 935 940

Thr Gln Ile Ser His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn

945 950 955 960

Val Asn Ser Gln Ala Asp Leu His Arg Ser Leu Leu Gln Lys Ala Gly

965 970 975

Leu Ser Pro Ala Asp Ile Arg Phe Ile Glu Ala His Gly Thr Gly Thr

980 985 990

Ser Leu Gly Asp Leu Ser Glu Ile Gln Ala Ile Asn Asp Ala Tyr Thr

995 1000 1005

Ser Ser Gln Pro Arg Thr Thr Gly Pro Leu Ile Val Ser Ala Ser

1010 1015 1020

Lys Thr Val Ile Gly His Thr Glu Pro Ala Gly Pro Leu Val Gly

1025 1030 1035

Met Leu Ser Val Leu Asn Ser Phe Lys Glu Gly Ala Val Pro Gly

1040 1045 1050

Leu Ala His Leu Thr Ala Asp Asn Leu Asn Pro Ser Leu Asp Cys

1055 1060 1065

Ser Ser Val Pro Leu Leu Ile Pro Tyr Gln Pro Val His Leu Ala

1070 1075 1080

Ala Pro Lys Pro His Arg Ala Ala Val Arg Ser Tyr Gly Phe Ser

1085 1090 1095

Gly Thr Leu Gly Gly Ile Val Leu Glu Ala Pro Asp Glu Glu Arg

1100 1105 1110

Leu Glu Glu Glu Leu Pro Asn Asp Lys Pro Met Leu Phe Val Val

1115 1120 1125

Ser Ala Lys Thr His Thr Ala Leu Ile Glu Tyr Leu Gly Arg Tyr

1130 1135 1140

Leu Glu Phe Leu Leu Gln Ala Asn Pro Gln Asp Phe Cys Asp Ile

1145 1150 1155

Cys Tyr Thr Ser Cys Val Gly Arg Glu His Tyr Arg Tyr Arg Tyr

1160 1165 1170

Ala Cys Val Ala Asn Asp Met Glu Asp Leu Ile Gly Gln Leu Gln

1175 1180 1185

Lys Arg Leu Gly Ser Lys Val Pro Pro Lys Pro Ser Tyr Lys Arg

1190 1195 1200

Gly Ala Leu Ala Phe Ala Phe Ser Gly Gln Gly Thr Gln Phe Arg

1205 1210 1215

Gly Met Ala Thr Glu Leu Ala Lys Ala Tyr Ser Gly Phe Arg Lys

1220 1225 1230

Ile Val Ser Asp Leu Ala Lys Arg Ala Ser Glu Leu Ser Gly His

1235 1240 1245

Ala Ile Asp Arg Phe Leu Leu Ala Tyr Asp Ile Gly Ala Glu Asn

1250 1255 1260

Val Ala Pro Asp Ser Glu Ala Asp Gln Ile Cys Ile Phe Val Tyr

1265 1270 1275

Gln Cys Ser Val Leu Arg Trp Leu Gln Thr Met Gly Ile Arg Pro

1280 1285 1290

Ser Ala Val Ile Gly His Ser Leu Gly Glu Ile Ser Ala Ser Val

1295 1300 1305

Ala Ala Gly Ala Leu Ser Leu Asp Ser Ala Leu Asp Leu Val Ile

1310 1315 1320

Ser Arg Ala Arg Leu Leu Arg Ser Ser Ala Ser Ala Pro Ala Gly

1325 1330 1335

Met Ala Ala Met Ser Ala Ser Gln Asp Glu Val Val Glu Leu Ile

1340 1345 1350

Gly Lys Leu Asp Leu Asp Lys Ala Asn Ser Leu Ser Val Ser Val

1355 1360 1365

Ile Asn Gly Pro Gln Asn Thr Val Val Ser Gly Ser Ser Ala Ala

1370 1375 1380

Ile Glu Ser Ile Val Ala Leu Ala Lys Gly Arg Lys Ile Lys Ala

1385 1390 1395

Ser Ala Leu Asn Ile Asn Gln Ala Phe His Ser Pro Tyr Val Asp

1400 1405 1410

Ser Ala Val Pro Gly Leu Arg Ala Trp Ser Glu Lys His Ile Ser

1415 1420 1425

Ser Ala Arg Pro Leu Gln Ile Pro Leu Tyr Ser Thr Leu Leu Gly

1430 1435 1440

Ala Gln Ile Ser Glu Gly Glu Met Leu Asn Pro Asp His Trp Val

1445 1450 1455

Asp His Ala Arg Lys Pro Val Gln Phe Ala Gln Ala Ala Thr Thr

1460 1465 1470

Met Lys Glu Ser Phe Thr Gly Val Ile Ile Asp Ile Gly Pro Gln

1475 1480 1485

Val Val Ala Trp Ser Leu Leu Leu Ser Asn Gly Leu Thr Ser Val

1490 1495 1500

Thr Ala Leu Ala Ala Lys Arg Gly Arg Ser Gln Gln Val Ala Phe

1505 1510 1515

Leu Ser Ala Leu Ala Asp Leu Tyr Gln Asp Tyr Gly Val Val Pro

1520 1525 1530

Asp Phe Val Gly Leu Tyr Ala Gln Gln Glu Asp Ala Ser Arg Leu

1535 1540 1545

Lys Lys Thr Asp Ile Leu Thr Tyr Pro Phe Gln Arg Gly Glu Glu

1550 1555 1560

Thr Leu Ser Ser Gly Ser Ser Thr Pro Thr Leu Glu Asn Thr Asp

1565 1570 1575

Leu Asp Ser Gly Lys Glu Leu Leu Met Gly Pro Thr Arg Gly Leu

1580 1585 1590

Leu Arg Ala Asp Asp Leu Arg Asp Ser Ile Val Ser Ser Val Lys

1595 1600 1605

Asp Val Leu Glu Leu Lys Ser Asn Glu Asp Leu Asp Leu Ser Glu

1610 1615 1620

Ser Leu Asn Ala Leu Gly Met Asp Ser Ile Met Phe Ala Gln Leu

1625 1630 1635

Arg Lys Arg Ile Gly Glu Gly Leu Gly Leu Asn Val Pro Met Val

1640 1645 1650

Phe Leu Ser Asp Ala Phe Ser Ile Gly Glu Met Val Ser Asn Leu

1655 1660 1665

Val Glu Gln Ala Glu Ala Ser Glu Asp Asn

1670 1675

<210> 36

<211> 4512

<212> DNA

<213> Panellus stipticus

<400> 36

atgcctcctg ctccttcctc catccttgat gtgttcaccg agactgcata caatcctcgc 60

acctccaacc gtcctgtagt agaatgtggc gagcacctct ggacatactc acaacttgaa 120

gcagtttcca atgccatgtc gcaagacctg gagaactcga tcggtcttta cgcgaaggtc 180

gcttttgtcg gtgaaaacca tccatttgtt ttcgctctca tgcttgcggt ctggaaaatt 240

gctggcacat tcatccccat tgatgcgcac attcctcccg ccctcctaga tggcatggtc 300

gacattgtga agccgatgcg catctatgta tcatcggccg atacctctaa ttcctcctgg 360

gcctcagaac tcgcggtcga gtccaaactc gcgtggtggc gacgcatgca gccaagcatc 420

aacctagaca acacccgagt cctcggctgg gccccgtggt cgcatgtatt atcccacatg 480

caggacattg gcaccgctac cattctcacc gcaggctgtt acgtctttgc ctctatccct 540

tccacatatc agcttcagca ggcgccaaca gatcttacga cccaagtcat caacggtatc 600

ctcaataaaa acatctcggc gcttgctgcc cttccatttg ttttcggcgg tatcaaagca 660

gcgtgcgagt cgggcgatct ggatgtggag gcacttctgg gtgccctgcg ccgcatgacg 720

atgctcgaat gtggcggggc tgcgctcgac cctgcaattg caaattggac ggatataaat 780

ggtgtatcgc tcatggttgg gattggaatg acggaaacgg gtggtgcaat cttcgcgggg 840

agggcgaaag actcgctctc cgggtttctt gctgaggggcc tgatctcaga tgccagaatt 900

gagcttgaca agggtaaatc tgatggctcg gatgagggag agcttgttgt cacaagcaag 960

ctacttccac atggctacat tggcttcgat gacgggtcat tttccgtgga tgcgcaaggg 1020

tgggtgacgt tcaagacaag ggattgctat cgtgtcaagg actcgaagtt tatttggcta 1080

ggccgggcca ctgattacat tcaggcgagc acatctctcc ttcctcagcg ctttaaacat 1140

gaactgcgct atcagatgac aagcggcgaa tcccttgacc cgcgtcccct cgaaaatttc 1200

ttacgctccg ccgagttcat ctccaatgtg tgcgtcatag gtgacaactt cctccggggt 1260

gcctccactt ctgtgtgggc cgttatcgag ctcacggaca gcgcgcaccg tctggatgcc 1320

gcttcagcga gaaagcaggt ggcgcgcgtg ctggcgccgc tcaatggtgg cctcccaccc 1380

gctctccgga tctcgatgtc ctcggtgttg atattgagcg gatcacagaa aattccgcgc 1440

acgaagaagg gcgaaatctt ccgtaagcag attgaggatc tgtttggtgc cgcgatgagc 1500

atgccgcctg aggctgagcc ggctctggag gatgaactaa caagtattgt tggaaatgtc 1560

ctcaacatat cggacagcga catgttgacc tcgatgactt ttgccgaact tggaatgacc 1620

tctgttctcg cagtaaaaat ctcagcaagg ctcaacgagc accttgctgg gcgggccgtc 1680

ctcccggcca acatctgtta catctacatc gatactccat cactgattgc cggcatccgg 1740

aacttccttt cgccggcatc ttctgatcaa gagccatcga actttgccac cacggcagac 1800

aagaaggacg aaatcgtcat cataggcaaa gccttccggc ttcccggcgg gatcaacgac 1860

gactctgctc tctgggctgc tcttatgggc aagaacgact ctgttattgc agacatcccg 1920

ccagaccgtt gggatcatgc tagcttctat cccgcccaca tctgcttgcg caaggccggg 1980

cttatagata tgtccagcta tgactatggg ttctttggcc tttcggcgac tgaggcgtac 2040

tatgtgtcgc ccaccatgcg cgcagcgctc gaggtggcgt tcgaagcgct ggagatgcg 2100

aacatcccgg tgtcaagaat caaggggaca aatacgagtg tttttgttgc aacgaaagat 2160

gatggatttg agacattgct gaatgcggcg catggatttg atgatgctga cggatacgtc 2220

ccttcagaag gcgccgtcgc ttttatactg aagacacgga cggcggcgga gagggatggg 2280

gatcgaatta tggccatcat caaagccacg gaagtctcgc ataatggaag atctcaggga 2340

ctcgccgcgc cgaacgtcaa ggcgcaagcc gctcttcata gagcagtctt gcgcaaagct 2400

aagctcgacc ctctggacat ccatttcatt gaagcccacg gaactggaac accgctgggc 2460

gatctttgtg aaatacaagg cataaatgaa gcctttgtct ccgcacgtcc acgtgcggag 2520

gatccactca ttgtcagcgc ctcaaagtct tcccttgggc acaccgaacc ttcggctgga 2580

ttggttggga tgctatctgt gttgatggcc ttgaagcacg gcatcgtgcc tgggttgctc 2640

catctccgag cggacaatgc taaccaccaa ctggacctga cacaagttcc acttcgtata 2700

tcaccggaac ctgtagtcat cgccgcatcc aagccgcatc acgccatggt gctatcctat 2760

ggattttcag gtacattggc agacattgtt ttggagagtc cggaagaacc atcgtcccca 2820

aacccgggag ccgcgggccc aatgatattt gtcctcagcg cgaagacttc cgcggctctg 2880

tcggcatata tcaaggctta cattgcattt ctacagaatg cagacgcgca cgagttttac 2940

aacatctgct acaccgcctg cgttggaaga gaacattaca aacacagatt tgcttgcgtt 3000

gcaaatgacc ttgctgatct gattcgccaa ttacaagact gtgcgagtgc gctggctcca 3060

accaaaagta gtactggtgc cttggcgttc gcgttccctg gccaaggcgt gcaatttcca 3120

ggcatggccg ctgcacttgc taaaaggcat tccatatttc gcgactatgt catggaattc 3180

ggtgatagag cacaagatct ttgtggcttg cccatcgcaa agatgttgct ggacgtggat 3240

gcagcagagg aagaagatat ccacagtgac gtagaccaga tttgcgtctt tgtctatcag 3300

tattcgatgt gtcgatggct tagggagctc gggctcgagg caagtgcggt tatcgggcac 3360

agtttgggtg aaataacggc cgcacttatc ggggacgcat ttacatttga agtggctctc 3420

gatctcgttg tcactcgagg acggctactc cgcccttctc aagggaatcc aggcgggatg 3480

gctgcgctgg catgccccga ggagaatgtc ccggcgattt tggaccaatg tcgtgtggac 3540

agtaccatta gcgtctccgt tatcaatggt ccgaggagcc tttgtgtatc cggagcttcc 3600

aacgacatcg acgagtttgt caagatggca aaacggcaaa acatcaaagc gactcgactg 3660

agggtggacc aaggatttca tagccctcga gttgatagtg ccgcggttgg actgcgtgca 3720

tggtcaggtt ctttttccaa atcatttcag ccgttgcgca tcacattata ctctacttct 3780

ctgggtgctg caatctcgaa aggagagatt ttgaatcaga cgcattgggc cgatcacgtc 3840

cgccgtccgg tcatattctc aaaagcagca gcagccatcc tcgaggacaa gtccatggc 3900

gcgatcctgg atatcggacc agacggtg gcatggtctc tccttctggc gaacggctgc 3960

aacgtcgcgt cagctgttgc cctgtccggc cgaagagtac aagatcagga aacagccttt 4020

ttatctgcac tggcgaatct gtatcaaaat cacggggtga cgccgaattt tcgcgtattt 4080

tatgctcacc aggcagtcca ggcgcgctat agaaccgtgg acatcccgaa gtatcccttc 4140

caacgccgac atcgatatcc atcctacatt ccatcgcgca atgccacggg tgccaacaga 4200

ctgaaagaac cattccgtag cgacctggat gaaccggctc aagacacgga gcacaccgcg 4260

gaactgagag tggacatgac tccggagaag ctccgggacg ccctgatgca ctgtgtgcgg 4320

gacacattgg aaggcgaaga ttttgatgaa tcggaatccc tcgtttcgcg tggaattgac 4380

tccattactt ttgcgggttt acggaagcgt gttcaagaac gacacggact taatctttcc 4440

atcattttct ggtctgatgg gttttctgtg aaagacatgg tcgacagcct catcgaacag 4500

cattttgtcc ac 4512

<210> 37

<211> 1504

<212>PRT

<213> Panellus stipticus

<400> 37

Met Pro Pro Ala Pro Ser Ser Ile Leu Asp Val Phe Thr Glu Thr Ala

1 5 10 15

Tyr Asn Pro Arg Thr Ser Asn Arg Pro Val Val Glu Cys Gly Glu His

20 25 30

Leu Trp Thr Tyr Ser Gln Leu Glu Ala Val Ser Asn Ala Met Ser Gln

35 40 45

Asp Leu Glu Asn Ser Ile Gly Leu Tyr Ala Lys Val Ala Phe Val Gly

50 55 60

Glu Asn His Pro Phe Val Phe Ala Leu Met Leu Ala Val Trp Lys Ile

65 70 75 80

Ala Gly Thr Phe Ile Pro Ile Asp Ala His Ile Pro Pro Ala Leu Leu

85 90 95

Asp Gly Met Val Asp Ile Val Lys Pro Met Arg Ile Tyr Val Ser Ser

100 105 110

Ala Asp Thr Ser Asn Ser Ser Trp Ala Ser Glu Leu Ala Val Glu Ser

115 120 125

Lys Leu Ala Trp Trp Arg Arg Met Gln Pro Ser Ile Asn Leu Asp Asn

130 135 140

Thr Arg Val Leu Gly Trp Ala Pro Trp Ser His Val Leu Ser His Met

145 150 155 160

Gln Asp Ile Gly Thr Ala Thr Ile Leu Thr Ala Gly Cys Tyr Val Phe

165 170 175

Ala Ser Ile Pro Ser Thr Tyr Gln Leu Gln Gln Ala Pro Thr Asp Leu

180 185 190

Thr Thr Gln Val Ile Asn Gly Ile Leu Asn Lys Asn Ile Ser Ala Leu

195 200 205

Ala Ala Leu Pro Phe Val Phe Gly Gly Ile Lys Ala Ala Cys Glu Ser

210 215 220

Gly Asp Leu Asp Val Glu Ala Leu Leu Gly Ala Leu Arg Arg Met Thr

225 230 235 240

Met Leu Glu Cys Gly Gly Ala Ala Leu Asp Pro Ala Ile Ala Asn Trp

245 250 255

Thr Asp Ile Asn Gly Val Ser Leu Met Val Gly Ile Gly Met Thr Glu

260 265 270

Thr Gly Gly Ala Ile Phe Ala Gly Arg Ala Lys Asp Ser Leu Ser Gly

275 280 285

Phe Leu Ala Glu Gly Leu Ile Ser Asp Ala Arg Ile Glu Leu Asp Lys

290 295 300

Gly Lys Ser Asp Gly Ser Asp Glu Gly Glu Leu Val Val Thr Ser Lys

305 310 315 320

Leu Leu Pro His Gly Tyr Ile Gly Phe Asp Asp Gly Ser Phe Ser Val

325 330 335

Asp Ala Gln Gly Trp Val Thr Phe Lys Thr Arg Asp Cys Tyr Arg Val

340 345 350

Lys Asp Ser Lys Phe Ile Trp Leu Gly Arg Ala Thr Asp Tyr Ile Gln

355 360 365

Ala Ser Thr Ser Leu Leu Pro Gln Arg Phe Lys His Glu Leu Arg Tyr

370 375 380

Gln Met Thr Ser Gly Glu Ser Leu Asp Pro Arg Pro Leu Glu Asn Phe

385 390 395 400

Leu Arg Ser Ala Glu Phe Ile Ser Asn Val Cys Val Ile Gly Asp Asn

405 410 415

Phe Leu Arg Gly Ala Ser Thr Ser Val Trp Ala Val Ile Glu Leu Thr

420 425 430

Asp Ser Ala His Arg Leu Asp Ala Ala Ser Ala Arg Lys Gln Val Ala

435 440 445

Arg Val Leu Ala Pro Leu Asn Gly Gly Leu Pro Pro Ala Leu Arg Ile

450 455 460

Ser Met Ser Ser Val Leu Ile Leu Ser Gly Ser Gln Lys Ile Pro Arg

465 470 475 480

Thr Lys Lys Gly Glu Ile Phe Arg Lys Gln Ile Glu Asp Leu Phe Gly

485 490 495

Ala Ala Met Ser Met Pro Pro Glu Ala Glu Pro Ala Leu Glu Asp Glu

500 505 510

Leu Thr Ser Ile Val Gly Asn Val Leu Asn Ile Ser Asp Ser Asp Met

515 520 525

Leu Thr Ser Met Thr Phe Ala Glu Leu Gly Met Thr Ser Val Leu Ala

530 535 540

Val Lys Ile Ser Ala Arg Leu Asn Glu His Leu Ala Gly Arg Ala Val

545 550 555 560

Leu Pro Ala Asn Ile Cys Tyr Ile Tyr Ile Asp Thr Pro Ser Leu Ile

565 570 575

Ala Gly Ile Arg Asn Phe Leu Ser Pro Ala Ser Ser Asp Gln Glu Pro

580 585 590

Ser Asn Phe Ala Thr Thr Ala Asp Lys Lys Asp Glu Ile Val Ile Ile

595 600 605

Gly Lys Ala Phe Arg Leu Pro Gly Gly Ile Asn Asp Asp Ser Ala Leu

610 615 620

Trp Ala Ala Leu Met Gly Lys Asn Asp Ser Val Ile Ala Asp Ile Pro

625 630 635 640

Pro Asp Arg Trp Asp His Ala Ser Phe Tyr Pro Ala His Ile Cys Leu

645 650 655

Arg Lys Ala Gly Leu Ile Asp Met Ser Ser Tyr Asp Tyr Gly Phe Phe

660 665 670

Gly Leu Ser Ala Thr Glu Ala Tyr Tyr Val Ser Pro Thr Met Arg Ala

675 680 685

Ala Leu Glu Val Ala Phe Glu Ala Leu Glu Asp Ala Asn Ile Pro Val

690 695 700

Ser Arg Ile Lys Gly Thr Asn Thr Ser Val Phe Val Ala Thr Lys Asp

705 710 715 720

Asp Gly Phe Glu Thr Leu Leu Asn Ala Ala His Gly Phe Asp Asp Ala

725 730 735

Asp Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys Thr

740 745 750

Arg Thr Ala Ala Glu Arg Asp Gly Asp Arg Ile Met Ala Ile Ile Lys

755 760 765

Ala Thr Glu Val Ser His Asn Gly Arg Ser Gln Gly Leu Ala Ala Pro

770 775 780

Asn Val Lys Ala Gln Ala Ala Leu His Arg Ala Val Leu Arg Lys Ala

785 790 795 800

Lys Leu Asp Pro Leu Asp Ile His Phe Ile Glu Ala His Gly Thr Gly

805 810 815

Thr Pro Leu Gly Asp Leu Cys Glu Ile Gln Gly Ile Asn Glu Ala Phe

820 825 830

Val Ser Ala Arg Pro Arg Ala Glu Asp Pro Leu Ile Val Ser Ala Ser

835 840 845

Lys Ser Ser Leu Gly His Thr Glu Pro Ser Ala Gly Leu Val Gly Met

850 855 860

Leu Ser Val Leu Met Ala Leu Lys His Gly Ile Val Pro Gly Leu Leu

865 870 875 880

His Leu Arg Ala Asp Asn Ala Asn His Gln Leu Asp Leu Thr Gln Val

885 890 895

Pro Leu Arg Ile Ser Pro Glu Pro Val Val Ile Ala Ala Ser Lys Pro

900 905 910

His His Ala Met Val Leu Ser Tyr Gly Phe Ser Gly Thr Leu Ala Asp

915 920 925

Ile Val Leu Glu Ser Pro Glu Glu Pro Ser Ser Pro Asn Pro Gly Ala

930 935 940

Ala Gly Pro Met Ile Phe Val Leu Ser Ala Lys Thr Ser Ala Ala Leu

945 950 955 960

Ser Ala Tyr Ile Lys Ala Tyr Ile Ala Phe Leu Gln Asn Ala Asp Ala

965 970 975

His Glu Phe Tyr Asn Ile Cys Tyr Thr Ala Cys Val Gly Arg Glu His

980 985 990

Tyr Lys His Arg Phe Ala Cys Val Ala Asn Asp Leu Ala Asp Leu Ile

995 1000 1005

Arg Gln Leu Gln Asp Cys Ala Ser Ala Leu Ala Pro Thr Lys Ser

1010 1015 1020

Ser Thr Gly Ala Leu Ala Phe Ala Phe Pro Gly Gln Gly Val Gln

1025 1030 1035

Phe Pro Gly Met Ala Ala Ala Leu Ala Lys Arg His Ser Ile Phe

1040 1045 1050

Arg Asp Tyr Val Met Glu Phe Gly Asp Arg Ala Gln Asp Leu Cys

1055 1060 1065

Gly Leu Pro Ile Ala Lys Met Leu Leu Asp Val Asp Ala Ala Glu

1070 1075 1080

Glu Glu Asp Ile His Ser Asp Val Asp Gln Ile Cys Val Phe Val

1085 1090 1095

Tyr Gln Tyr Ser Met Cys Arg Trp Leu Arg Glu Leu Gly Leu Glu

1100 1105 1110

Ala Ser Ala Val Ile Gly His Ser Leu Gly Glu Ile Thr Ala Ala

1115 1120 1125

Leu Ile Gly Asp Ala Phe Thr Phe Glu Val Ala Leu Asp Leu Val

1130 1135 1140

Val Thr Arg Gly Arg Leu Leu Arg Pro Ser Gln Gly Asn Pro Gly

1145 1150 1155

Gly Met Ala Ala Leu Ala Cys Pro Glu Glu Asn Val Pro Ala Ile

1160 1165 1170

Leu Asp Gln Cys Arg Val Asp Ser Thr Ile Ser Val Ser Val Ile

1175 1180 1185

Asn Gly Pro Arg Ser Leu Cys Val Ser Gly Ala Ser Asn Asp Ile

1190 1195 1200

Asp Glu Phe Val Lys Met Ala Lys Arg Gln Asn Ile Lys Ala Thr

1205 1210 1215

Arg Leu Arg Val Asp Gln Gly Phe His Ser Pro Arg Val Asp Ser

1220 1225 1230

Ala Ala Val Gly Leu Arg Ala Trp Ser Gly Ser Phe Ser Lys Ser

1235 1240 1245

Phe Gln Pro Leu Arg Ile Thr Leu Tyr Ser Thr Ser Leu Gly Ala

1250 1255 1260

Ala Ile Ser Lys Gly Glu Ile Leu Asn Gln Thr His Trp Ala Asp

1265 1270 1275

His Val Arg Arg Pro Val Ile Phe Ser Lys Ala Ala Ala Ala Ile

1280 1285 1290

Leu Glu Asp Lys Ser Ile Gly Ala Ile Leu Asp Ile Gly Pro Gln

1295 1300 1305

Thr Val Ala Trp Ser Leu Leu Leu Ala Asn Gly Cys Asn Val Ala

1310 1315 1320

Ser Ala Val Ala Leu Ser Gly Arg Arg Val Gln Asp Gln Glu Thr

1325 1330 1335

Ala Phe Leu Ser Ala Leu Ala Asn Leu Tyr Gln Asn His Gly Val

1340 1345 1350

Thr Pro Asn Phe Arg Val Phe Tyr Ala His Gln Ala Val Gln Ala

1355 1360 1365

Arg Tyr Arg Thr Val Asp Ile Pro Lys Tyr Pro Phe Gln Arg Arg

1370 1375 1380

His Arg Tyr Pro Ser Tyr Ile Pro Ser Arg Asn Ala Thr Gly Ala

1385 1390 1395

Asn Arg Leu Lys Glu Pro Phe Arg Ser Asp Leu Asp Glu Pro Ala

1400 1405 1410

Gln Asp Thr Glu His Thr Ala Glu Leu Arg Val Asp Met Thr Pro

1415 1420 1425

Glu Lys Leu Arg Asp Ala Leu Met His Cys Val Arg Asp Thr Leu

1430 1435 1440

Glu Gly Glu Asp Phe Asp Glu Ser Glu Ser Leu Val Ser Arg Gly

1445 1450 1455

Ile Asp Ser Ile Thr Phe Ala Gly Leu Arg Lys Arg Val Gln Glu

1460 1465 1470

Arg His Gly Leu Asn Leu Ser Ile Ile Phe Trp Ser Asp Gly Phe

1475 1480 1485

Ser Val Lys Asp Met Val Asp Ser Leu Ile Glu Gln His Phe Val

1490 1495 1500

His

<210> 38

<211> 4803

<212> DNA

<213> Panellus stipticus

<400> 38

atggcactgg tctctcccac ttgttcgtct cagatgcctc ctgctccttc ctccatcctt 60

gatgtgttca ccgagactgc atacaatcct cgcacctcca accgtcctgt agtagaatgt 120

ggcgagcacg tctggaccta ctcacaactt gaagcagttt ccaatgccat gtcgcaagac 180

ctggagaact cgatcggtct ttacgcgaag gtcgcttttg tcggtgaaaa ccatccatt 240

gttttcgctc tcatgcttgc agtctggaaa attgctggca cattcatccc cattgatgcg 300

cacattcctc ccgccctcct agatggcatg gtcgacattg tgaagccgat gtgcatctat 360

gtatcatcgg ccgatacctc taattcctcc tgggcctcag aactcgcggt cgaggttcgg 420

gtattccgtc cggaagaatc tacgattccg gctttgaacg aacactatgg aagatccagc 480

atcactcccg ctcagcatcg gccaatactc aacgtggtcc aggcagttcc cctgacacac 540

aaatttatcc tcagcaactg tcagtccaaa ctcgcgtggt ggcgacgcat gcagccaagc 600

atcaacctag acaacacccg agtcctcggc tgggccccgt ggtcgcatgt attatcccac 660

atgcaggaca ttggcaccgc taccattctc accgcaggct gttacgtctt tgcctctatc 720

ccttccacat atcagcttca gcaggcgcca acagatctta cgacccaagt catcaacggt 780

atcctcaata aaaacatctc ggcgcttgct gcccttccat ttgttttcgg cggtatcaaa 840

gcagcgtgcg agtcgggcga tctggatgtg gaggcacttc tgggtgccct gcgccgcatg 900

acgatgctcg aatgtggcgg ggctgcgctc gaccctgcaa ttgcaaattg gacggatata 960

aacggtgtat cgctcatggt tgggattgga atgacagaaa cgggcggtgc aatcttcgcg 1020

ggggagggcga aagactcgct ctccgggttt cttgctgagg gtctgatctc agacgccaga 1080

attgagcttg acaagggtga atctgatggc tcggatggta cgacgttctc catcttctcg 1140

accactgccg gaattaaacc tgattgtcag agggagagct tgttgtcaca agcaagctac 1200

ttccacatgg ctacattggc ttcgatgacg ggtcattttc cgtggatgcg caagggtgac 1260

aacttcctcc ggggtgcctc cacttctgtg tgggccatta tcgagctcac ggacagcgcg 1320

caccgcctgg atgccgcttc agcgagaaag caggtggcgc gcgtactggc gccgctcaat 1380

ggtggcctcc cacccgctct ccggatctca atgtcctcgg tgttgatatt gagcggatca 1440

cagaaaattc cgcgcacgaa gaagggcgag atcttccgta agcagattga ggatctgttt 1500

ggtgccgcga tgagcatgcc gcctgaggct gagccggctc tggaggatga actaacaagt 1560

attgttggaa atgtcctcaa catatcggac agcgacatgt tgacctcgat gacttttgcc 1620

gaacttggaa tgacctctgt tctggcagta aaaatctcag caaggctcaa cgagcacctt 1680

gctgggcggg ccgtcctccc ggccaacatc tgttacatct acatcgatac tccatcactg 1740

attgccggca tccggaactt cctttcgccg acatcttctg atcaagagcc atcgaacttt 1800

gccaccacgg cagacaagaa ggacgagatc gtcatcatag gcaaagcctt tcggcttccc 1860

ggcgggatca acgacgactc tgctctctgg gctgctctta tgggcaagaa cgactctgtt 1920

attgcagaca tcccgccaga ccgttgggat catgctagct tctatcccgc ccacatctgc 1980

ttgcgcaagg ctgggcttat agatatgtcc agctatgact atgggttctt tggcctttcg 2040

gcgactgagg cgtactatgt gtcgcccacc atgcgcgcag cgctcgaggt ggcgttcgaa 2100

gcgctggagg atgcgaacat cccggtgtca agaattaagg ggacaaatac gagtgttttt 2160

gttgcaacga aagatgatgg atttgagaca ttgctgaatg cggcgcatgg atttgatgcg 2220

tatacccggt tctacgggac tgggcgggct ccaagtactg ccagtgggcg cataagctac 2280

cttcttgaca tccatgggcc ctcactcaca gtagatacgg cctgtagcgg aggaattgtc 2340

tgcattgatc aagatatacc agcgaattta tgtatcatag cgatcgcata cctccagtct 2400

ggcgccggtg aatcagccat cgaacgacaa tacaggtttc tcacggcgca gaacatggca 2460

tcgcccacta gccgctgttc caccttcact gcagatgctg acggatatgt cccttcagaa 2520

ggcgccgtcg cttttatact gaagacacgg actgcggcgg agagggatgg ggatcgaatt 2580

atggccatca tcaaagccac agaagtctcg cataatggaa gatctcaggg actcgccgcg 2640

ccgaacgtca aagcgcaagc cgctcttcat agagcagtct tgcgcaaagc taagctcgac 2700

cctctggaca tccattcat tgaagcccac ggaactgttt taggaacacc gctgggtgat 2760

ctttgtgaaa tacaaggcat aaatgaagcc tttgtctccg cacgtccacg tgcggagaggat 2820

ccactcattg tcagcgcctc caagtcttcc cttgggcaca ccgaaccttc ggctggattg 2880

gttgggatgc tatctgtgtt gatggccttg aagcacggca tcgtgcctgg gttgctccat 2940

ctccgagcgg acaatgctaa ccaccaactg gacctgacac gagtcccact gcgtatatca 3000

ccggaacctg tagtcatcgc cgcatccaag ccgcatcacg ccatggtgct gctcacagtc 3060

cgtacattgg cagacattgt tttggagagt ccggaagaac caccgtccca gaacccggga 3120

gccgcgggcc caatgatatt tgtcctcagc gcgaagactt ccgcggctct gtcggcatat 3180

atcaaggctt acattgcatt tctacagaat gcagacgcgc acgagtttta caacatctgc 3240

tacaccgcct gtgttggaag agaacattac aaacacagat ttgcttgcgt tgcaaatgac 3300

cttgctgatc tgattcgcca attacaagac tgtgcgagtg cgctggctcc aaccaaaagt 3360

agtactggtg ccttggcgtt cgcgttccct ggccaaggcg tgcaatttcc aggcatggcc 3420

gctgcacttg ctaaaaggca ttccatattt cgcgactatg tcatggaatt tggtcataga 3480

gcacaagatc tttgtggctt gcccatcgca aagatgttgc tggacgtgga tgcagcagag 3540

gaagaagata tccacagtga cgtagaccag atttgcgtct ttgtctatca gtattcgatg 3600

tgtcgatggc ttagggagct cgggctcgag gcaagtgcgg ttatcgggca cagtttgggc 3660

gaaataacag ccgcacttat cggggacgca tttacatttg aagtggctct cgatctcgtt 3720

gtcactcgag gacggctact ccgcccttct caagggaatc caggcgggat ggctgcgctg 3780

gcatgccccg aggagaatgt cccagcgatt ttggaccaat gtcgtgtgga cagtaccatt 3840

agcgtctccg ttatcaatgg tccgaggagc ctttgtgtat ccggagcttc caacgacatc 3900

gacgagtttg tcaagatggc aaaacggcaa aacatcaaag cgactcgact gagggtggac 3960

caaggatttc atagccctcg agttgatagt gccgcggttg gactgcgtga atggtcaggt 4020

tctttttcga aatcatttca gccgttgcgc atcacattat actctacgtc tctgggtgct 4080

gcaatctcga aaggagagat tttgactcag acacattggg ccgatcacgt ccgccgtccg 4140

gtcatattct caaaagcagc agcagccatc ctcgaggaca agtccattgg cgcgatcctg 4200

gatatcggac cacagacggt ggcatggtct ctccttctgg cgaacggctg caacgtcgcg 4260

tcagctgttg ccctgtccgg ccgaagagta caagatcagg aaacggcctt tttatctgca 4320

ctggcgaatc tgtatcaaaa tcacggggtg acgccgaatt ttcgcgtatt ttacgctcac 4380

caggcagtcc aggcgcgcta tagaaccgtg gacattccga agtatccctt ccaacgccga 4440

catcgatatc catcctacat tccatcgcgc aatgccacgg gtgccaacag actgaaagaa 4500

ccattccgta gcgacctgga tgaaccggct caagacacgg agcacaccgc ggaaatgaga 4560

gtggacatga ctccggagaa gctccgggac gccctgatgc actgtgtgcg ggacacactg 4620

gaaggcgaag attttgatga atcggaatcc ctcgtttcgc gtggaattga ctccattact 4680

tttgcgggtc tacggaagcg tgttcaagaa cgacacggac ttaatctttc catcattttc 4740

tggtctgatg ggttttctgt gaaagacatg gtcgacagcc tcatcgaaca gcattttgtc 4800

ca.4803

<210> 39

<211> 1601

<212>PRT

<213> Panellus stipticus

<400> 39

Met Ala Leu Val Ser Pro Thr Cys Ser Ser Gln Met Pro Pro Ala Pro

1 5 10 15

Ser Ser Ile Leu Asp Val Phe Thr Glu Thr Ala Tyr Asn Pro Arg Thr

20 25 30

Ser Asn Arg Pro Val Val Glu Cys Gly Glu His Val Trp Thr Tyr Ser

35 40 45

Gln Leu Glu Ala Val Ser Asn Ala Met Ser Gln Asp Leu Glu Asn Ser

50 55 60

Ile Gly Leu Tyr Ala Lys Val Ala Phe Val Gly Glu Asn His Pro Phe

65 70 75 80

Val Phe Ala Leu Met Leu Ala Val Trp Lys Ile Ala Gly Thr Phe Ile

85 90 95

Pro Ile Asp Ala His Ile Pro Pro Ala Leu Leu Asp Gly Met Val Asp

100 105 110

Ile Val Lys Pro Met Cys Ile Tyr Val Ser Ser Ala Asp Thr Ser Asn

115 120 125

Ser Ser Trp Ala Ser Glu Leu Ala Val Glu Val Arg Val Phe Arg Pro

130 135 140

Glu Glu Ser Thr Ile Pro Ala Leu Asn Glu His Tyr Gly Arg Ser Ser

145 150 155 160

Ile Thr Pro Ala Gln His Arg Pro Ile Leu Asn Val Val Gln Ala Val

165 170 175

Pro Leu Thr His Lys Phe Ile Leu Ser Asn Cys Gln Ser Lys Leu Ala

180 185 190

Trp Trp Arg Arg Met Gln Pro Ser Ile Asn Leu Asp Asn Thr Arg Val

195 200 205

Leu Gly Trp Ala Pro Trp Ser His Val Leu Ser His Met Gln Asp Ile

210 215 220

Gly Thr Ala Thr Ile Leu Thr Ala Gly Cys Tyr Val Phe Ala Ser Ile

225 230 235 240

Pro Ser Thr Tyr Gln Leu Gln Gln Ala Pro Thr Asp Leu Thr Thr Gln

245 250 255

Val Ile Asn Gly Ile Leu Asn Lys Asn Ile Ser Ala Leu Ala Ala Leu

260 265 270

Pro Phe Val Phe Gly Gly Ile Lys Ala Ala Cys Glu Ser Gly Asp Leu

275 280 285

Asp Val Glu Ala Leu Leu Gly Ala Leu Arg Arg Met Thr Met Leu Glu

290 295 300

Cys Gly Gly Ala Ala Leu Asp Pro Ala Ile Ala Asn Trp Thr Asp Ile

305 310 315 320

Asn Gly Val Ser Leu Met Val Gly Ile Gly Met Thr Glu Thr Gly Gly

325 330 335

Ala Ile Phe Ala Gly Arg Ala Lys Asp Ser Leu Ser Gly Phe Leu Ala

340 345 350

Glu Gly Leu Ile Ser Asp Ala Arg Ile Glu Leu Asp Lys Gly Glu Ser

355 360 365

Asp Gly Ser Asp Gly Thr Thr Phe Ser Ile Phe Ser Thr Thr Ala Gly

370 375 380

Ile Lys Pro Asp Cys Gln Arg Glu Ser Leu Leu Ser Gln Ala Ser Tyr

385 390 395 400

Phe His Met Ala Thr Leu Ala Ser Met Thr Gly His Phe Pro Trp Met

405 410 415

Arg Lys Gly Asp Asn Phe Leu Arg Gly Ala Ser Thr Ser Val Trp Ala

420 425 430

Ile Ile Glu Leu Thr Asp Ser Ala His Arg Leu Asp Ala Ala Ser Ala

435 440 445

Arg Lys Gln Val Ala Arg Val Leu Ala Pro Leu Asn Gly Gly Leu Pro

450 455 460

Pro Ala Leu Arg Ile Ser Met Ser Ser Val Leu Ile Leu Ser Gly Ser

465 470 475 480

Gln Lys Ile Pro Arg Thr Lys Lys Gly Glu Ile Phe Arg Lys Gln Ile

485 490 495

Glu Asp Leu Phe Gly Ala Ala Met Ser Met Pro Pro Glu Ala Glu Pro

500 505 510

Ala Leu Glu Asp Glu Leu Thr Ser Ile Val Gly Asn Val Leu Asn Ile

515 520 525

Ser Asp Ser Asp Met Leu Thr Ser Met Thr Phe Ala Glu Leu Gly Met

530 535 540

Thr Ser Val Leu Ala Val Lys Ile Ser Ala Arg Leu Asn Glu His Leu

545 550 555 560

Ala Gly Arg Ala Val Leu Pro Ala Asn Ile Cys Tyr Ile Tyr Ile Asp

565 570 575

Thr Pro Ser Leu Ile Ala Gly Ile Arg Asn Phe Leu Ser Pro Thr Ser

580 585 590

Ser Asp Gln Glu Pro Ser Asn Phe Ala Thr Thr Ala Asp Lys Lys Asp

595 600 605

Glu Ile Val Ile Ile Gly Lys Ala Phe Arg Leu Pro Gly Gly Ile Asn

610 615 620

Asp Asp Ser Ala Leu Trp Ala Ala Leu Met Gly Lys Asn Asp Ser Val

625 630 635 640

Ile Ala Asp Ile Pro Pro Asp Arg Trp Asp His Ala Ser Phe Tyr Pro

645 650 655

Ala His Ile Cys Leu Arg Lys Ala Gly Leu Ile Asp Met Ser Ser Tyr

660 665 670

Asp Tyr Gly Phe Phe Gly Leu Ser Ala Thr Glu Ala Tyr Tyr Val Ser

675 680 685

Pro Thr Met Arg Ala Ala Leu Glu Val Ala Phe Glu Ala Leu Glu Asp

690 695 700

Ala Asn Ile Pro Val Ser Arg Ile Lys Gly Thr Asn Thr Ser Val Phe

705 710 715 720

Val Ala Thr Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Ala His

725 730 735

Gly Phe Asp Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser

740 745 750

Thr Ala Ser Gly Arg Ile Ser Tyr Leu Leu Asp Ile His Gly Pro Ser

755 760 765

Leu Thr Val Asp Thr Ala Cys Ser Gly Gly Ile Val Cys Ile Asp Gln

770 775 780

Asp Ile Pro Ala Asn Leu Cys Ile Ile Ala Ile Ala Tyr Leu Gln Ser

785 790 795 800

Gly Ala Gly Glu Ser Ala Ile Glu Arg Gln Tyr Arg Phe Leu Thr Ala

805 810 815

Gln Asn Met Ala Ser Pro Thr Ser Arg Cys Ser Thr Phe Thr Ala Asp

820 825 830

Ala Asp Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys

835 840 845

Thr Arg Thr Ala Ala Glu Arg Asp Gly Asp Arg Ile Met Ala Ile Ile

850 855 860

Lys Ala Thr Glu Val Ser His Asn Gly Arg Ser Gln Gly Leu Ala Ala

865 870 875 880

Pro Asn Val Lys Ala Gln Ala Ala Leu His Arg Ala Val Leu Arg Lys

885 890 895

Ala Lys Leu Asp Pro Leu Asp Ile His Phe Ile Glu Ala His Gly Thr

900 905 910

Val Leu Gly Thr Pro Leu Gly Asp Leu Cys Glu Ile Gln Gly Ile Asn

915 920 925

Glu Ala Phe Val Ser Ala Arg Pro Arg Ala Glu Asp Pro Leu Ile Val

930 935 940

Ser Ala Ser Lys Ser Ser Leu Gly His Thr Glu Pro Ser Ala Gly Leu

945 950 955 960

Val Gly Met Leu Ser Val Leu Met Ala Leu Lys His Gly Ile Val Pro

965 970 975

Gly Leu Leu His Leu Arg Ala Asp Asn Ala Asn His Gln Leu Asp Leu

980 985 990

Thr Arg Val Pro Leu Arg Ile Ser Pro Glu Pro Val Val Ile Ala Ala

995 1000 1005

Ser Lys Pro His His Ala Met Val Leu Leu Thr Val Arg Thr Leu

1010 1015 1020

Ala Asp Ile Val Leu Glu Ser Pro Glu Glu Pro Pro Ser Gln Asn

1025 1030 1035

Pro Gly Ala Ala Gly Pro Met Ile Phe Val Leu Ser Ala Lys Thr

1040 1045 1050

Ser Ala Ala Leu Ser Ala Tyr Ile Lys Ala Tyr Ile Ala Phe Leu

1055 1060 1065

Gln Asn Ala Asp Ala His Glu Phe Tyr Asn Ile Cys Tyr Thr Ala

1070 1075 1080

Cys Val Gly Arg Glu His Tyr Lys His Arg Phe Ala Cys Val Ala

1085 1090 1095

Asn Asp Leu Ala Asp Leu Ile Arg Gln Leu Gln Asp Cys Ala Ser

1100 1105 1110

Ala Leu Ala Pro Thr Lys Ser Ser Thr Gly Ala Leu Ala Phe Ala

1115 1120 1125

Phe Pro Gly Gln Gly Val Gln Phe Pro Gly Met Ala Ala Ala Leu

1130 1135 1140

Ala Lys Arg His Ser Ile Phe Arg Asp Tyr Val Met Glu Phe Gly

1145 1150 1155

His Arg Ala Gln Asp Leu Cys Gly Leu Pro Ile Ala Lys Met Leu

1160 1165 1170

Leu Asp Val Asp Ala Ala Glu Glu Glu Asp Ile His Ser Asp Val

1175 1180 1185

Asp Gln Ile Cys Val Phe Val Tyr Gln Tyr Ser Met Cys Arg Trp

1190 1195 1200

Leu Arg Glu Leu Gly Leu Glu Ala Ser Ala Val Ile Gly His Ser

1205 1210 1215

Leu Gly Glu Ile Thr Ala Ala Leu Ile Gly Asp Ala Phe Thr Phe

1220 1225 1230

Glu Val Ala Leu Asp Leu Val Val Thr Arg Gly Arg Leu Leu Arg

1235 1240 1245

Pro Ser Gln Gly Asn Pro Gly Gly Met Ala Ala Leu Ala Cys Pro

1250 1255 1260

Glu Glu Asn Val Pro Ala Ile Leu Asp Gln Cys Arg Val Asp Ser

1265 1270 1275

Thr Ile Ser Val Ser Val Ile Asn Gly Pro Arg Ser Leu Cys Val

1280 1285 1290

Ser Gly Ala Ser Asn Asp Ile Asp Glu Phe Val Lys Met Ala Lys

1295 1300 1305

Arg Gln Asn Ile Lys Ala Thr Arg Leu Arg Val Asp Gln Gly Phe

1310 1315 1320

His Ser Pro Arg Val Asp Ser Ala Ala Val Gly Leu Arg Glu Trp

1325 1330 1335

Ser Gly Ser Phe Ser Lys Ser Phe Gln Pro Leu Arg Ile Thr Leu

1340 1345 1350

Tyr Ser Thr Ser Leu Gly Ala Ala Ile Ser Lys Gly Glu Ile Leu

1355 1360 1365

Thr Gln Thr His Trp Ala Asp His Val Arg Arg Pro Val Ile Phe

1370 1375 1380

Ser Lys Ala Ala Ala Ala Ile Leu Glu Asp Lys Ser Ile Gly Ala

1385 1390 1395

Ile Leu Asp Ile Gly Pro Gln Thr Val Ala Trp Ser Leu Leu Leu

1400 1405 1410

Ala Asn Gly Cys Asn Val Ala Ser Ala Val Ala Leu Ser Gly Arg

1415 1420 1425

Arg Val Gln Asp Gln Glu Thr Ala Phe Leu Ser Ala Leu Ala Asn

1430 1435 1440

Leu Tyr Gln Asn His Gly Val Thr Pro Asn Phe Arg Val Phe Tyr

1445 1450 1455

Ala His Gln Ala Val Gln Ala Arg Tyr Arg Thr Val Asp Ile Pro

1460 1465 1470

Lys Tyr Pro Phe Gln Arg Arg His Arg Tyr Pro Ser Tyr Ile Pro

1475 1480 1485

Ser Arg Asn Ala Thr Gly Ala Asn Arg Leu Lys Glu Pro Phe Arg

1490 1495 1500

Ser Asp Leu Asp Glu Pro Ala Gln Asp Thr Glu His Thr Ala Glu

1505 1510 1515

Met Arg Val Asp Met Thr Pro Glu Lys Leu Arg Asp Ala Leu Met

1520 1525 1530

His Cys Val Arg Asp Thr Leu Glu Gly Glu Asp Phe Asp Glu Ser

1535 1540 1545

Glu Ser Leu Val Ser Arg Gly Ile Asp Ser Ile Thr Phe Ala Gly

1550 1555 1560

Leu Arg Lys Arg Val Gln Glu Arg His Gly Leu Asn Leu Ser Ile

1565 1570 1575

Ile Phe Trp Ser Asp Gly Phe Ser Val Lys Asp Met Val Asp Ser

1580 1585 1590

Leu Ile Glu Gln His Phe Val His

1595 1600

<210> 40

<211> 5073

<212> DNA

<213> Neonothopanus gardneri

<400> 40

atgaattcca aagtgaatct tccctccact ttgcttgatg ttttcctcga gatcgctggc 60

gagccatcta ccgcttcgcg tgatgttttg gaatgtggcg agcacagatg gacttaccag 120

cagcttgacg ttgtttcatc tgctttagcc cagcatctca agtacactgt cggtctatct 180

cctacagtcg cagtgatcag cgaaaatcat ccttatgttt ttgccttgat cctggctgtg 240

tggaaagttg ggggcatttt cgcgcccctc gacgcacatg ctcctgctga gttggtagct 300

ggcatgttaa gcataatctc tccttcgtgc ttagtacttc agagcacaga tgtagctaat 360

caaactcttg cgtgtgatct cgatattcct gttgaggtat tccactcgcg tcaatccact 420

attcctgaac taaacaagaa atatctcacc gattctgggt taccggcggg ttttccgctc 480

tcagattcaa acaaaccggc tctgtatctc ttcacctcgt ctgccacttc tcggagcaat 540

cttaaatgcg tgcctctcac tcacgctttc atcctgagca atagcctctc gaaacgcgca 600

tggtgccaac gtatgcggcc agagacagac ttcgatggca tacgcgttct tggatgggct 660

ccatggtctc atgtcttggc gcacatgcag gacatcgggc ccgtcacttt actcaatgct 720

ggatgctacg tctttgctac tatcccttcc tcgtacccca cggatgtgca gagtgacagc 780

aatttgatat ctcatgtcgc aaatgctatc atacacaagg gtgtaaaatc gtttgcttgt 840

ctcccttttg tactcggagg gctgaaagca ttatgcgagt ccaagccatc cgtcaaagca 900

gatctacagg tcgaagagca agctcagctt ttgatctctc taaggcgcat ggaaattctt 960

gaatgcggag gtgcgatgct cgaagcgaat gttgcgtctt gggctatcga gaatcatatt 1020

ccggtctcta ttggtatcgg tatgacagaa actggtggcg cgcttttcgc tggacctgtt 1080

caggatatcc ggaccggttt ttctgcggac aataaattca tcaaggacgc tacctacttc 1140

ctagttgcaa atggagatga atctgggaac gatgtaactg aaggggagct agttgtgaaa 1200

agtaaaatgc tcccgcgagg ctatcttggc tacaatgatt cttccttttc cgttgacgat 1260

gacggattgg ttacgttcaa gacaggcgac agatacagtg ttacacgcga cggaagattc 1320

tcttggctag gcaggaatac tgatttcatt cagatgacta gtggagagac attagatccc 1380

cgacctgtcg agagttcgct ctgccaaagt cctctcatct ctcaagcatg cgttattgga 1440

gacaagtttc tcaacgggcc tgccactgct gtttgcgcga tcgttgagct tgagccgaca 1500

atggtagaaa cgggagaggc taactcgcgg gacatagtcc aagtctttgc acccatcaac 1560

cgagacctgc ctcctccttt aaggatagca tggtcgcaca ttttaattct caagcattcc 1620

gagaagatac cgatgacgaa gaagggaacg attttccgta agaagattga gcagatgttt 1680

ggcgctgcat tgggcattgc ttccatccct acaaacagtg tcaatggatc cgttggctcc 1740

ggagaagagc ctcaagctac tgcagatgag actgttctac aagatgaact atcaaatacc 1800

gtgaagaaaa taatcagccg tgttctagga gtaactgatg aggaattgct ttggacactg 1860

tcattcgcag agctgggaat gacctcaaca ctggctattc gtatcgtaaa cgagctgaac 1920

gaaactctcg ttggggggcga tctccctatc aatgcttgct acatttatgt cgacctttcc 1980

tctctgagca aggctgtgta tgcgaaactg gctcatctcg agctgtcaga tcatgttcct 2040

gagctcaaac aagctccctt caagtcttct ggcgggaaag agattgtcat tgtcggccaa 2100

gcgtttcgtc tacccggctc aatcaatgac gttgcctctc ttcgggatgc attcctagca 2160

agacagacat cgtccatcat cgctggaata ccttccgatc gctgggacca tgccagcttc 2220

tatcccaagg acatatgttt cgacaaggct ggtcttgtgg atatagctca ttatgatcat 2280

agcttcttcg gaatcacagc aacggaagcg ctccatctgt ctccaaccat gcgtcttgca 2340

ctggaagttt cgtttgaggc gcttgagaat gctaatatcc caacgtcgcg attgaaaggc 2400

tcgcagacgg cagtttatgt tgcaacaaca gatgacggat ttgagaccct gttgaatgct 2460

gaggctggct atgaagctta tacacggttt tatggtactg gtcgggcagc aagtacggcg 2520

agtgggcgca taagctatct tcttgatata catggaccct ccgtcactat tgacacggca 2580

tgcagtggtg gagttgtttg tatcgatcaa gcaatcaact atttacaatc gtcgagttca 2640

gcggacaccg ctatcgtgtt cctttctgct caagggatgg tctccccgag gggacgatgt 2700

gcgacattta cggctgatgc tgacggctat gtcccttctg agggtgcagt tgctttcgta 2760

ttgaaaactc gcgaagccgc tatacgcgac aaagacaaca ttctcgctac aatcaaagcg 2820

acacagattt cgcacaacgg ccgctcacaa ggacttgtag caccgaacgt aaactcgcaa 2880

gttgaccttc atcgttcgtt gctcgagaaa gctcgtctta gtcctgctga tgtccaattc 2940

attgaagctc atgggacagg aacgtcattg ggagatctct cagagattca agctataaac 3000

gctgcttaca cttcctctca gccacgtacg accggcccac tcatagtcag cgcttccaag 3060

actgtcgttg gccataccga accagctggt cctttggttg gtatgttgtc ggtcttgctg 3120

tctttccagg aaggcgcagt ccctggtctc gctcatctta ccacacgtaa cctcaatcct 3180

acgttggact attcttcagt gccgcttctc attccctctg aacctgttcg tctacaaaca 3240

ccaaagcctt atagagctgc cgtaatgtcc tacggctttt cgggtaccct ggccggcctc 3300

gttctagaga gccctgacga acgtagctca gaagaagagc cgcctgatga caagccgatg 3360

ctgttcgtcg ttagcgcaaa gacacacacg gcactaattg aatacctgca acggtatctc 3420

gagttcctct tacatgcgaa tccccgcgat ttctgtgata tctgctacac aagctgtgtc 3480

gggagggaac actatagata tcggtttgct tgtgttgcaa atgacatgga agacctcatt 3540

ggccaacttc aaagacgttt gtctagcaag ttaccatcga agccgctata caaacgcggt 3600

gctctggcct ttgcgttttc tggacagggg acacagtttc aaggaatggc gacagatctt 3660

gcgaagaggt actctggctt ccgaaaaatc gtttccggac tcgcaaagag tgctggcgag 3720

ctctcgggtt acgctattga ccgttatctt ctcgcgtatg acgttggtag cagtattgct 3780

actcctaata gtgaggtgga ccagatctgc attttcgtat accaatgctc tgtccttcgc 3840

tggttgggga gtattggaat taaaccaaac gtggtaatcg gccatagcct tggagagatt 3900

tcggcttctg tggcggcagg ggcactttct cttgacattg ctctggatct tgtcatctca 3960

cgagctgggt tgttgcgccc ctcgacagat gttcctgcgg gaatggctgc tgtggccgct 4020

tcacaacagg aggtcattga gttgattgat gcgctggacc ccgacaaggc aaattcgctc 4080

agtgtttcgg ttataaatgg acctcaaaat atcgttgtgt caggcgcttc agcagctatt 4140

gagagaatgg tcgcttctgc gaaggagaag aagatcaaag cttctgttct gaatattagc 4200

caagcttttc acagttcgta tgttgacagt gccattacgg gtcttcgagc ttggtcagaa 4260

aaacatattt cctcagcgat accactgcag attccgctgt attcgacatt gctgggagct 4320

cggatatcaa agggccaaaa actgaaccca gaccactggg tcgaccacgc acggaagccc 4380

gtacagttcg cacaagcagc tacggcgatg aaagaaacct tcaccggagt catcatggat 4440

atcggacccc aagcagtagc atggtcactt ctgcttgcaa acggactcac atccgtaacc 4500

gcgcttgctg cgaagagagg gaggagtcag caggtggctt tcttgagcgc cttggcggag 4560

ttgtatcagg attatggcat tgttcctgat tttgtggcgc tttatgctca ggagggaggaa 4620

atagacaagt tgaggaagac ggatattttg acatatccgt ttcagcgtgt taggaggtat 4680

ccgagtttta taccttcaag gcgtgtgaga ggtggagaaa ttccttcgag ccatcccagc 4740

gagtctgcga cgttggagaa cacgaatgag ggtacggctt tgcgtgctga gtcgaggtg 4800

ttgtgcaggg aggatctgca tgatagcatc gttacctctg tgaaggatgt tctagagctc 4860

aaaccaaatg aagatctaga tttgtctgaa agcctgaacg cgcttggtgt ggactctata 4920

atgtttgctc agttaaggaa acgtattggg gagggactcg ggttgagtat cccgacagtg 4980

ttcctttcgg atgccttttc tattaatgag atggttaata atcttatgga acaggcggag 5040

acgcctggtg aagagggcgt aatgcaggag aat 5073

<210> 41

<211> 1691

<212>PRT

<213> Neonothopanus gardneri

<400> 41

Met Asn Ser Lys Val Asn Leu Pro Ser Thr Leu Leu Asp Val Phe Leu

1 5 10 15

Glu Ile Ala Gly Glu Pro Ser Thr Ala Ser Arg Asp Val Leu Glu Cys

20 25 30

Gly Glu His Arg Trp Thr Tyr Gln Gln Leu Asp Val Val Ser Ser Ala

35 40 45

Leu Ala Gln His Leu Lys Tyr Thr Val Gly Leu Ser Pro Thr Val Ala

50 55 60

Val Ile Ser Glu Asn His Pro Tyr Val Phe Ala Leu Ile Leu Ala Val

65 70 75 80

Trp Lys Val Gly Gly Ile Phe Ala Pro Leu Asp Ala His Ala Pro Ala

85 90 95

Glu Leu Val Ala Gly Met Leu Ser Ile Ile Ser Pro Ser Cys Leu Val

100 105 110

Leu Gln Ser Thr Asp Val Ala Asn Gln Thr Leu Ala Cys Asp Leu Asp

115 120 125

Ile Pro Val Glu Val Phe His Ser Arg Gln Ser Thr Ile Pro Glu Leu

130 135 140

Asn Lys Lys Tyr Leu Thr Asp Ser Gly Leu Pro Ala Gly Phe Pro Leu

145 150 155 160

Ser Asp Ser Asn Lys Pro Ala Leu Tyr Leu Phe Thr Ser Ser Ala Thr

165 170 175

Ser Arg Ser Asn Leu Lys Cys Val Pro Leu Thr His Ala Phe Ile Leu

180 185 190

Ser Asn Ser Leu Ser Lys Arg Ala Trp Cys Gln Arg Met Arg Pro Glu

195 200 205

Thr Asp Phe Asp Gly Ile Arg Val Leu Gly Trp Ala Pro Trp Ser His

210 215 220

Val Leu Ala His Met Gln Asp Ile Gly Pro Val Thr Leu Leu Asn Ala

225 230 235 240

Gly Cys Tyr Val Phe Ala Thr Ile Pro Ser Ser Tyr Pro Thr Asp Val

245 250 255

Gln Ser Asp Ser Asn Leu Ile Ser His Val Ala Asn Ala Ile Ile His

260 265 270

Lys Gly Val Lys Ser Phe Ala Cys Leu Pro Phe Val Leu Gly Gly Leu

275 280 285

Lys Ala Leu Cys Glu Ser Lys Pro Ser Val Lys Ala Asp Leu Gln Val

290 295 300

Glu Glu Gln Ala Gln Leu Leu Ile Ser Leu Arg Arg Met Glu Ile Leu

305 310 315 320

Glu Cys Gly Gly Ala Met Leu Glu Ala Asn Val Ala Ser Trp Ala Ile

325 330 335

Glu Asn His Ile Pro Val Ser Ile Gly Ile Gly Met Thr Glu Thr Gly

340 345 350

Gly Ala Leu Phe Ala Gly Pro Val Gln Asp Ile Arg Thr Gly Phe Ser

355 360 365

Ala Asp Asn Lys Phe Ile Lys Asp Ala Thr Tyr Phe Leu Val Ala Asn

370 375 380

Gly Asp Glu Ser Gly Asn Asp Val Thr Glu Gly Glu Leu Val Val Lys

385 390 395 400

Ser Lys Met Leu Pro Arg Gly Tyr Leu Gly Tyr Asn Asp Ser Ser Phe

405 410 415

Ser Val Asp Asp Asp Gly Leu Val Thr Phe Lys Thr Gly Asp Arg Tyr

420 425 430

Ser Val Thr Arg Asp Gly Arg Phe Ser Trp Leu Gly Arg Asn Thr Asp

435 440 445

Phe Ile Gln Met Thr Ser Gly Glu Thr Leu Asp Pro Arg Pro Val Glu

450 455 460

Ser Ser Leu Cys Gln Ser Pro Leu Ile Ser Gln Ala Cys Val Ile Gly

465 470 475 480

Asp Lys Phe Leu Asn Gly Pro Ala Thr Ala Val Cys Ala Ile Val Glu

485 490 495

Leu Glu Pro Thr Met Val Glu Thr Gly Glu Ala Asn Ser Arg Asp Ile

500 505 510

Val Gln Val Phe Ala Pro Ile Asn Arg Asp Leu Pro Pro Pro Leu Arg

515 520 525

Ile Ala Trp Ser His Ile Leu Ile Leu Lys His Ser Glu Lys Ile Pro

530 535 540

Met Thr Lys Lys Gly Thr Ile Phe Arg Lys Lys Ile Glu Gln Met Phe

545 550 555 560

Gly Ala Ala Leu Gly Ile Ala Ser Ile Pro Thr Asn Ser Val Asn Gly

565 570 575

Ser Val Gly Ser Gly Glu Glu Pro Gln Ala Thr Ala Asp Glu Thr Val

580 585 590

Leu Gln Asp Glu Leu Ser Asn Thr Val Lys Lys Ile Ile Ser Arg Val

595 600 605

Leu Gly Val Thr Asp Glu Glu Leu Leu Trp Thr Leu Ser Phe Ala Glu

610 615 620

Leu Gly Met Thr Ser Thr Leu Ala Ile Arg Ile Val Asn Glu Leu Asn

625 630 635 640

Glu Thr Leu Val Gly Gly Asp Leu Pro Ile Asn Ala Cys Tyr Ile Tyr

645 650 655

Val Asp Leu Ser Ser Leu Ser Lys Ala Val Tyr Ala Lys Leu Ala His

660 665 670

Leu Glu Leu Ser Asp His Val Pro Glu Leu Lys Gln Ala Pro Phe Lys

675 680 685

Ser Ser Gly Gly Lys Glu Ile Val Ile Val Gly Gln Ala Phe Arg Leu

690 695 700

Pro Gly Ser Ile Asn Asp Val Ala Ser Leu Arg Asp Ala Phe Leu Ala

705 710 715 720

Arg Gln Thr Ser Ser Ile Ile Ala Gly Ile Pro Ser Asp Arg Trp Asp

725 730 735

His Ala Ser Phe Tyr Pro Lys Asp Ile Cys Phe Asp Lys Ala Gly Leu

740 745 750

Val Asp Ile Ala His Tyr Asp His Ser Phe Phe Gly Ile Thr Ala Thr

755 760 765

Glu Ala Leu His Leu Ser Pro Thr Met Arg Leu Ala Leu Glu Val Ser

770 775 780

Phe Glu Ala Leu Glu Asn Ala Asn Ile Pro Thr Ser Arg Leu Lys Gly

785 790 795 800

Ser Gln Thr Ala Val Tyr Val Ala Thr Thr Asp Asp Gly Phe Glu Thr

805 810 815

Leu Leu Asn Ala Glu Ala Gly Tyr Glu Ala Tyr Thr Arg Phe Tyr Gly

820 825 830

Thr Gly Arg Ala Ala Ser Thr Ala Ser Gly Arg Ile Ser Tyr Leu Leu

835 840 845

Asp Ile His Gly Pro Ser Val Thr Ile Asp Thr Ala Cys Ser Gly Gly

850 855 860

Val Val Cys Ile Asp Gln Ala Ile Asn Tyr Leu Gln Ser Ser Ser Ser

865 870 875 880

Ala Asp Thr Ala Ile Val Phe Leu Ser Ala Gln Gly Met Val Ser Pro

885 890 895

Arg Gly Arg Cys Ala Thr Phe Thr Ala Asp Ala Asp Gly Tyr Val Pro

900 905 910

Ser Glu Gly Ala Val Ala Phe Val Leu Lys Thr Arg Glu Ala Ala Ile

915 920 925

Arg Asp Lys Asp Asn Ile Leu Ala Thr Ile Lys Ala Thr Gln Ile Ser

930 935 940

His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn Val Asn Ser Gln

945 950 955 960

Val Asp Leu His Arg Ser Leu Leu Glu Lys Ala Arg Leu Ser Pro Ala

965 970 975

Asp Val Gln Phe Ile Glu Ala His Gly Thr Gly Thr Ser Leu Gly Asp

980 985 990

Leu Ser Glu Ile Gln Ala Ile Asn Ala Ala Tyr Thr Ser Ser Gln Pro

995 1000 1005

Arg Thr Thr Gly Pro Leu Ile Val Ser Ala Ser Lys Thr Val Val

1010 1015 1020

Gly His Thr Glu Pro Ala Gly Pro Leu Val Gly Met Leu Ser Val

1025 1030 1035

Leu Leu Ser Phe Gln Glu Gly Ala Val Pro Gly Leu Ala His Leu

1040 1045 1050

Thr Thr Arg Asn Leu Asn Pro Thr Leu Asp Tyr Ser Ser Val Pro

1055 1060 1065

Leu Leu Ile Pro Ser Glu Pro Val Arg Leu Gln Thr Pro Lys Pro

1070 1075 1080

Tyr Arg Ala Ala Val Met Ser Tyr Gly Phe Ser Gly Thr Leu Ala

1085 1090 1095

Gly Leu Val Leu Glu Ser Pro Asp Glu Arg Ser Ser Glu Glu Glu

1100 1105 1110

Pro Pro Asp Asp Lys Pro Met Leu Phe Val Val Ser Ala Lys Thr

1115 1120 1125

His Thr Ala Leu Ile Glu Tyr Leu Gln Arg Tyr Leu Glu Phe Leu

1130 1135 1140

Leu His Ala Asn Pro Arg Asp Phe Cys Asp Ile Cys Tyr Thr Ser

1145 1150 1155

Cys Val Gly Arg Glu His Tyr Arg Tyr Arg Phe Ala Cys Val Ala

1160 1165 1170

Asn Asp Met Glu Asp Leu Ile Gly Gln Leu Gln Arg Arg Leu Ser

1175 1180 1185

Ser Lys Leu Pro Ser Lys Pro Leu Tyr Lys Arg Gly Ala Leu Ala

1190 1195 1200

Phe Ala Phe Ser Gly Gln Gly Thr Gln Phe Gln Gly Met Ala Thr

1205 1210 1215

Asp Leu Ala Lys Arg Tyr Ser Gly Phe Arg Lys Ile Val Ser Gly

1220 1225 1230

Leu Ala Lys Ser Ala Gly Glu Leu Ser Gly Tyr Ala Ile Asp Arg

1235 1240 1245

Tyr Leu Leu Ala Tyr Asp Val Gly Ser Ser Ile Ala Thr Pro Asn

1250 1255 1260

Ser Glu Val Asp Gln Ile Cys Ile Phe Val Tyr Gln Cys Ser Val

1265 1270 1275

Leu Arg Trp Leu Gly Ser Ile Gly Ile Lys Pro Asn Val Val Ile

1280 1285 1290

Gly His Ser Leu Gly Glu Ile Ser Ala Ser Val Ala Ala Gly Ala

1295 1300 1305

Leu Ser Leu Asp Ile Ala Leu Asp Leu Val Ile Ser Arg Ala Gly

1310 1315 1320

Leu Leu Arg Pro Ser Thr Asp Val Pro Ala Gly Met Ala Ala Val

1325 1330 1335

Ala Ala Ser Gln Gln Glu Val Ile Glu Leu Ile Asp Ala Leu Asp

1340 1345 1350

Pro Asp Lys Ala Asn Ser Leu Ser Val Ser Val Ile Asn Gly Pro

1355 1360 1365

Gln Asn Ile Val Val Ser Gly Ala Ser Ala Ala Ile Glu Arg Met

1370 1375 1380

Val Ala Ser Ala Lys Glu Lys Lys Ile Lys Ala Ser Val Leu Asn

1385 1390 1395

Ile Ser Gln Ala Phe His Ser Ser Tyr Val Asp Ser Ala Ile Thr

1400 1405 1410

Gly Leu Arg Ala Trp Ser Glu Lys His Ile Ser Ser Ala Ile Pro

1415 1420 1425

Leu Gln Ile Pro Leu Tyr Ser Thr Leu Leu Gly Ala Arg Ile Ser

1430 1435 1440

Lys Gly Gln Lys Leu Asn Pro Asp His Trp Val Asp His Ala Arg

1445 1450 1455

Lys Pro Val Gln Phe Ala Gln Ala Ala Thr Ala Met Lys Glu Thr

1460 1465 1470

Phe Thr Gly Val Ile Met Asp Ile Gly Pro Gln Ala Val Ala Trp

1475 1480 1485

Ser Leu Leu Leu Ala Asn Gly Leu Thr Ser Val Thr Ala Leu Ala

1490 1495 1500

Ala Lys Arg Gly Arg Ser Gln Gln Val Ala Phe Leu Ser Ala Leu

1505 1510 1515

Ala Glu Leu Tyr Gln Asp Tyr Gly Ile Val Pro Asp Phe Val Ala

1520 1525 1530

Leu Tyr Ala Gln Glu Glu Glu Ile Asp Lys Leu Arg Lys Thr Asp

1535 1540 1545

Ile Leu Thr Tyr Pro Phe Gln Arg Val Arg Arg Tyr Pro Ser Phe

1550 1555 1560

Ile Pro Ser Arg Arg Val Arg Gly Gly Glu Ile Pro Ser Ser His

1565 1570 1575

Pro Ser Glu Ser Ala Thr Leu Glu Asn Thr Asn Glu Gly Thr Ala

1580 1585 1590

Leu Arg Ala Glu Ser Arg Val Leu Cys Arg Glu Asp Leu His Asp

1595 1600 1605

Ser Ile Val Thr Ser Val Lys Asp Val Leu Glu Leu Lys Pro Asn

1610 1615 1620

Glu Asp Leu Asp Leu Ser Glu Ser Leu Asn Ala Leu Gly Val Asp

1625 1630 1635

Ser Ile Met Phe Ala Gln Leu Arg Lys Arg Ile Gly Glu Gly Leu

1640 1645 1650

Gly Leu Ser Ile Pro Thr Val Phe Leu Ser Asp Ala Phe Ser Ile

1655 1660 1665

Asn Glu Met Val Asn Asn Leu Met Glu Gln Ala Glu Thr Pro Gly

1670 1675 1680

Glu Glu Gly Val Met Gln Glu Asn

1685 1690

<210> 42

<211> 5298

<212> DNA

<213> Guyanagaster necrorhiza

<400> 42

atggcggccg ataggcactt ttctcttctc gatgtctttc tcgacgttgc ccataatgct 60

gagacgtcac aacgcaatat tttggaatgc ggcctggaca cctggacata ctcagatttg 120

gacatcatct cgtcggccct ggcccaggat ctcagtacta ccttgggttg ctctcccagg 180

gttgcagtcg tcagcgagaa ccatccctat gtatttgctc tcatgctggc cgtatggaag 240

cttggaggaa tattcatccc catcgacgtc catgttcccg ccgagctttt aacgggcatg 300

ctacgcatcg ttgctccaga ctgcgtggtg attcctgaga ctgatctttc taatcagcgc 360

gtcatctctg cactttacct ccacgttatt cccttcaatg tcaatgcgtc gacaatgtct 420

acacttcgac agaaatatgt cctatctact cagaaaccct tgctgtctga gttccctctt 480

cctcatgttg atcgtgcatg cctctatctc tttacgtcct ccgcgtcttc caccgccaac 540

ttgaaatgcg tacctttgac tcatgcactt atcctcagca actgtcgttc caagcttgca 600

tggtggcggc gtgttcgtcc agaagaaaac atggatggga tgcgtgtttt ggggtgggcg 660

ccttggtcac atattctcgc gtacatgcag gatattggga cagcaacgct cctgaatgca 720

ggctgctatg tctttgcatc tgttccgtcc acatatccta cgcagcacgt agccaatggc 780

ctgcaagacc ccacttcaaa tataatcaag tcgcttctca accgccgtat cacggcgttc 840

gcatgtgtgc cgttcattct tagtgaactg aaagctatgt gcgacccagc ttccggtccg 900

gacgccaagg atcaaatgtg cttgggagct ggggagaaag tgcgtcttat aagcacactg 960

cagaatctca tcatgttcga gtgtggaggc gctgcgctcg aggcagatat cacggattgg 1020

accgtcgaaa atggtatatc agtcatggtc ggtattggaa tgacggagac cgccggcacg 1080

gttttggcag cgcgtgcaca agacgctcgt tcgaatggct attctgccga agaggctctc 1140

atcgctgatg gcatactatc attggtcgga cctgacgagg aggaaatggc ctttgacgga 1200

gaacttgtcg tgaaaagcaa gctcatccca catggataca tcaagtacca cgattcggca 1260

ttttcagtgg actcagatga ctgggtgacg ttcaaaactg gggacaaata tcagcgtaca 1320

ccagatggac atttcaagtg gcttggaaga aagaccgatt ttattcagat gacaagcagc 1380

gaaaccttag atcccagacc tatcgaaaaa gccctctgca tgaaccccat tatcgcaaat 1440

gcgtgtgtca ttggtgacag gtttctaagg gaacctgcga cgggtgtatg tgccattgtc 1500

gagatcaggt cagatgtgga tatggcttcc gccgaggttg acagggagat tgcgaatgtc 1560

ctcgctccaa tcaatcgtga tcttcctccg gctcttcgaa tagcatggtc tcgcgtactc 1620

ataatccgac ctcctcagaa aataccactc acaaagaaag gtgaggtgtt tcgcaagaag 1680

attgaagata tatttggcac cttcttgggt gtcggtgtta ctaccaaggt ggaagtggac 1740

catgaaagta aagaagatga tacggaacac atcgtgagac aggttgtgag caatcttctt 1800

ggagtccatg atcctgagct attgtctact ttatctttcg ctgagctcgg gatgacctca 1860

tttatggccg ttagcatcgt taacacttta aacaagcata ttggcggcct cgcacttcca 1920

cttaactcat gctacattca tattgatctt gattctcttg tagatgccat ttcacttgaa 1980

cgtggtcatc gaaagaaccc tacgcccttt tcttctaacc ctttcctcgc ctttgaatcc 2040

agccagcaaa aggatatgga gattgtgatt gttggtaaag cattccgttt gcccggctca 2100

ctcgacaata gcacctctct ctgggaagct ttgttgtcaa agagtgattc agtcatcggt 2160

gacatcccac ccgatcgctg ggatcatgca agtttttacc cccacgatat atgctttacg 2220

aaagcaggcc ttgtcgatgt ttcccattat gactatagat ttttcggtct cactgctaca 2280

gaggcgttgt acgtatctcc gacaatgcgc ctcgccttgg aagtgtcatt tgaggctctg 2340

gagaatgcaa atattccctt atccaagttg aaggggacac gaactgctgt ttatgttgcc 2400

actaaagacg atgggttcga gacactctta aatgctgaac aaggctacga tgcctacacg 2460

cgattctacg gcacgggacg tgctccgagt accgcaagtg gccgtataag ctatcttctt 2520

gatattcatg gaccttctgt taccgttgac acagcatgca gcggaggcat tgtatgtata 2580

gaccaagcca tcacttttct gcaatccgga ggggcagata ctgctattgt ttgttcgagc 2640

aatacgcact gttggccggg atcatttatg ttcctgacag cgcaaggcat ggtctctcca 2700

aatggaagat gtgccacatt cactacaaat gcagatggat atgtaccttc agaaggcgca 2760

gtggctttcg ttctcaagac acacagcgca gcaacacgcg acaaagacaa tatactggcc 2820

gtgataaagt caacagacgt gtctcataac ggccgttctc aagggctagt tgcaccaaac 2880

gtaaaggcac agacaaacct acaccagtcg ctgttacgaa aagctgggct gcatcctgat 2940

ctaattaact ttatcgaagc tcatggaaca ggtacatctt taggagacct ttcagaaatc 3000

cagggtatca ataatgccta cacctcatct cgacctcgtc cagccggtcc tctgatcatt 3060

agcgcgtcaa aaacggtttt gggacatagt gaaccaactg caggaatggc cggcatcctc 3120

acaaccttgc tcgcctttga gaaagagacg gttcctggtc tcaaccactt aacggaacat 3180

aatttaaacc cttcgcttga ttgctctgca gtgcctctcc tgattcctca cgagcctgtt 3240

catatcagtg gtgcaaagcc ccatcgagct gcggttctat catacggatt cgcggggaca 3300

ctggctggta ccatcttaga gagtccacct tgcgacctta caaggccctt gtcaaacgac 3360

atacaagaac atcctatgat tttcgtcgtc agtgggaaaa cggtgccctc tctggaagca 3420

tatctagagc ggtatttggt atttttacgg gtcgcaaaac caagcgaatt ccatgacatc 3480

tgctacacca cttgcatcgg gagggagctg tataaatacc ggttctcctg cgttgcccga 3540

aacatggccg acctcatttc tcaaattgaa catcgactga cgacttgttc cacttcgaaa 3600

gagaagcccc gtggctcgtt aggattcgtg ttctcgggtc aaggtaccca gttccccggc 3660

atggcagcag cacttgccga acgatattca gggttccgag cgctcgtctc caagtttggg 3720

cagatcgccc aagagcagtc gggctacccg atcgataggc tgctcctcga agttaccgat 3780

acattaccag aagcaaacag cgaggtcgac caaatttgca tttttgtcta ccaatattct 3840

gttctgcaat ggctgcaacg tctaggcatt caaccgaaaa cagtccttgg tcacagtcta 3900

ggagaaatta ccgcctcagt cgcagttggc gccttctctt ttaggtctgc attggacctt 3960

gtggtcaccc gcgctcgtct tcttcgtcct caaccaaaat tctctgcggg aatggctgca 4020

gtagcagcgt ccaaggaaga agttgaagga cttatagata tgctcaaact tgcggagtcg 4080

ctgagcgttg cggttcataa cagtccacgg agtatcgttg tatcaggcgc atcagctgcg 4140

gtcgatgcca tggttgtcgc tgctaaaaag cagggcttga aggcctcccg cttaaaggtt 4200

gaccaagcct ttcacagccc ttacgttgat tctgctgtat cggggttgct cgactggtcc 4260

aataagcatc gttcgacctt cctcccattg aacatccctt tatactcaac tttgactggc 4320

gcgcgtattc cgaaaggagg gaagttctgc tgggatcact gggttaatca tgctcgaaag 4380

cccgtccagt ttgcggcggc agctgcagca atggacgaag atcaatccat cggtgttctt 4440

gttgatgttg gaccccaacc tgttgcgtgg accctccttc aagcgaataa cctcctcaat 4500

acctcttcga ttgctctatc agcaaaaatt ggaaaggatc aggagatggc gttgctctct 4560

gctttgagct acctcgtcca agagcacaac ctttctctca gcttttatga gctttactct 4620

cagcgtcacg gtactctgaa gaagacagac gttcctacct acccgttccg tcgcgtccac 4680

cgctacccga ctttcatacc gtcacggaat agaagtcctg ctgacatgag ggtagctata 4740

ccacctaccg acctctctgt ccgaaagaat gtggatgcaa caccgcagtc tcgtcgtgct 4800

ggcctgatag cctgtcttaa agttatcctc gagttaacac caggagagga atttgacctt 4860

tctgagactc tcaatgctcg tggggtcgat tcagttatgt tcgcacagtt gcggaagcgt 4920

gttggggaag aattcgacct agatatacca atgatctatt tatcagacgt gttcacgatg 4980

gaacagatga ttgactacct cgtcgaacag tccagcccca catcaaagtt ggtagagatt 5040

tcggttaatc aatcattaga cggagaaggc ctccggacag ggcttgtatc atgccttagg 5100

gacgtactgg aaatctcctc cgacgaagaa cttgactttt ccgaaacttt gaacgctcgt 5160

ggtacggact caattatgtt cgctcagcta cgaaaacgtg tcggggaagg atttggtctt 5220

gaaattccga tgatatatct gtctgatgtg tttaccatgg aagacatgat caatttcctt 5280

gtctcggagc gctcgtgc 5298

<210> 43

<211> 1766

<212>PRT

<213> Guyanagaster necrorhiza

<400> 43

Met Ala Ala Asp Arg His Phe Ser Leu Leu Asp Val Phe Leu Asp Val

1 5 10 15

Ala His Asn Ala Glu Thr Ser Gln Arg Asn Ile Leu Glu Cys Gly Leu

20 25 30

Asp Thr Trp Thr Tyr Ser Asp Leu Asp Ile Ile Ser Ser Ala Leu Ala

35 40 45

Gln Asp Leu Ser Thr Thr Leu Gly Cys Ser Pro Arg Val Ala Val Val

50 55 60

Ser Glu Asn His Pro Tyr Val Phe Ala Leu Met Leu Ala Val Trp Lys

65 70 75 80

Leu Gly Gly Ile Phe Ile Pro Ile Asp Val His Val Pro Ala Glu Leu

85 90 95

Leu Thr Gly Met Leu Arg Ile Val Ala Pro Asp Cys Val Val Ile Pro

100 105 110

Glu Thr Asp Leu Ser Asn Gln Arg Val Ile Ser Ala Leu Tyr Leu His

115 120 125

Val Ile Pro Phe Asn Val Asn Ala Ser Thr Met Ser Thr Leu Arg Gln

130 135 140

Lys Tyr Val Leu Ser Thr Gln Lys Pro Leu Leu Ser Glu Phe Pro Leu

145 150 155 160

Pro His Val Asp Arg Ala Cys Leu Tyr Leu Phe Thr Ser Ser Ala Ser

165 170 175

Ser Thr Ala Asn Leu Lys Cys Val Pro Leu Thr His Ala Leu Ile Leu

180 185 190

Ser Asn Cys Arg Ser Lys Leu Ala Trp Trp Arg Arg Val Arg Pro Glu

195 200 205

Glu Asn Met Asp Gly Met Arg Val Leu Gly Trp Ala Pro Trp Ser His

210 215 220

Ile Leu Ala Tyr Met Gln Asp Ile Gly Thr Ala Thr Leu Leu Asn Ala

225 230 235 240

Gly Cys Tyr Val Phe Ala Ser Val Pro Ser Thr Tyr Pro Thr Gln His

245 250 255

Val Ala Asn Gly Leu Gln Asp Pro Thr Ser Asn Ile Ile Lys Ser Leu

260 265 270

Leu Asn Arg Arg Ile Thr Ala Phe Ala Cys Val Pro Phe Ile Leu Ser

275 280 285

Glu Leu Lys Ala Met Cys Asp Pro Ala Ser Gly Pro Asp Ala Lys Asp

290 295 300

Gln Met Cys Leu Gly Ala Gly Glu Lys Val Arg Leu Ile Ser Thr Leu

305 310 315 320

Gln Asn Leu Ile Met Phe Glu Cys Gly Gly Ala Ala Leu Glu Ala Asp

325 330 335

Ile Thr Asp Trp Thr Val Glu Asn Gly Ile Ser Val Met Val Gly Ile

340 345 350

Gly Met Thr Glu Thr Ala Gly Thr Val Leu Ala Ala Arg Ala Gln Asp

355 360 365

Ala Arg Ser Asn Gly Tyr Ser Ala Glu Glu Ala Leu Ile Ala Asp Gly

370 375 380

Ile Leu Ser Leu Val Gly Pro Asp Glu Glu Glu Met Ala Phe Asp Gly

385 390 395 400

Glu Leu Val Val Lys Ser Lys Leu Ile Pro His Gly Tyr Ile Lys Tyr

405 410 415

His Asp Ser Ala Phe Ser Val Asp Ser Asp Asp Trp Val Thr Phe Lys

420 425 430

Thr Gly Asp Lys Tyr Gln Arg Thr Pro Asp Gly His Phe Lys Trp Leu

435 440 445

Gly Arg Lys Thr Asp Phe Ile Gln Met Thr Ser Ser Glu Thr Leu Asp

450 455 460

Pro Arg Pro Ile Glu Lys Ala Leu Cys Met Asn Pro Ile Ile Ala Asn

465 470 475 480

Ala Cys Val Ile Gly Asp Arg Phe Leu Arg Glu Pro Ala Thr Gly Val

485 490 495

Cys Ala Ile Val Glu Ile Arg Ser Asp Val Asp Met Ala Ser Ala Glu

500 505 510

Val Asp Arg Glu Ile Ala Asn Val Leu Ala Pro Ile Asn Arg Asp Leu

515 520 525

Pro Pro Ala Leu Arg Ile Ala Trp Ser Arg Val Leu Ile Ile Arg Pro

530 535 540

Pro Gln Lys Ile Pro Leu Thr Lys Lys Gly Glu Val Phe Arg Lys Lys

545 550 555 560

Ile Glu Asp Ile Phe Gly Thr Phe Leu Gly Val Gly Val Thr Thr Lys

565 570 575

Val Glu Val Asp His Glu Ser Lys Glu Asp Asp Thr Glu His Ile Val

580 585 590

Arg Gln Val Val Ser Asn Leu Leu Gly Val His Asp Pro Glu Leu Leu

595 600 605

Ser Thr Leu Ser Phe Ala Glu Leu Gly Met Thr Ser Phe Met Ala Val

610 615 620

Ser Ile Val Asn Thr Leu Asn Lys His Ile Gly Gly Leu Ala Leu Pro

625 630 635 640

Leu Asn Ser Cys Tyr Ile His Ile Asp Leu Asp Ser Leu Val Asp Ala

645 650 655

Ile Ser Leu Glu Arg Gly His Arg Lys Asn Pro Thr Pro Phe Ser Ser

660 665 670

Asn Pro Phe Leu Ala Phe Glu Ser Ser Gln Gln Lys Asp Met Glu Ile

675 680 685

Val Ile Val Gly Lys Ala Phe Arg Leu Pro Gly Ser Leu Asp Asn Ser

690 695 700

Thr Ser Leu Trp Glu Ala Leu Leu Ser Lys Ser Asp Ser Val Ile Gly

705 710 715 720

Asp Ile Pro Pro Asp Arg Trp Asp His Ala Ser Phe Tyr Pro His Asp

725 730 735

Ile Cys Phe Thr Lys Ala Gly Leu Val Asp Val Ser His Tyr Asp Tyr

740 745 750

Arg Phe Phe Gly Leu Thr Ala Thr Glu Ala Leu Tyr Val Ser Pro Thr

755 760 765

Met Arg Leu Ala Leu Glu Val Ser Phe Glu Ala Leu Glu Asn Ala Asn

770 775 780

Ile Pro Leu Ser Lys Leu Lys Gly Thr Arg Thr Ala Val Tyr Val Ala

785 790 795 800

Thr Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Glu Gln Gly Tyr

805 810 815

Asp Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser Thr Ala

820 825 830

Ser Gly Arg Ile Ser Tyr Leu Leu Asp Ile His Gly Pro Ser Val Thr

835 840 845

Val Asp Thr Ala Cys Ser Gly Gly Ile Val Cys Ile Asp Gln Ala Ile

850 855 860

Thr Phe Leu Gln Ser Gly Gly Ala Asp Thr Ala Ile Val Cys Ser Ser

865 870 875 880

Asn Thr His Cys Trp Pro Gly Ser Phe Met Phe Leu Thr Ala Gln Gly

885 890 895

Met Val Ser Pro Asn Gly Arg Cys Ala Thr Phe Thr Thr Asn Ala Asp

900 905 910

Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Val Leu Lys Thr His

915 920 925

Ser Ala Ala Thr Arg Asp Lys Asp Asn Ile Leu Ala Val Ile Lys Ser

930 935 940

Thr Asp Val Ser His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn

945 950 955 960

Val Lys Ala Gln Thr Asn Leu His Gln Ser Leu Leu Arg Lys Ala Gly

965 970 975

Leu His Pro Asp Leu Ile Asn Phe Ile Glu Ala His Gly Thr Gly Thr

980 985 990

Ser Leu Gly Asp Leu Ser Glu Ile Gln Gly Ile Asn Asn Ala Tyr Thr

995 1000 1005

Ser Ser Arg Pro Arg Pro Ala Gly Pro Leu Ile Ile Ser Ala Ser

1010 1015 1020

Lys Thr Val Leu Gly His Ser Glu Pro Thr Ala Gly Met Ala Gly

1025 1030 1035

Ile Leu Thr Thr Leu Leu Ala Phe Glu Lys Glu Thr Val Pro Gly

1040 1045 1050

Leu Asn His Leu Thr Glu His Asn Leu Asn Pro Ser Leu Asp Cys

1055 1060 1065

Ser Ala Val Pro Leu Leu Ile Pro His Glu Pro Val His Ile Ser

1070 1075 1080

Gly Ala Lys Pro His Arg Ala Ala Val Leu Ser Tyr Gly Phe Ala

1085 1090 1095

Gly Thr Leu Ala Gly Thr Ile Leu Glu Ser Pro Pro Cys Asp Leu

1100 1105 1110

Thr Arg Pro Leu Ser Asn Asp Ile Gln Glu His Pro Met Ile Phe

1115 1120 1125

Val Val Ser Gly Lys Thr Val Pro Ser Leu Glu Ala Tyr Leu Glu

1130 1135 1140

Arg Tyr Leu Val Phe Leu Arg Val Ala Lys Pro Ser Glu Phe His

1145 1150 1155

Asp Ile Cys Tyr Thr Thr Cys Ile Gly Arg Glu Leu Tyr Lys Tyr

1160 1165 1170

Arg Phe Ser Cys Val Ala Arg Asn Met Ala Asp Leu Ile Ser Gln

1175 1180 1185

Ile Glu His Arg Leu Thr Thr Cys Ser Thr Ser Lys Glu Lys Pro

1190 1195 1200

Arg Gly Ser Leu Gly Phe Val Phe Ser Gly Gln Gly Thr Gln Phe

1205 1210 1215

Pro Gly Met Ala Ala Ala Leu Ala Glu Arg Tyr Ser Gly Phe Arg

1220 1225 1230

Ala Leu Val Ser Lys Phe Gly Gln Ile Ala Gln Glu Gln Ser Gly

1235 1240 1245

Tyr Pro Ile Asp Arg Leu Leu Leu Glu Val Thr Asp Thr Leu Pro

1250 1255 1260

Glu Ala Asn Ser Glu Val Asp Gln Ile Cys Ile Phe Val Tyr Gln

1265 1270 1275

Tyr Ser Val Leu Gln Trp Leu Gln Arg Leu Gly Ile Gln Pro Lys

1280 1285 1290

Thr Val Leu Gly His Ser Leu Gly Glu Ile Thr Ala Ser Val Ala

1295 1300 1305

Val Gly Ala Phe Ser Phe Arg Ser Ala Leu Asp Leu Val Val Thr

1310 1315 1320

Arg Ala Arg Leu Leu Arg Pro Gln Pro Lys Phe Ser Ala Gly Met

1325 1330 1335

Ala Ala Val Ala Ala Ser Lys Glu Glu Val Glu Gly Leu Ile Asp

1340 1345 1350

Met Leu Lys Leu Ala Glu Ser Leu Ser Val Ala Val His Asn Ser

1355 1360 1365

Pro Arg Ser Ile Val Val Ser Gly Ala Ser Ala Ala Val Asp Ala

1370 1375 1380

Met Val Val Ala Ala Lys Lys Gln Gly Leu Lys Ala Ser Arg Leu

1385 1390 1395

Lys Val Asp Gln Ala Phe His Ser Pro Tyr Val Asp Ser Ala Val

1400 1405 1410

Ser Gly Leu Leu Asp Trp Ser Asn Lys His Arg Ser Thr Phe Leu

1415 1420 1425

Pro Leu Asn Ile Pro Leu Tyr Ser Thr Leu Thr Gly Ala Arg Ile

1430 1435 1440

Pro Lys Gly Gly Lys Phe Cys Trp Asp His Trp Val Asn His Ala

1445 1450 1455

Arg Lys Pro Val Gln Phe Ala Ala Ala Ala Ala Ala Met Asp Glu

1460 1465 1470

Asp Gln Ser Ile Gly Val Leu Val Asp Val Gly Pro Gln Pro Val

1475 1480 1485

Ala Trp Thr Leu Leu Gln Ala Asn Asn Leu Leu Asn Thr Ser Ser

1490 1495 1500

Ile Ala Leu Ser Ala Lys Ile Gly Lys Asp Gln Glu Met Ala Leu

1505 1510 1515

Leu Ser Ala Leu Ser Tyr Leu Val Gln Glu His Asn Leu Ser Leu

1520 1525 1530

Ser Phe Tyr Glu Leu Tyr Ser Gln Arg His Gly Thr Leu Lys Lys

1535 1540 1545

Thr Asp Val Pro Thr Tyr Pro Phe Arg Arg Val His Arg Tyr Pro

1550 1555 1560

Thr Phe Ile Pro Ser Arg Asn Arg Ser Pro Ala Asp Met Arg Val

1565 1570 1575

Ala Ile Pro Pro Thr Asp Leu Ser Val Arg Lys Asn Val Asp Ala

1580 1585 1590

Thr Pro Gln Ser Arg Arg Ala Gly Leu Ile Ala Cys Leu Lys Val

1595 1600 1605

Ile Leu Glu Leu Thr Pro Gly Glu Glu Phe Asp Leu Ser Glu Thr

1610 1615 1620

Leu Asn Ala Arg Gly Val Asp Ser Val Met Phe Ala Gln Leu Arg

1625 1630 1635

Lys Arg Val Gly Glu Glu Phe Asp Leu Asp Ile Pro Met Ile Tyr

1640 1645 1650

Leu Ser Asp Val Phe Thr Met Glu Gln Met Ile Asp Tyr Leu Val

1655 1660 1665

Glu Gln Ser Ser Pro Thr Ser Lys Leu Val Glu Ile Ser Val Asn

1670 1675 1680

Gln Ser Leu Asp Gly Glu Gly Leu Arg Thr Gly Leu Val Ser Cys

1685 1690 1695

Leu Arg Asp Val Leu Glu Ile Ser Ser Asp Glu Glu Leu Asp Phe

1700 1705 1710

Ser Glu Thr Leu Asn Ala Arg Gly Thr Asp Ser Ile Met Phe Ala

1715 1720 1725

Gln Leu Arg Lys Arg Val Gly Glu Gly Phe Gly Leu Glu Ile Pro

1730 1735 1740

Met Ile Tyr Leu Ser Asp Val Phe Thr Met Glu Asp Met Ile Asn

1745 1750 1755

Phe Leu Val Ser Glu Arg Ser Cys

1760 1765

<210> 44

<211> 5034

<212> DNA

<213> Mycena citricolor

<400> 44

atgtccgcct cttcctccta ttctgaactg gagtctccga cgtcgctgct tgacgtattc 60

gtgcacgccg cgcgagaccc ccacaccgcg tcgcgtcgcg tgctggagtg cggctcggac 120

acatggacat acgcggcctt ggatgcggta tccgatggca tagcgaggga gctcgcgccc 180

ttcggtctgg cgcccaaggt cgctgtggtc agcgagaacc atccatttgt gttcgcgctg 240

ctgttcgcag tctggaagct cgggggaacg ttcatcccca tcgacgcgca tgtccccttt 300

gccatgctca cggggatggt gaacatcgtg aagccgacgt gtctctacct ccccgcctcc 360

gccacatcca acatatctct cgtgaaggcc ttcgacatac ggaccgtcgt atttgggcac 420

aaggagaatt ccatgcaagc tttgttcgac aagtactcat tgcacgcggc gccgttgcac 480

gcggcaccat tgcattacgc gcctcccagt gccgaccatg cctgtctgta tctcttcacg 540

tcgtcagcgt cctcgaccaa aaacctcaag gcggtcccgc tgacgcatac actcgtcctg 600

cggggggtgtc aatccaaact cgcttggtgg cgccgagtcc aacccggaaa gaatctcgac 660

gctatccgga tcctcggctg ggcaccctgg tctcacgttc tcgcctatat gcaggatatc 720

gggacagcga cactgctcaa cgcgggctgc tacgtctttg cgaccatccc ttcgtcctac 780

ccgcagctgc ctaccactac ccccgtcgat ctgacgactt cgctcatcga cgcgctcgtc 840

acgcggcgcg tcgcggcatt cgcctgtgtc cccttcgtcc tggcgaacct caaagccgcg 900

tgccagagca accaccccgc gcgctcccag ctgctcgagg cgctgcagaa gacgatcatg 960

ctcgagtgcg gtggggcggt gctggacgac gcgacggtgg actgggccga gcgcaacggc 1020

atccgcatct tcacgggaat cgggatgacc gagactggcg gggccgtttt cgtcggcctc 1080

gcgagcgaat ctagacgcgg gtttctaccg gaaggcctac tcggtgatgc atccttctcg 1140

atctcgagtg acacggacgc tcttgacgaa ggggaacttg tcgtgaaaag taaactcatc 1200

gcgcccggat atgtcggcta cgacgacgga gcgcactccg tggactgcga cggctgggtc 1260

acattcagga caggagatcg atatcgccag acccagccag acgggcgctt tacgtggcta 1320

ggaaggatca cagacttcat tcagatggtc agcggcgagt acctcgatcc gcggcccctc 1380

gagggagagtc tgcgtgcgtc cccgctgatc gccaacgcct gcatcgtggg cgacgcgttc 1440

ctcagcagcg cctcgacgag catcctcgcc atcatcgagc tcgcgacccc ggatctggcg 1500

cacacggcct ccatccgggc gcagctcgcg cgcgtcctcg cgcccctcaa ccgcgatctg 1560

cctccaccgc ttcggatcgc agcgtcttcg atcctggttc tggacgggat gcggaagatc 1620

ccgaagacga agaagggcga catcttccgc aaaaagctgg aagacacgtt cggtgcggag 1680

tttgagcaga tgctgcggac cgagaaagtc gggctgggtg acttggcgga tgtggatgcc 1740

ggtatcactc gtatcatcgg caacttgctc ggcatctccg acgacgagct tctgtcgacc 1800

atgtcgtttg ctgagctcgg gatgacctcg cttctcgccg tcaagatcgc gagcgaacta 1860

aacaagtttc tggacggccg ggctgtgctg cccacgaaca tctgttacat acactttgac 1920

gtgccctcgc tcacgagtag cgtccgagaa aggctatcct ccgcgccctc ctcgatcaca 1980

tccgccgcca gcgagcccgc ggcctcgtcc cccggcaccc cccgcgccga cgaaatcgtg 2040

atcgtgggca aggccttccg gctgccggac ggcgtgaacg acgacgccgc gctgtgggac 2100

gtcctgacgg gcgagtccgc gtcgatcatc aaggacatcc ccgccgaccg ctgggaccac 2160

gcgagcttct acccgaagga catccacttt ggcagagctg gtctggtgga tgtcgcgcgg 2220

ttcgactacg gctttttcgg catgacggcg agcgaggcgt attcgctgtc gccgacgatg 2280

cgcttggcgc tcgaagtcgc gtatgaggcg ttagaggacg cgaacattcc gttccgggcc 2340

gtcaagggct cgcgcatggg tgtgttcgtc gctgtgaaag acgatggatt cgagaccctg 2400

ttgcatgcgg ggcagggtta tgacgcttac acgcggttct atggaactgg aagggcgccg 2460

agcaccgcca gcggccgcat caactatctg ctcgatctcc acgggccgtc gatcaccgtc 2520

gacacggcct gcagcggcgg catcgtgtgc atcgaccagg ctgtcaccta tcttcaatcg 2580

ggcgctgcag agacggcgat cgtgtgttcg agcaacacac actgctggcc cggatccttc 2640

atgttcttga ccgcgcaagg catggcctcc cccaacggcc ggtgcgcatc cttcacatca 2700

gacgccgacg gatacgcgcc gtcggagggc gcggtgggct tcgtgctgaa gacgcggtct 2760

gccgctgtgc gcgacggcga tcggatactg gctacgatca gggccacgga gatcggacac 2820

aatggacggt ctcagggact cgctgcgccg aatgtcaggt cccaggcggc tgctcatcgg 2880

gcggtgctgc ggagagcgag gctggacccg tccgagattg acttcatcga agcgcatggg 2940

actggcacca cattgggcga tctgtgcgaa gtgcagggca tcaacgacag ctttgtctcg 3000

cccaagaaac gcgccaaccc tcttgtggtc agcgcgtcca agagcaccat cggccacacc 3060

gagccgtcgg cgggtctcgt cgggatcctg tccgcgctga tgtcgttcga gaagcgcatc 3120

gtgccgagat tggcatatct gactgagagc aatgtgaacc cggcgctcga cgcgagtgtc 3180

gttccgctgc actttcccac aaagcacatc gagctgcgcg ctgatgtgcc gtacaaggcc 3240

gtagtgatgt cgtacggatt cgcaggtacc ctagccgaca tcgtcctcga gagcgaggta 3300

ccccaaccca cgcccgcggt cgctcaggac acggccggcc aacagccaat gctcttcgtg 3360

ctcagcgcca aaaccccacg cgcactcgca gcctacatcg agctgtacct aggcttcctg 3420

cggcacgcgg acccggggcct cttcgcgcgc atctgctaca cggcgtgtgt cgcgcgcgag 3480

cactacaagc accgcgtcgc gtgcgttgcg acggacctcg tcgacctcat cgcgcagctc 3540

gagacgcgtc tggtgcagac ggcgtatgcg ggcggcggcg gggcccgggc cgcgcgcacc 3600

gggccgctgg tgtttgcgtt ttcggggcag ggcacgcagt tcccggcgat ggcggcgccg 3660

ctggcgcggc ggtacgcgcg cttcggggag atcgtggggg gctgtgcgcg catggcgcgc 3720

gagctgagcg ggttccccgt ggatggcatt ctcctcggag acgatgtgac gcccgtgaag 3780

gacaacagcg ctgcggcgga ggtgcacagc gaggtggacc agatctgcat atttgtgtac 3840

cagtatgcga tgtgtcggtg gttgggcgag ctgggtgtgg agcccaaggc tgccataggc 3900

cacagcttgg gagagatcac agcagcggtc atcgcaggag cactcccttt cgaagccgcc 3960

ctcgatctgg tcgtcacgcg cgcccggctg ctcaagccgt gcgccgagca accgagcggc 4020

atggcggcgc tcgcctgcac cccagacgtc gcatcgaagc tcacactcgg cgcgtcggtg 4080

tcggtgtcgg tctacaacgg cccgcagagc atctgcctgt ccggcgcgtc ggccgagctc 4140

gacgatgcgg tgcgggccgc gaaggcgcgg aacatcaagg cgacgcggct gcaggtcgac 4200

cagggcttcc actcgccgtg cgtcgacgcc gcggtgccgg ggctgcaggc gtggtgcgcg 4260

gcgcaccgtg cgtcagctgc gccgctgaag atgccgctgt actcgaccgt gcgcggggac 4320

gtcgttccga agggagcggc gctggatccg gagcactggg tggcgcacgc gcggaacccg 4380

gtgctgttcg cgcagacggc gcaggctctg aaagagtccc ttccgcatgg gatcactttg 4440

gacgtcggcc cgcaggcggt ggcgtggtcg ctgctgctgc tgaacgggct cagcgcgacg 4500

cgcacggtcg ctgccggggc gaagaagggc gcagaccagg agcgcgcgct gctggggggcg 4560

ctggggggcgc tgttcgagca gcacaaggtc acgccggact ttgggcggct gtacgcgccg 4620

ctcgagaaga cgcggatccc gacgtacccc tttgagcgcg cgcggtgcta cccgacgttc 4680

ataccctcgc gcttcgcgca cggggctgcg gctgctgcga agacagatgg cgaggtcctg 4740

tcggccgaag aagaaaatgt ggcacccgtg ggattgctga caaagggagga tctgcgtgcg 4800

gctctcgtcg cgtgtctccg agcgacgctc gagctgcgcc ccgacgaaga gctggacgaa 4860

gcagagccgc tgaccgtgca cggcgtggac tcgatcgggt ttgcgaagct gcgcaagcac 4920

gtcgaggacc gctggggggct ggacatcccc gtcgtgtact ggtccgacgc gttcaccgtc 4980

ggcgagatgc tcggcaactt ggtcggccag tatgacgtag tgtctactgc tgcg 5034

<210> 45

<211> 1678

<212>PRT

<213> Mycena citricolor

<400> 45

Met Ser Ala Ser Ser Ser Tyr Ser Glu Leu Glu Ser Pro Thr Ser Leu

1 5 10 15

Leu Asp Val Phe Val His Ala Ala Arg Asp Pro His Thr Ala Ser Arg

20 25 30

Arg Val Leu Glu Cys Gly Ser Asp Thr Trp Thr Tyr Ala Ala Leu Asp

35 40 45

Ala Val Ser Asp Gly Ile Ala Arg Glu Leu Ala Pro Phe Gly Leu Ala

50 55 60

Pro Lys Val Ala Val Val Ser Glu Asn His Pro Phe Val Phe Ala Leu

65 70 75 80

Leu Phe Ala Val Trp Lys Leu Gly Gly Thr Phe Ile Pro Ile Asp Ala

85 90 95

His Val Pro Phe Ala Met Leu Thr Gly Met Val Asn Ile Val Lys Pro

100 105 110

Thr Cys Leu Tyr Leu Pro Ala Ser Ala Thr Ser Asn Ile Ser Leu Val

115 120 125

Lys Ala Phe Asp Ile Arg Thr Val Val Phe Gly His Lys Glu Asn Ser

130 135 140

Met Gln Ala Leu Phe Asp Lys Tyr Ser Leu His Ala Ala Pro Leu His

145 150 155 160

Ala Ala Pro Leu His Tyr Ala Pro Pro Ser Ala Asp His Ala Cys Leu

165 170 175

Tyr Leu Phe Thr Ser Ser Ala Ser Ser Thr Lys Asn Leu Lys Ala Val

180 185 190

Pro Leu Thr His Thr Leu Val Leu Arg Gly Cys Gln Ser Lys Leu Ala

195 200 205

Trp Trp Arg Arg Val Gln Pro Gly Lys Asn Leu Asp Ala Ile Arg Ile

210 215 220

Leu Gly Trp Ala Pro Trp Ser His Val Leu Ala Tyr Met Gln Asp Ile

225 230 235 240

Gly Thr Ala Thr Leu Leu Asn Ala Gly Cys Tyr Val Phe Ala Thr Ile

245 250 255

Pro Ser Ser Tyr Pro Gln Leu Pro Thr Thr Thr Pro Val Asp Leu Thr

260 265 270

Thr Ser Leu Ile Asp Ala Leu Val Thr Arg Arg Val Ala Ala Phe Ala

275 280 285

Cys Val Pro Phe Val Leu Ala Asn Leu Lys Ala Ala Cys Gln Ser Asn

290 295 300

His Pro Ala Arg Ser Gln Leu Leu Glu Ala Leu Gln Lys Thr Ile Met

305 310 315 320

Leu Glu Cys Gly Gly Ala Val Leu Asp Asp Ala Thr Val Asp Trp Ala

325 330 335

Glu Arg Asn Gly Ile Arg Ile Phe Thr Gly Ile Gly Met Thr Glu Thr

340 345 350

Gly Gly Ala Val Phe Val Gly Leu Ala Ser Glu Ser Arg Arg Gly Phe

355 360 365

Leu Pro Glu Gly Leu Leu Gly Asp Ala Ser Phe Ser Ile Ser Ser Asp

370 375 380

Thr Asp Ala Leu Asp Glu Gly Glu Leu Val Val Lys Ser Lys Leu Ile

385 390 395 400

Ala Pro Gly Tyr Val Gly Tyr Asp Asp Gly Ala His Ser Val Asp Cys

405 410 415

Asp Gly Trp Val Thr Phe Arg Thr Gly Asp Arg Tyr Arg Gln Thr Gln

420 425 430

Pro Asp Gly Arg Phe Thr Trp Leu Gly Arg Ile Thr Asp Phe Ile Gln

435 440 445

Met Val Ser Gly Glu Tyr Leu Asp Pro Arg Pro Leu Glu Glu Ser Leu

450 455 460

Arg Ala Ser Pro Leu Ile Ala Asn Ala Cys Ile Val Gly Asp Ala Phe

465 470 475 480

Leu Ser Ser Ala Ser Thr Ser Ile Leu Ala Ile Ile Glu Leu Ala Thr

485 490 495

Pro Asp Leu Ala His Thr Ala Ser Ile Arg Ala Gln Leu Ala Arg Val

500 505 510

Leu Ala Pro Leu Asn Arg Asp Leu Pro Pro Pro Leu Arg Ile Ala Ala

515 520 525

Ser Ser Ile Leu Val Leu Asp Gly Met Arg Lys Ile Pro Lys Thr Lys

530 535 540

Lys Gly Asp Ile Phe Arg Lys Lys Leu Glu Asp Thr Phe Gly Ala Glu

545 550 555 560

Phe Glu Gln Met Leu Arg Thr Glu Lys Val Gly Leu Gly Asp Leu Ala

565 570 575

Asp Val Asp Ala Gly Ile Thr Arg Ile Ile Gly Asn Leu Leu Gly Ile

580 585 590

Ser Asp Asp Glu Leu Leu Ser Thr Met Ser Phe Ala Glu Leu Gly Met

595 600 605

Thr Ser Leu Leu Ala Val Lys Ile Ala Ser Glu Leu Asn Lys Phe Leu

610 615 620

Asp Gly Arg Ala Val Leu Pro Thr Asn Ile Cys Tyr Ile His Phe Asp

625 630 635 640

Val Pro Ser Leu Thr Ser Ser Val Arg Glu Arg Leu Ser Ser Ala Pro

645 650 655

Ser Ser Ile Thr Ser Ala Ala Ser Glu Pro Ala Ala Ser Ser Pro Gly

660 665 670

Thr Pro Arg Ala Asp Glu Ile Val Ile Val Gly Lys Ala Phe Arg Leu

675 680 685

Pro Asp Gly Val Asn Asp Asp Ala Ala Leu Trp Asp Val Leu Thr Gly

690 695 700

Glu Ser Ala Ser Ile Ile Lys Asp Ile Pro Ala Asp Arg Trp Asp His

705 710 715 720

Ala Ser Phe Tyr Pro Lys Asp Ile His Phe Gly Arg Ala Gly Leu Val

725 730 735

Asp Val Ala Arg Phe Asp Tyr Gly Phe Phe Gly Met Thr Ala Ser Glu

740 745 750

Ala Tyr Ser Leu Ser Pro Thr Met Arg Leu Ala Leu Glu Val Ala Tyr

755 760 765

Glu Ala Leu Glu Asp Ala Asn Ile Pro Phe Arg Ala Val Lys Gly Ser

770 775 780

Arg Met Gly Val Phe Val Ala Val Lys Asp Asp Gly Phe Glu Thr Leu

785 790 795 800

Leu His Ala Gly Gln Gly Tyr Asp Ala Tyr Thr Arg Phe Tyr Gly Thr

805 810 815

Gly Arg Ala Pro Ser Thr Ala Ser Gly Arg Ile Asn Tyr Leu Leu Asp

820 825 830

Leu His Gly Pro Ser Ile Thr Val Asp Thr Ala Cys Ser Gly Gly Ile

835 840 845

Val Cys Ile Asp Gln Ala Val Thr Tyr Leu Gln Ser Gly Ala Ala Glu

850 855 860

Thr Ala Ile Val Cys Ser Ser Asn Thr His Cys Trp Pro Gly Ser Phe

865 870 875 880

Met Phe Leu Thr Ala Gln Gly Met Ala Ser Pro Asn Gly Arg Cys Ala

885 890 895

Ser Phe Thr Ser Asp Ala Asp Gly Tyr Ala Pro Ser Glu Gly Ala Val

900 905 910

Gly Phe Val Leu Lys Thr Arg Ser Ala Ala Val Arg Asp Gly Asp Arg

915 920 925

Ile Leu Ala Thr Ile Arg Ala Thr Glu Ile Gly His Asn Gly Arg Ser

930 935 940

Gln Gly Leu Ala Ala Pro Asn Val Arg Ser Gln Ala Ala Ala His Arg

945 950 955 960

Ala Val Leu Arg Arg Ala Arg Leu Asp Pro Ser Glu Ile Asp Phe Ile

965 970 975

Glu Ala His Gly Thr Gly Thr Thr Leu Gly Asp Leu Cys Glu Val Gln

980 985 990

Gly Ile Asn Asp Ser Phe Val Ser Pro Lys Lys Arg Ala Asn Pro Leu

995 1000 1005

Val Val Ser Ala Ser Lys Ser Thr Ile Gly His Thr Glu Pro Ser

1010 1015 1020

Ala Gly Leu Val Gly Ile Leu Ser Ala Leu Met Ser Phe Glu Lys

1025 1030 1035

Arg Ile Val Pro Arg Leu Ala Tyr Leu Thr Glu Ser Asn Val Asn

1040 1045 1050

Pro Ala Leu Asp Ala Ser Val Val Pro Leu His Phe Pro Thr Lys

1055 1060 1065

His Ile Glu Leu Arg Ala Asp Val Pro Tyr Lys Ala Val Val Met

1070 1075 1080

Ser Tyr Gly Phe Ala Gly Thr Leu Ala Asp Ile Val Leu Glu Ser

1085 1090 1095

Glu Val Pro Gln Pro Thr Pro Ala Val Ala Gln Asp Thr Ala Gly

1100 1105 1110

Gln Gln Pro Met Leu Phe Val Leu Ser Ala Lys Thr Pro Arg Ala

1115 1120 1125

Leu Ala Ala Tyr Ile Glu Leu Tyr Leu Gly Phe Leu Arg His Ala

1130 1135 1140

Asp Pro Gly Leu Phe Ala Arg Ile Cys Tyr Thr Ala Cys Val Ala

1145 1150 1155

Arg Glu His Tyr Lys His Arg Val Ala Cys Val Ala Thr Asp Leu

1160 1165 1170

Val Asp Leu Ile Ala Gln Leu Glu Thr Arg Leu Val Gln Thr Ala

1175 1180 1185

Tyr Ala Gly Gly Gly Gly Ala Arg Ala Ala Arg Thr Gly Pro Leu

1190 1195 1200

Val Phe Ala Phe Ser Gly Gln Gly Thr Gln Phe Pro Ala Met Ala

1205 1210 1215

Ala Pro Leu Ala Arg Arg Tyr Ala Arg Phe Gly Glu Ile Val Gly

1220 1225 1230

Gly Cys Ala Arg Met Ala Arg Glu Leu Ser Gly Phe Pro Val Asp

1235 1240 1245

Gly Ile Leu Leu Gly Asp Asp Val Thr Pro Val Lys Asp Asn Ser

1250 1255 1260

Ala Ala Ala Glu Val His Ser Glu Val Asp Gln Ile Cys Ile Phe

1265 1270 1275

Val Tyr Gln Tyr Ala Met Cys Arg Trp Leu Gly Glu Leu Gly Val

1280 1285 1290

Glu Pro Lys Ala Ala Ile Gly His Ser Leu Gly Glu Ile Thr Ala

1295 1300 1305

Ala Val Ile Ala Gly Ala Leu Pro Phe Glu Ala Ala Leu Asp Leu

1310 1315 1320

Val Val Thr Arg Ala Arg Leu Leu Lys Pro Cys Ala Glu Gln Pro

1325 1330 1335

Ser Gly Met Ala Ala Leu Ala Cys Thr Pro Asp Val Ala Ser Lys

1340 1345 1350

Leu Thr Leu Gly Ala Ser Val Ser Val Ser Val Tyr Asn Gly Pro

1355 1360 1365

Gln Ser Ile Cys Leu Ser Gly Ala Ser Ala Glu Leu Asp Asp Ala

1370 1375 1380

Val Arg Ala Ala Lys Ala Arg Asn Ile Lys Ala Thr Arg Leu Gln

1385 1390 1395

Val Asp Gln Gly Phe His Ser Pro Cys Val Asp Ala Ala Val Pro

1400 1405 1410

Gly Leu Gln Ala Trp Cys Ala Ala His Arg Ala Ser Ala Ala Pro

1415 1420 1425

Leu Lys Met Pro Leu Tyr Ser Thr Val Arg Gly Asp Val Val Pro

1430 1435 1440

Lys Gly Ala Ala Leu Asp Pro Glu His Trp Val Ala His Ala Arg

1445 1450 1455

Asn Pro Val Leu Phe Ala Gln Thr Ala Gln Ala Leu Lys Glu Ser

1460 1465 1470

Leu Pro His Gly Ile Thr Leu Asp Val Gly Pro Gln Ala Val Ala

1475 1480 1485

Trp Ser Leu Leu Leu Leu Asn Gly Leu Ser Ala Thr Arg Thr Val

1490 1495 1500

Ala Ala Gly Ala Lys Lys Gly Ala Asp Gln Glu Arg Ala Leu Leu

1505 1510 1515

Gly Ala Leu Gly Ala Leu Phe Glu Gln His Lys Val Thr Pro Asp

1520 1525 1530

Phe Gly Arg Leu Tyr Ala Pro Leu Glu Lys Thr Arg Ile Pro Thr

1535 1540 1545

Tyr Pro Phe Glu Arg Ala Arg Cys Tyr Pro Thr Phe Ile Pro Ser

1550 1555 1560

Arg Phe Ala His Gly Ala Ala Ala Ala Ala Lys Thr Asp Gly Glu

1565 1570 1575

Val Leu Ser Ala Glu Glu Glu Asn Val Ala Pro Val Gly Leu Leu

1580 1585 1590

Thr Lys Glu Asp Leu Arg Ala Ala Leu Val Ala Cys Leu Arg Ala

1595 1600 1605

Thr Leu Glu Leu Arg Pro Asp Glu Glu Leu Asp Glu Ala Glu Pro

1610 1615 1620

Leu Thr Val His Gly Val Asp Ser Ile Gly Phe Ala Lys Leu Arg

1625 1630 1635

Lys His Val Glu Asp Arg Trp Gly Leu Asp Ile Pro Val Val Tyr

1640 1645 1650

Trp Ser Asp Ala Phe Thr Val Gly Glu Met Leu Gly Asn Leu Val

1655 1660 1665

Gly Gln Tyr Asp Val Val Ser Thr Ala Ala

1670 1675

<210> 46

<211> 4956

<212> DNA

<213> Mycena chlorophos

<400> 46

atgaatccgc cctcgtctat cctcgaagtc ttccagcgta ccgccctcga cccctcgggc 60

gccgaccggc gcgttctgga atgcgggccc gacttttgga cctacgcggg tctcgacgcg 120

gtttccacag gccttgcggc ggacttggct gctctcggag attctcccat cgtggctgtc 180

gtcgctgaga accatccctt cgtcttcgca ctcatgtttg ccgtctggaa attgcacggg 240

acgtttgttc ccatcgacgc gcatattccg tggaacctgc tcgacggcat gttggacatc 300

gtcaagccga cttgcatgtt tctcgtcgag tcggatacca acaatatctc gaacaccaag 360

gcccagggag tcgacttcgc tgtgcgcctc ttcggaggag aaggattcac catcccggcg 420

ctctcggcca aatatgcggg gaacgtctcg aatggcgccc ccgagtcact cccttctcca 480

gacgcgactg ctctgtatct gttcacgtcg tctgcctcgt cgcggcacaa tttgaaggcc 540

gttccgctca cgcatcgatt cattgccgct ggctgcgaag cgaaactcgc cttctggcac 600

cgtcttcatc ctcacaaccc caccgatgcg attcgcgtgc tgggatgggc tccgttgtcg 660

cacgtcctcg cgcatatgca ggatatcggc accgcggccc tcctcaacgc cggctgctat 720

gtcttttcga cgatcccctt gtcttacacc tcagcagaaa ctcagcccgc gcaagatatc 780

acctcggctc tcattcactc cgtgctccat tacgaggtca aagcatttgc gggcctgcct 840

tttgttattg cggcattcaa ggctgcttgt gaaggcggga acgaccgtct cctagcgcaa 900

ctacgctcca tgaccatgct cgagtgtggc ggggcgcagt tggacaagga catcgtggat 960

tgggcagtca agcaagcgat cccgctcgtg gttggggtcg ggatgacgga aacgggtggc 1020

gcgatactgg cgggccccgt cggggatgcg tcggatgggt ttcaccccca agggctgctg 1080

ctagatgcac agttctccct tatcggcaat gacgatgaat cggaaggcga gctggtcatc 1140

aagagtccca atcttccgcg cggatatctg aagtacgagg acggctcgtt cgacatcgac 1200

gcgcagggtg tcgtcacgtt caagactgga gacatctacc gcaaaagtgt cgagggtaaa 1260

ctcctttggg ttgggcgtag tacggatttc atccagatgg ctaccggcga gacgctcgat 1320

ccccgtcgtg ttgaacgggc gctacgcttc gcatcgggga tcaacgatgc ctgcatcatc 1380

gggaatgcgt tcctgaatgg ctcctctacc gcaatttacg ccatcatcga gctcgctccg 1440

cgcaccgtca acatcaataa tgattcaaat gtctccatc tgcaggtggt tgcccgcgcc 1500

ctgtctccta tcaaccgcga ccttcctccc gcattacgca ttgtgttgtc ttctgtattg 1560

attctggctg aaggcatgaa gattccgagg acaaagaaag gcgaaatctt ccggaagaag 1620

attgatgaag ttttcggagc tgctctccgg gctttaggtc actcggcaac tccaacagag 1680

gttgttcttg agcaggaacc agcggcagcc agcaaaccca tttttgacaa gaacaagctg 1740

cagactgcta ttgcgcatag cttgggtctg gatattctgg agattgacct gctggacaag 1800

ctgacctttg ctgagcttgg catgacatct attcttgcaa ttcgggttgc agaagatctc 1860

aacaaattgc tgcagggaca agttaccctg ccagtcaata tctgctacct ctatccagat 1920

gcccagctgt tgtttgcagc agttcaggaa cagctgctca agcagcagca cccttcaacc 1980

ccaactgctc cctccgtgcc ggctttgctg tcagcaactt cctctgttcc aattctcttg 2040

caggaactag atgatgttgt cattgttggc aaatcattcc ggttgcctgg cggaatctac 2100

gatgatcgag cactctgggc agctctcacc aatcaagcta cccgaaaccc catctcatat 2160

atttcgggcc agcgctggga ccatacaagc ttttacccag ctgatattgc attcttgcag 2220

gcagggttgc ttgactccga ccactttacg gattttgatg cagctttctt cgggatgacc 2280

gagaaagagg catactatct gtccccgacc atgcggcttg ctcttgaagt agcctttgag 2340

gctctagaag atgcaaatat tcctgttggt caggtgaaag gcactagcat gggagtatat 2400

gcagctgtca aggatgatgg attcgaaacc cttttgaatg ctgctcatgg gtatgatgcc 2460

tacacacgat tctacggaac tggacgggca ccaagtacca ctagcgggcg gatcagtcaa 2520

ggagaatcag cgattgtctg ctccagcaat acacactgtt ggcctggctc cttcatgttc 2580

ttgactgccc aaggaatggt gtctcctcat ggacgttgtg cctccttcag tgctcaagca 2640

gatggatatg ttccttcaga gggtgctgtt gcatttatcc tgaagacccg caaagcagca 2700

gttcgggatg gaaaccaaat ttttgccaca attcgggctg cggtggtatc acacaatggt 2760

cgatcacaag gtcttgcagc accaaacatt caagcccaat ccgagttgca tcaacaagca 2820

ttgcagaagg caaatatcca acccactgat attcattttg tggaaactca tggaacaggg 2880

acttcgcttg gggatgtctg tgagattcat gggataaatg ctgcttttgc agcaggtcac 2940

cgtccctctg gacctctcat cattagtgca agcaaaggca ctattggaca tacagagcct 3000

tctgcaggtc ttgtgggcat catggcggca ctgctctcct tcaagcatgg ccttgttcct 3060

gggctgatcc atacatctca tgggcaactc aacccggcac ttgatcaatc caaagttccg 3120

cttatcttca gcccacaaac aatttccctg ggcggagaaa agccttacag atctgtggtc 3180

atgtcatatg gctttgcagg cacactagcg gatattgtgc ttgaaggccc tgctgaggag 3240

gctttttccg ggccaggcaa aaacagcagt gctcctccgc ctatgatctt tgccctcagt 3300

gccaaatctg catcagccct ccaagaatac aagcagaagt acatcacctt cctgcagaat 3360

gttggctctg gaggccaact gttcagcaag atctgtttga cttcttgcat tgcccgagag 3420

cactacaagc atagattctc ctgtgctgct cagaacacac tggatcttct tctgcagcta 3480

gagcactctg ttgctgccag ccacaaacct ccaacaactc gtaccggaac agtcaccttt 3540

gctttctctg gacagggagc ccaattcccc agcatggatg cagctctggc tcaaggctac 3600

tctgccttca aatccatctt gctggaactt ggaaacaagg ctgccaaact ctctggattc 3660

cccatcactg attgcctgtt ggcaacaaca gcatcagctg atgaagaagc cgtccatagt 3720

gaggtggacc aaatttgcat ttttgttcat caatatgcaa tggctctttt cctcgagatg 3780

ctaggaattg tccccggtgc tgccataggc cacagcttgg gagagatcac agcagcggtc 3840

gttgctggtg gactttcgtt tgaacttggc ctagagttgg tcatcctccg tgcacatctg 3900

ctccgtccag agcagaacaa gcccgctggc atggctgcct tggcatgctc agaagcggac 3960

ttcctcaagt ttccgtccac cgatgcaact atttctgttt tcaactctcc tcggagcatt 4020

gcagtctctg gagcagcaag ctccattgag acagttctta ctgctgccaa agagcagaat 4080

atcaaggcca cgaagctcag ggttgatcaa ggattccata gcagctatgt ggagcatgcg 4140

cttcccgggc tcaagcactg gtcagcaatg aattcaggct ccttccaggc actcaggatt 4200

ccactctatt caactgcact tggccatgtt gttcctgctg gagagaccct tcagccagat 4260

cactggatga accatacccg caatgctgtt cattttacgc aaactgcgca ggctctgaaa 4320

gagtcccttc cgcatgggat cactttggat cttggtcctc aggctgtagc tcaaactctc 4380

ctgctggcca atgaccatcc tgttggccgc accattggat tgtgtggcaa acgcacagga 4440

gatcaaagac atgcattcct gctcgctctt gctgagcttt accagcagca tggtcttgtg 4500

cccaactttc atgcacttta tggcgtagct gcccaggatc tcaaggacca tctcaccagc 4560

ctgccaacat atccattcca acgtgtccgc tgctatccca gctacattcc atcccgtcac 4620

tccaacactc ccgggaccac cgtggtgatt gatgcaaagc cgcgggatga agtgaaacct 4680

gtggcagagg tctcaaagtc ggacacggat tcttccacat cattttcttc gaccattctc 4740

ttccacattc gctccatcct tgagcttcgc ccccatgagg ttctggatac gtctgaatcc 4800

ctcttgacgt acggggtcga ctcgattggg tttgcagcac tgcagaaggc cctggagcag 4860

cagcatgggc taaacttgtc gattgtgttc tggagcgacg tgttttccat tgccgacatt 4920

gtgaagaatc ttgaggagca gaagagcttg aagatg 4956

<210> 47

<211> 1652

<212>PRT

<213> Mycena chlorophos

<400> 47

Met Asn Pro Pro Ser Ser Ile Leu Glu Val Phe Gln Arg Thr Ala Leu

1 5 10 15

Asp Pro Ser Gly Ala Asp Arg Arg Val Leu Glu Cys Gly Pro Asp Phe

20 25 30

Trp Thr Tyr Ala Gly Leu Asp Ala Val Ser Thr Gly Leu Ala Ala Asp

35 40 45

Leu Ala Ala Leu Gly Asp Ser Pro Ile Val Ala Val Val Ala Glu Asn

50 55 60

His Pro Phe Val Phe Ala Leu Met Phe Ala Val Trp Lys Leu His Gly

65 70 75 80

Thr Phe Val Pro Ile Asp Ala His Ile Pro Trp Asn Leu Leu Asp Gly

85 90 95

Met Leu Asp Ile Val Lys Pro Thr Cys Met Phe Leu Val Glu Ser Asp

100 105 110

Thr Asn Asn Ile Ser Asn Thr Lys Ala Gln Gly Val Asp Phe Ala Val

115 120 125

Arg Leu Phe Gly Gly Glu Gly Phe Thr Ile Pro Ala Leu Ser Ala Lys

130 135 140

Tyr Ala Gly Asn Val Ser Asn Gly Ala Pro Glu Ser Leu Pro Ser Pro

145 150 155 160

Asp Ala Thr Ala Leu Tyr Leu Phe Thr Ser Ser Ala Ser Ser Arg His

165 170 175

Asn Leu Lys Ala Val Pro Leu Thr His Arg Phe Ile Ala Ala Gly Cys

180 185 190

Glu Ala Lys Leu Ala Phe Trp His Arg Leu His Pro His Asn Pro Thr

195 200 205

Asp Ala Ile Arg Val Leu Gly Trp Ala Pro Leu Ser His Val Leu Ala

210 215 220

His Met Gln Asp Ile Gly Thr Ala Ala Leu Leu Asn Ala Gly Cys Tyr

225 230 235 240

Val Phe Ser Thr Ile Pro Leu Ser Tyr Thr Ser Ala Glu Thr Gln Pro

245 250 255

Ala Gln Asp Ile Thr Ser Ala Leu Ile His Ser Val Leu His Tyr Glu

260 265 270

Val Lys Ala Phe Ala Gly Leu Pro Phe Val Ile Ala Ala Phe Lys Ala

275 280 285

Ala Cys Glu Gly Gly Asn Asp Arg Leu Leu Ala Gln Leu Arg Ser Met

290 295 300

Thr Met Leu Glu Cys Gly Gly Ala Gln Leu Asp Lys Asp Ile Val Asp

305 310 315 320

Trp Ala Val Lys Gln Ala Ile Pro Leu Val Val Gly Val Gly Met Thr

325 330 335

Glu Thr Gly Gly Ala Ile Leu Ala Gly Pro Val Gly Asp Ala Ser Asp

340 345 350

Gly Phe His Pro Gln Gly Leu Leu Leu Asp Ala Gln Phe Ser Leu Ile

355 360 365

Gly Asn Asp Asp Glu Ser Glu Gly Glu Leu Val Ile Lys Ser Pro Asn

370 375 380

Leu Pro Arg Gly Tyr Leu Lys Tyr Glu Asp Gly Ser Phe Asp Ile Asp

385 390 395 400

Ala Gln Gly Val Val Thr Phe Lys Thr Gly Asp Ile Tyr Arg Lys Ser

405 410 415

Val Glu Gly Lys Leu Leu Trp Val Gly Arg Ser Thr Asp Phe Ile Gln

420 425 430

Met Ala Thr Gly Glu Thr Leu Asp Pro Arg Arg Val Glu Arg Ala Leu

435 440 445

Arg Phe Ala Ser Gly Ile Asn Asp Ala Cys Ile Ile Gly Asn Ala Phe

450 455 460

Leu Asn Gly Ser Ser Thr Ala Ile Tyr Ala Ile Ile Glu Leu Ala Pro

465 470 475 480

Arg Thr Val Asn Ile Asn Asn Asp Ser Asn Val Ser His Leu Gln Val

485 490 495

Val Ala Arg Ala Leu Ser Pro Ile Asn Arg Asp Leu Pro Pro Ala Leu

500 505 510

Arg Ile Val Leu Ser Ser Val Leu Ile Leu Ala Glu Gly Met Lys Ile

515 520 525

Pro Arg Thr Lys Lys Gly Glu Ile Phe Arg Lys Lys Ile Asp Glu Val

530 535 540

Phe Gly Ala Ala Leu Arg Ala Leu Gly His Ser Ala Thr Pro Thr Glu

545 550 555 560

Val Val Leu Glu Gln Glu Pro Ala Ala Ala Ser Lys Pro Ile Phe Asp

565 570 575

Lys Asn Lys Leu Gln Thr Ala Ile Ala His Ser Leu Gly Leu Asp Ile

580 585 590

Leu Glu Ile Asp Leu Leu Asp Lys Leu Thr Phe Ala Glu Leu Gly Met

595 600 605

Thr Ser Ile Leu Ala Ile Arg Val Ala Glu Asp Leu Asn Lys Leu Leu

610 615 620

Gln Gly Gln Val Thr Leu Pro Val Asn Ile Cys Tyr Leu Tyr Pro Asp

625 630 635 640

Ala Gln Leu Leu Phe Ala Ala Val Gln Glu Gln Leu Leu Lys Gln Gln

645 650 655

His Pro Ser Thr Pro Thr Ala Pro Ser Val Pro Ala Leu Leu Ser Ala

660 665 670

Thr Ser Ser Val Pro Ile Leu Leu Gln Glu Leu Asp Asp Val Val Ile

675 680 685

Val Gly Lys Ser Phe Arg Leu Pro Gly Gly Ile Tyr Asp Asp Arg Ala

690 695 700

Leu Trp Ala Ala Leu Thr Asn Gln Ala Thr Arg Asn Pro Ile Ser Tyr

705 710 715 720

Ile Ser Gly Gln Arg Trp Asp His Thr Ser Phe Tyr Pro Ala Asp Ile

725 730 735

Ala Phe Leu Gln Ala Gly Leu Leu Asp Ser Asp His Phe Thr Asp Phe

740 745 750

Asp Ala Ala Phe Phe Gly Met Thr Glu Lys Glu Ala Tyr Tyr Leu Ser

755 760 765

Pro Thr Met Arg Leu Ala Leu Glu Val Ala Phe Glu Ala Leu Glu Asp

770 775 780

Ala Asn Ile Pro Val Gly Gln Val Lys Gly Thr Ser Met Gly Val Tyr

785 790 795 800

Ala Ala Val Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Ala His

805 810 815

Gly Tyr Asp Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser

820 825 830

Thr Thr Ser Gly Arg Ile Ser Gln Gly Glu Ser Ala Ile Val Cys Ser

835 840 845

Ser Asn Thr His Cys Trp Pro Gly Ser Phe Met Phe Leu Thr Ala Gln

850 855 860

Gly Met Val Ser Pro His Gly Arg Cys Ala Ser Phe Ser Ala Gln Ala

865 870 875 880

Asp Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys Thr

885 890 895

Arg Lys Ala Ala Val Arg Asp Gly Asn Gln Ile Phe Ala Thr Ile Arg

900 905 910

Ala Ala Val Val Ser His Asn Gly Arg Ser Gln Gly Leu Ala Ala Pro

915 920 925

Asn Ile Gln Ala Gln Ser Glu Leu His Gln Gln Ala Leu Gln Lys Ala

930 935 940

Asn Ile Gln Pro Thr Asp Ile His Phe Val Glu Thr His Gly Thr Gly

945 950 955 960

Thr Ser Leu Gly Asp Val Cys Glu Ile His Gly Ile Asn Ala Ala Phe

965 970 975

Ala Ala Gly His Arg Pro Ser Gly Pro Leu Ile Ile Ser Ala Ser Lys

980 985 990

Gly Thr Ile Gly His Thr Glu Pro Ser Ala Gly Leu Val Gly Ile Met

995 1000 1005

Ala Ala Leu Leu Ser Phe Lys His Gly Leu Val Pro Gly Leu Ile

1010 1015 1020

His Thr Ser His Gly Gln Leu Asn Pro Ala Leu Asp Gln Ser Lys

1025 1030 1035

Val Pro Leu Ile Phe Ser Pro Gln Thr Ile Ser Leu Gly Gly Glu

1040 1045 1050

Lys Pro Tyr Arg Ser Val Val Met Ser Tyr Gly Phe Ala Gly Thr

1055 1060 1065

Leu Ala Asp Ile Val Leu Glu Gly Pro Ala Glu Glu Ala Phe Ser

1070 1075 1080

Gly Pro Gly Lys Asn Ser Ser Ala Pro Pro Pro Met Ile Phe Ala

1085 1090 1095

Leu Ser Ala Lys Ser Ala Ser Ala Leu Gln Glu Tyr Lys Gln Lys

1100 1105 1110

Tyr Ile Thr Phe Leu Gln Asn Val Gly Ser Gly Gly Gln Leu Phe

1115 1120 1125

Ser Lys Ile Cys Leu Thr Ser Cys Ile Ala Arg Glu His Tyr Lys

1130 1135 1140

His Arg Phe Ser Cys Ala Ala Gln Asn Thr Leu Asp Leu Leu Leu

1145 1150 1155

Gln Leu Glu His Ser Val Ala Ala Ser His Lys Pro Pro Thr Thr

1160 1165 1170

Arg Thr Gly Thr Val Thr Phe Ala Phe Ser Gly Gln Gly Ala Gln

1175 1180 1185

Phe Pro Ser Met Asp Ala Ala Leu Ala Gln Gly Tyr Ser Ala Phe

1190 1195 1200

Lys Ser Ile Leu Leu Glu Leu Gly Asn Lys Ala Ala Lys Leu Ser

1205 1210 1215

Gly Phe Pro Ile Thr Asp Cys Leu Leu Ala Thr Thr Ala Ser Ala

1220 1225 1230

Asp Glu Glu Ala Val His Ser Glu Val Asp Gln Ile Cys Ile Phe

1235 1240 1245

Val His Gln Tyr Ala Met Ala Leu Phe Leu Glu Met Leu Gly Ile

1250 1255 1260

Val Pro Gly Ala Ala Ile Gly His Ser Leu Gly Glu Ile Thr Ala

1265 1270 1275

Ala Val Val Ala Gly Gly Leu Ser Phe Glu Leu Gly Leu Glu Leu

1280 1285 1290

Val Ile Leu Arg Ala His Leu Leu Arg Pro Glu Gln Asn Lys Pro

1295 1300 1305

Ala Gly Met Ala Ala Leu Ala Cys Ser Glu Ala Asp Phe Leu Lys

1310 1315 1320

Phe Pro Ser Thr Asp Ala Thr Ile Ser Val Phe Asn Ser Pro Arg

1325 1330 1335

Ser Ile Ala Val Ser Gly Ala Ala Ser Ser Ile Glu Thr Val Leu

1340 1345 1350

Thr Ala Ala Lys Glu Gln Asn Ile Lys Ala Thr Lys Leu Arg Val

1355 1360 1365

Asp Gln Gly Phe His Ser Ser Tyr Val Glu His Ala Leu Pro Gly

1370 1375 1380

Leu Lys His Trp Ser Ala Met Asn Ser Gly Ser Phe Gln Ala Leu

1385 1390 1395

Arg Ile Pro Leu Tyr Ser Thr Ala Leu Gly His Val Val Pro Ala

1400 1405 1410

Gly Glu Thr Leu Gln Pro Asp His Trp Met Asn His Thr Arg Asn

1415 1420 1425

Ala Val His Phe Thr Gln Thr Ala Gln Ala Leu Lys Glu Ser Leu

1430 1435 1440

Pro His Gly Ile Thr Leu Asp Leu Gly Pro Gln Ala Val Ala Gln

1445 1450 1455

Thr Leu Leu Leu Ala Asn Asp His Pro Val Gly Arg Thr Ile Gly

1460 1465 1470

Leu Cys Gly Lys Arg Thr Gly Asp Gln Arg His Ala Phe Leu Leu

1475 1480 1485

Ala Leu Ala Glu Leu Tyr Gln Gln His Gly Leu Val Pro Asn Phe

1490 1495 1500

His Ala Leu Tyr Gly Val Ala Ala Gln Asp Leu Lys Asp His Leu

1505 1510 1515

Thr Ser Leu Pro Thr Tyr Pro Phe Gln Arg Val Arg Cys Tyr Pro

1520 1525 1530

Ser Tyr Ile Pro Ser Arg His Ser Asn Thr Pro Gly Thr Thr Val

1535 1540 1545

Val Ile Asp Ala Lys Pro Arg Asp Glu Val Lys Pro Val Ala Glu

1550 1555 1560

Val Ser Lys Ser Asp Thr Asp Ser Ser Thr Ser Phe Ser Ser Thr

1565 1570 1575

Ile Leu Phe His Ile Arg Ser Ile Leu Glu Leu Arg Pro His Glu

1580 1585 1590

Val Leu Asp Thr Ser Glu Ser Leu Leu Thr Tyr Gly Val Asp Ser

1595 1600 1605

Ile Gly Phe Ala Ala Leu Gln Lys Ala Leu Glu Gln Gln His Gly

1610 1615 1620

Leu Asn Leu Ser Ile Val Phe Trp Ser Asp Val Phe Ser Ile Ala

1625 1630 1635

Asp Ile Val Lys Asn Leu Glu Glu Gln Lys Ser Leu Lys Met

1640 1645 1650

<210> 48

<211> 5313

<212> DNA

<213> Armillaria gallica

<400> 48

atggaggccg acggtcacca ctctcttctc gatgtctttc tcagcgttgc acatgattct 60

gagaagtcca aacgtaatgt cttggaatgc ggccaggata cctggacata ctcagatttg 120

gacatcatct cgtcggcctt ggcacaggat ctcaaagcta ccttgggttg ttttcccaaa 180

gttgcagtcg tcagcgagaa ccatccctac gtgtttgctc tcatgctggc cgtttggaag 240

cttgaaggga tattcatccc catcgacgtc catattacag ctgaccttct aaagggcatg 300

ctacgcattg ttgcccccac ttgtctggcg atcccagaga ccgatatttc caaccagcgt 360

gttgcctctg cgattggtat acatgttctc cccttcaacg tgaatgcgtc gaccatgaat 420

gcacttcgac agaaatacga cccatttact cagaatgcct cgctatctgg atgcgcactt 480

ccttacgttg atcgcgcatg cctctatctc tttacatcct ccgcgtcctc tactgccaac 540

ttgaaatgtg tacctttgac ccatactctt atcctcagta actgccgttc caagctcgca 600

tggtggcggc gcgttcgtcc ggaaggcgaa atggatggga tacgtgttct agggtgggca 660

ccttggtcgc atatccttgc ctacatgcaa gatattggca cggcaacgtt cctgaatgcc 720

ggctgctatg tctttgcgtc cgttccatcc acatatcctt cacagctggc agcgaatggc 780

ctacaaggcc ccaccatgaa tatcatcgat tcacttcttg aacggcgagt cgccgcattt 840

gcttgcgtac cgttcatttt gagcgaacta aaagctatgt gcgagacggc tgccagtcca 900

gatgacaagc atctcatgtg cttgagagct gaggagaaag ttcgccttgt cagtgcgctg 960

cagcggctta tgatgctcga gtgcggaggc gctgcgctcg agtcggatgt cacacgttgg 1020

gccgtcgaaa atggcatatc ggtcatggtc ggcatcggga tgacggagac agtcggtacg 1080

ctgtttgcag agcgcgcgca agacgcctgt tccaatggct attctgcgca ggacgccctc 1140

attgctgatg gcatcatgtc actggtcggg tctgacaacg aggaagccac cttcgaaggg 1200

gaactagtcg tgaagagcaa gctcatccca cacggataca tcaactaccg tgattcgtcg 1260

ttttcggtgg actcggacgg ctgggtaacg ttcaaaacgg gagacaaata tcagcgcacg 1320

ccagatggac gattcaaatg gcttggaaga aagaccgatt ttattcagat gacaagcagc 1380

gaaacattgg atcccagacc cattgagcaa gccctctgtg cgaatccaag tatcgcaaat 1440

gcatgcgtca ttggtgacag gttcctgaga gagcctgcga ctagcgtatg cgccattgtc 1500

gagatcgggc ctgaagtgga catgccttcg tccaagatcg acagagaaat tgcgaatgcc 1560

ctcgctccaa tcaatcgcga ccttcctcct gctcttcgca tatcatggtc tcgcgtactc 1620

ataattcgac ctcctcagaa aataccagtt acgaggaaag gcgatgtgtt ccgtaagaag 1680

attgaagata tgtttggggcc tttccttggt gtcggcgttt ctaccgaggt cgaagccggc 1740

catgaaacta aagaagatga cacggaacac atcgtgagac aggttgtgag caatcttctt 1800

ggcgtccatg atcctgagct attgtctgct ttatctttcg ctgagctggg catgacctca 1860

tttatggctg ttagcatcgt caacgctctc aacaaacgca tcggcggcct cacccttccg 1920

tctaatgcat gctacatcca tattgatctt gattctcttg tggacgcgat ttcacttgaa 1980

tatggtcatg gaagtatccc tgcagagttg ccttctaacc ctttccccga catcgagtcc 2040

catcagcata atgataagga cattgtgata gtcggcaagg cattccgttt acccggctca 2100

ctcaacagta ctgcaactct ctgggaagct ttgttatcga ataacaattc ggtcatcagt 2160

gatatcccat ctgatcgctg ggatcatgca agcttttacc cccacaatat atgtttcacg 2220

aaagcaggcc tcgtcgatgt tgcacattac gactacagat tttttggcct catggcgaca 2280

gaggcgttgt acgtatctcc gacgatgcgc ctcgccttgg aagtgtcatt tgaagccctg 2340

gagaacgcaa atattccgct atccaagctg aaggggacac aaaccgctgt ctatgtcgcc 2400

actaaagacg atggcttcga gacactctta aatgccgagc aaggttacga tgcgtacacg 2460

cgattctacg gcacgggtcg cgctccgagt accgcaagcg gtcgcataag ctatcttctt 2520

gatattcatg ggccatctat taccgttgat acagcatgca gcggaggcat tgtatgtatg 2580

gatcaagcca tcactttctt gcaatccgga ggggccgata ccgcaattgt ctgttcgacc 2640

aatacgcact gttggccggg atcattcatg ttcctgacgg cacaaggcat ggtttctcca 2700

aatggaagat gcgctacatt cactaccgat gcagacggat atgtgccttc ggagggtgca 2760

gtggctttca ttctcaagac gcgcagcgct gcaatacgcg acaatgacaa tatactcgcc 2820

gtgatcaaat caacagatgt gtcccataac ggccgttctc aagggctagt tgcaccaaac 2880

gtaaaggcgc aggcaaacct acaccggtcg ttgctacgaa aagctgggct gtttcctgat 2940

caaattaact ttatcgaagc tcatggaaca ggtacatctc taggagacct ttcggaaatc 3000

cagggcatta ataacgccta catctcatca cgacctcgtc tggccggtcc ccttatcatt 3060

agtgcatcga aaacagtttt aggacacagt gaaccaacag cagggatggc cggcatcctc 3120

acagccttgc ttgcccttga gaaagagaca gttcctggtt taaatcactt aacggatcac 3180

aacctcaacc cttcgcttga ttgcagcgta gttcctctcc tgattcctca cgagtctatt 3240

cacattggtg gtgcaaagcc acatcgagct gcggttctgt catacggctt cgcgggtacg 3300

ctggccggtg ctatcttaga aggaccacct tctggtgtac catggccgtc gtcaaatgat 3360

atacaagaac accctatgat tttcgtcgtc agtgggaaaa ccgtgcctac actggaagcg 3420

tacctgggac ggtatttgac atttttgcgg gtcgcaaaaa cacacgactt caggacatc 3480

tgctacacca cttgcgtcgg gagggagcac tacaaatacc ggttctcctg cgttgcccga 3540

aacatggcag accttatttc tcaaattgaa catcgactga caactctttc cacttcgaaa 3600

cagaagcctc gcggctcgtt agggttcatg ttctcaggac aaggcacttg tttccctggt 3660

atggcttcag cacttgctga acaatattcg gggttccgaa tgctcgtctc taagtttggg 3720

caggctgccc aagagcggtc cggttatccg atcgataggc tgttgcttga agtttctgat 3780

acattaccag aaacaaacag cgaggtcgac caaatttgca tttttgtcta ccaatattct 3840

gttctgcaat ggttgcaatg tctaggcatt caaccgaaag cagtcctcgg tcacagcctg 3900

ggagaaatta ctgccgcagt cgcagctggc gccctttcgt tcgaatctgc gttggatctt 3960

gtggtcaccc gtgctcgtct tctccgtccc agaacaaaag actctgcagg aatggccgca 4020

gtagcagcgt ccaaggaaga agttgaagga gttatagaaa ccctccaact tgcaaactcg 4080

ctaagcgttg cggttcacaa cggtccgcgg agtgttgttg tgtcaggcgc atcagcagaa 4140

atcgatgccc tagttgtcgc agctaaagaa cggggcttga aagcctcccg cttaaaggtt 4200

gaccaaggct tccacagccc ttacgttgac tctgcggttc cgggtttact cgactggtca 4260

aacaagcatc gttcgacctt ctttccttta aacattcctc tatactcgac tttgaccggc 4320

gagcttattc cgaagggacg gaggttcgtc tcggatcact gggtaaacca tgctcgaaaa 4380

cctgttcagt ttgcggcggc agcggcagcg gtggatgaag atcgatccat tggtgtgctc 4440

gttgacgtcg gaccccaacc cgttgcgtgg accctccttc aagcaaacaa ccttctcaat 4500

acctctgcag ttgcgctatc cgcaaaggcc ggaaaggacc aggagatggc gctgctcact 4560

gctttgagct acctcttcca agagcacaac ctttctccca acttccacga gctttactct 4620

cagcgtcatg ggactctgca gaagacggac attcccacct acccattcca acgtgtccac 4680

cgctatccga ccttcatacc gtcacgaaat caaagtcctg ctattgcaac ggtagttata 4740

ccgccacctc gcttctctgt ccaaaaggct gcggatgtag catcacagtc gaaggaatca 4800

gactgtcgag ctggtttgat cagttgcctt agagccatcc tcgaattaac accggaagag 4860

gagtttgacc tttctgagac tctcaacgct cgtggtatgg attcgatcat gtttgcgcag 4920

ctacggaagc gggttgggga agaattcaac ctcgatatac ccatgatcta tctatcagac 4980

gtgttcacga tggaacagat ggtcgactac ctcgtcgaac agtccggatc cacacccgcg 5040

tcaaagcacg tagaaacttc agctaatcaa ccattagacg aagaagatct ccggacgggg 5100

ctcttgtcat gcctgaggaa cgtgctagaa attacccccg atgaagaact tgacctatct 5160

gaaactttga atgctcgtgg tgttgactcg atcatgttcg ctcagctgcg gaaacgcgtt 5220

ggggaaggtt ttggtgtgga aattccgatg atatatctgt ctgacgtgtt taccatggaa 5280

gacatgatca atttcctcgt ctccgagcgg tcg 5313

<210> 49

<211> 1771

<212>PRT

<213> Armillaria gallica

<400> 49

Met Glu Ala Asp Gly His His Ser Leu Leu Asp Val Phe Leu Ser Val

1 5 10 15

Ala His Asp Ser Glu Lys Ser Lys Arg Asn Val Leu Glu Cys Gly Gln

20 25 30

Asp Thr Trp Thr Tyr Ser Asp Leu Asp Ile Ile Ser Ser Ala Leu Ala

35 40 45

Gln Asp Leu Lys Ala Thr Leu Gly Cys Phe Pro Lys Val Ala Val Val

50 55 60

Ser Glu Asn His Pro Tyr Val Phe Ala Leu Met Leu Ala Val Trp Lys

65 70 75 80

Leu Glu Gly Ile Phe Ile Pro Ile Asp Val His Ile Thr Ala Asp Leu

85 90 95

Leu Lys Gly Met Leu Arg Ile Val Ala Pro Thr Cys Leu Ala Ile Pro

100 105 110

Glu Thr Asp Ile Ser Asn Gln Arg Val Ala Ser Ala Ile Gly Ile His

115 120 125

Val Leu Pro Phe Asn Val Asn Ala Ser Thr Met Asn Ala Leu Arg Gln

130 135 140

Lys Tyr Asp Pro Phe Thr Gln Asn Ala Ser Leu Ser Gly Cys Ala Leu

145 150 155 160

Pro Tyr Val Asp Arg Ala Cys Leu Tyr Leu Phe Thr Ser Ser Ala Ser

165 170 175

Ser Thr Ala Asn Leu Lys Cys Val Pro Leu Thr His Thr Leu Ile Leu

180 185 190

Ser Asn Cys Arg Ser Lys Leu Ala Trp Trp Arg Arg Val Arg Pro Glu

195 200 205

Gly Glu Met Asp Gly Ile Arg Val Leu Gly Trp Ala Pro Trp Ser His

210 215 220

Ile Leu Ala Tyr Met Gln Asp Ile Gly Thr Ala Thr Phe Leu Asn Ala

225 230 235 240

Gly Cys Tyr Val Phe Ala Ser Val Pro Ser Thr Tyr Pro Ser Gln Leu

245 250 255

Ala Ala Asn Gly Leu Gln Gly Pro Thr Met Asn Ile Ile Asp Ser Leu

260 265 270

Leu Glu Arg Arg Val Ala Ala Phe Ala Cys Val Pro Phe Ile Leu Ser

275 280 285

Glu Leu Lys Ala Met Cys Glu Thr Ala Ala Ser Pro Asp Asp Lys His

290 295 300

Leu Met Cys Leu Arg Ala Glu Glu Lys Val Arg Leu Val Ser Ala Leu

305 310 315 320

Gln Arg Leu Met Met Leu Glu Cys Gly Gly Ala Ala Leu Glu Ser Asp

325 330 335

Val Thr Arg Trp Ala Val Glu Asn Gly Ile Ser Val Met Val Gly Ile

340 345 350

Gly Met Thr Glu Thr Val Gly Thr Leu Phe Ala Glu Arg Ala Gln Asp

355 360 365

Ala Cys Ser Asn Gly Tyr Ser Ala Gln Asp Ala Leu Ile Ala Asp Gly

370 375 380

Ile Met Ser Leu Val Gly Ser Asp Asn Glu Glu Ala Thr Phe Glu Gly

385 390 395 400

Glu Leu Val Val Lys Ser Lys Leu Ile Pro His Gly Tyr Ile Asn Tyr

405 410 415

Arg Asp Ser Ser Phe Ser Val Asp Ser Asp Gly Trp Val Thr Phe Lys

420 425 430

Thr Gly Asp Lys Tyr Gln Arg Thr Pro Asp Gly Arg Phe Lys Trp Leu

435 440 445

Gly Arg Lys Thr Asp Phe Ile Gln Met Thr Ser Ser Glu Thr Leu Asp

450 455 460

Pro Arg Pro Ile Glu Gln Ala Leu Cys Ala Asn Pro Ser Ile Ala Asn

465 470 475 480

Ala Cys Val Ile Gly Asp Arg Phe Leu Arg Glu Pro Ala Thr Ser Val

485 490 495

Cys Ala Ile Val Glu Ile Gly Pro Glu Val Asp Met Pro Ser Ser Lys

500 505 510

Ile Asp Arg Glu Ile Ala Asn Ala Leu Ala Pro Ile Asn Arg Asp Leu

515 520 525

Pro Pro Ala Leu Arg Ile Ser Trp Ser Arg Val Leu Ile Ile Arg Pro

530 535 540

Pro Gln Lys Ile Pro Val Thr Arg Lys Gly Asp Val Phe Arg Lys Lys

545 550 555 560

Ile Glu Asp Met Phe Gly Pro Phe Leu Gly Val Gly Val Ser Thr Glu

565 570 575

Val Glu Ala Gly His Glu Thr Lys Glu Asp Asp Thr Glu His Ile Val

580 585 590

Arg Gln Val Val Ser Asn Leu Leu Gly Val His Asp Pro Glu Leu Leu

595 600 605

Ser Ala Leu Ser Phe Ala Glu Leu Gly Met Thr Ser Phe Met Ala Val

610 615 620

Ser Ile Val Asn Ala Leu Asn Lys Arg Ile Gly Gly Leu Thr Leu Pro

625 630 635 640

Ser Asn Ala Cys Tyr Ile His Ile Asp Leu Asp Ser Leu Val Asp Ala

645 650 655

Ile Ser Leu Glu Tyr Gly His Gly Ser Ile Pro Ala Glu Leu Pro Ser

660 665 670

Asn Pro Phe Pro Asp Ile Glu Ser His Gln His Asn Asp Lys Asp Ile

675 680 685

Val Ile Val Gly Lys Ala Phe Arg Leu Pro Gly Ser Leu Asn Ser Thr

690 695 700

Ala Thr Leu Trp Glu Ala Leu Leu Ser Asn Asn Asn Ser Val Ile Ser

705 710 715 720

Asp Ile Pro Ser Asp Arg Trp Asp His Ala Ser Phe Tyr Pro His Asn

725 730 735

Ile Cys Phe Thr Lys Ala Gly Leu Val Asp Val Ala His Tyr Asp Tyr

740 745 750

Arg Phe Phe Gly Leu Met Ala Thr Glu Ala Leu Tyr Val Ser Pro Thr

755 760 765

Met Arg Leu Ala Leu Glu Val Ser Phe Glu Ala Leu Glu Asn Ala Asn

770 775 780

Ile Pro Leu Ser Lys Leu Lys Gly Thr Gln Thr Ala Val Tyr Val Ala

785 790 795 800

Thr Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Glu Gln Gly Tyr

805 810 815

Asp Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser Thr Ala

820 825 830

Ser Gly Arg Ile Ser Tyr Leu Leu Asp Ile His Gly Pro Ser Ile Thr

835 840 845

Val Asp Thr Ala Cys Ser Gly Gly Ile Val Cys Met Asp Gln Ala Ile

850 855 860

Thr Phe Leu Gln Ser Gly Gly Ala Asp Thr Ala Ile Val Cys Ser Thr

865 870 875 880

Asn Thr His Cys Trp Pro Gly Ser Phe Met Phe Leu Thr Ala Gln Gly

885 890 895

Met Val Ser Pro Asn Gly Arg Cys Ala Thr Phe Thr Thr Asp Ala Asp

900 905 910

Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys Thr Arg

915 920 925

Ser Ala Ala Ile Arg Asp Asn Asp Asn Ile Leu Ala Val Ile Lys Ser

930 935 940

Thr Asp Val Ser His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn

945 950 955 960

Val Lys Ala Gln Ala Asn Leu His Arg Ser Leu Leu Arg Lys Ala Gly

965 970 975

Leu Phe Pro Asp Gln Ile Asn Phe Ile Glu Ala His Gly Thr Gly Thr

980 985 990

Ser Leu Gly Asp Leu Ser Glu Ile Gln Gly Ile Asn Asn Ala Tyr Ile

995 1000 1005

Ser Ser Arg Pro Arg Leu Ala Gly Pro Leu Ile Ile Ser Ala Ser

1010 1015 1020

Lys Thr Val Leu Gly His Ser Glu Pro Thr Ala Gly Met Ala Gly

1025 1030 1035

Ile Leu Thr Ala Leu Leu Ala Leu Glu Lys Glu Thr Val Pro Gly

1040 1045 1050

Leu Asn His Leu Thr Asp His Asn Leu Asn Pro Ser Leu Asp Cys

1055 1060 1065

Ser Val Val Pro Leu Leu Ile Pro His Glu Ser Ile His Ile Gly

1070 1075 1080

Gly Ala Lys Pro His Arg Ala Ala Val Leu Ser Tyr Gly Phe Ala

1085 1090 1095

Gly Thr Leu Ala Gly Ala Ile Leu Glu Gly Pro Pro Ser Gly Val

1100 1105 1110

Pro Trp Pro Ser Ser Asn Asp Ile Gln Glu His Pro Met Ile Phe

1115 1120 1125

Val Val Ser Gly Lys Thr Val Pro Thr Leu Glu Ala Tyr Leu Gly

1130 1135 1140

Arg Tyr Leu Thr Phe Leu Arg Val Ala Lys Thr His Asp Phe Gln

1145 1150 1155

Asp Ile Cys Tyr Thr Thr Cys Val Gly Arg Glu His Tyr Lys Tyr

1160 1165 1170

Arg Phe Ser Cys Val Ala Arg Asn Met Ala Asp Leu Ile Ser Gln

1175 1180 1185

Ile Glu His Arg Leu Thr Thr Leu Ser Thr Ser Lys Gln Lys Pro

1190 1195 1200

Arg Gly Ser Leu Gly Phe Met Phe Ser Gly Gln Gly Thr Cys Phe

1205 1210 1215

Pro Gly Met Ala Ser Ala Leu Ala Glu Gln Tyr Ser Gly Phe Arg

1220 1225 1230

Met Leu Val Ser Lys Phe Gly Gln Ala Ala Gln Glu Arg Ser Gly

1235 1240 1245

Tyr Pro Ile Asp Arg Leu Leu Leu Glu Val Ser Asp Thr Leu Pro

1250 1255 1260

Glu Thr Asn Ser Glu Val Asp Gln Ile Cys Ile Phe Val Tyr Gln

1265 1270 1275

Tyr Ser Val Leu Gln Trp Leu Gln Cys Leu Gly Ile Gln Pro Lys

1280 1285 1290

Ala Val Leu Gly His Ser Leu Gly Glu Ile Thr Ala Ala Val Ala

1295 1300 1305

Ala Gly Ala Leu Ser Phe Glu Ser Ala Leu Asp Leu Val Val Thr

1310 1315 1320

Arg Ala Arg Leu Leu Arg Pro Arg Thr Lys Asp Ser Ala Gly Met

1325 1330 1335

Ala Ala Val Ala Ala Ser Lys Glu Glu Val Glu Gly Val Ile Glu

1340 1345 1350

Thr Leu Gln Leu Ala Asn Ser Leu Ser Val Ala Val His Asn Gly

1355 1360 1365

Pro Arg Ser Val Val Val Ser Gly Ala Ser Ala Glu Ile Asp Ala

1370 1375 1380

Leu Val Val Ala Ala Lys Glu Arg Gly Leu Lys Ala Ser Arg Leu

1385 1390 1395

Lys Val Asp Gln Gly Phe His Ser Pro Tyr Val Asp Ser Ala Val

1400 1405 1410

Pro Gly Leu Leu Asp Trp Ser Asn Lys His Arg Ser Thr Phe Phe

1415 1420 1425

Pro Leu Asn Ile Pro Leu Tyr Ser Thr Leu Thr Gly Glu Leu Ile

1430 1435 1440

Pro Lys Gly Arg Arg Phe Val Ser Asp His Trp Val Asn His Ala

1445 1450 1455

Arg Lys Pro Val Gln Phe Ala Ala Ala Ala Ala Ala Val Asp Glu

1460 1465 1470

Asp Arg Ser Ile Gly Val Leu Val Asp Val Gly Pro Gln Pro Val

1475 1480 1485

Ala Trp Thr Leu Leu Gln Ala Asn Asn Leu Leu Asn Thr Ser Ala

1490 1495 1500

Val Ala Leu Ser Ala Lys Ala Gly Lys Asp Gln Glu Met Ala Leu

1505 1510 1515

Leu Thr Ala Leu Ser Tyr Leu Phe Gln Glu His Asn Leu Ser Pro

1520 1525 1530

Asn Phe His Glu Leu Tyr Ser Gln Arg His Gly Thr Leu Gln Lys

1535 1540 1545

Thr Asp Ile Pro Thr Tyr Pro Phe Gln Arg Val His Arg Tyr Pro

1550 1555 1560

Thr Phe Ile Pro Ser Arg Asn Gln Ser Pro Ala Ile Ala Thr Val

1565 1570 1575

Val Ile Pro Pro Pro Arg Phe Ser Val Gln Lys Ala Ala Asp Val

1580 1585 1590

Ala Ser Gln Ser Lys Glu Ser Asp Cys Arg Ala Gly Leu Ile Ser

1595 1600 1605

Cys Leu Arg Ala Ile Leu Glu Leu Thr Pro Glu Glu Glu Phe Asp

1610 1615 1620

Leu Ser Glu Thr Leu Asn Ala Arg Gly Met Asp Ser Ile Met Phe

1625 1630 1635

Ala Gln Leu Arg Lys Arg Val Gly Glu Glu Phe Asn Leu Asp Ile

1640 1645 1650

Pro Met Ile Tyr Leu Ser Asp Val Phe Thr Met Glu Gln Met Val

1655 1660 1665

Asp Tyr Leu Val Glu Gln Ser Gly Ser Thr Pro Ala Ser Lys His

1670 1675 1680

Val Glu Thr Ser Ala Asn Gln Pro Leu Asp Glu Glu Asp Leu Arg

1685 1690 1695

Thr Gly Leu Leu Ser Cys Leu Arg Asn Val Leu Glu Ile Thr Pro

1700 1705 1710

Asp Glu Glu Leu Asp Leu Ser Glu Thr Leu Asn Ala Arg Gly Val

1715 1720 1725

Asp Ser Ile Met Phe Ala Gln Leu Arg Lys Arg Val Gly Glu Gly

1730 1735 1740

Phe Gly Val Glu Ile Pro Met Ile Tyr Leu Ser Asp Val Phe Thr

1745 1750 1755

Met Glu Asp Met Ile Asn Phe Leu Val Ser Glu Arg Ser

1760 1765 1770

<210> 50

<211> 5310

<212> DNA

<213> Armillaria ostoyae

<400> 50

atggaggccg acggtcacta ctctcttctc gatgtctttc tcagcgttgc acatgattct 60

gagaagtcca aacgtaatgt cttggaatgc ggccaggata cttggacata ctcggatttg 120

gacattatct cgtcggccct ggcacaggat ctcaaagcta tcttgggttg ttttcccaaa 180

gttgcagtcg tcagcgagaa ccatccctac gtatttgctc tcatgctggc cgtatggaag 240

cttgaaggga tattcatccc tatcgacgtc cacgttacag ctgaccttct aaagggcatg 300

ctacgcattg ttgctcccac ttgtctggtg atcccagaga ccgatatttt taaccagcgt 360

gttgcctctg caattggtat acatgttctc cccttcaacg tgaatgcgtc gaccatgact 420

gcacttcgac agaaatacca cccatttact cagaaagcct cgctatctgg gtgcgcactg 480

ccttacgttg atcgcgcatg cctctatctc tttacatcct ccgcgtcctc tactgccaat 540

ttgaaatgcg tacctttgac ccatactctt atcctcagta actgccgttc caagctcgca 600

tggtggcggc gcgttcgtcc ggaaggcgaa atggatgaga tacgtgttct agggtgggca 660

ccttggtcgc atatccttgc ctacatgcaa gatattggca cggcaacgct cctgaacgcc 720

ggttgctatg tctttgcgtc cgttccatcc acatatccta cacaactggc agcgaatggc 780

ttacaaggcc ctatcatgaa tatcatcgat tcacttcttg aacgacgagt cgccgcattt 840

gcttgcgtac cgttcatttt gagcgaacta aaagctatgt gcgagacggc ttccagtcca 900

gacgacaagc atcaaatgtg cttgagagct gaggagaaag ttcgccttgt cagtgcgctg 960

cagcggctta taatgctcga gtgcggaggc gctgcgcttg agtcaggtgt cacacgttgg 1020

gccgtcgaga atggcatatc agtcatggtc ggcatcggga tgacggagac agtcggtacg 1080

ctgtttgcag agcgcgcgca agacgcccgt tccaacggct attctgcgca ggacgccctc 1140

atttctgatg ggatcatgtc actggtcggg tctgacaacg aggaagccac cctcgaaggg 1200

gaactagtcg tcaagagcaa gctcatccct catggataca tcaagtaccg tgattcgtcg 1260

ttttcggtgg actcggacgg ctgggtaact ttcaaaacgg gagacaaata tcagcgcacg 1320

ccagatggac gattcaaatg gcttggaaga aagaccgatt ttattcagat gacaagcagc 1380

gaaacactgg atcccagacc cattgaggaa gccctctgtg cgaatccaag tatcgcaaat 1440

gcatgcgtca ttggtgacag gttcctgagg gagcctgcga ctagcgtatg cgccattgtc 1500

gagatcgggc cggaagtgga catgccttcg tccaagatcg acagagaaat tgcgaatacc 1560

ctcactccaa tcaatcgcgg ccttcctcct gctcttcgca tatcatggtc tcgcgtactc 1620

ataattcgac ctcctcagaa aataccagtt acgaggaaag gtgatgtgtt ccgtaagaag 1680

attgaagata tgtttgggtc tttccttggt gtcggcgttt ctaccgaggt cgaagtcgac 1740

catgaaacta aagaagatga tacgaaacac gtcgtgagac aggttgtaag caatcttctt 1800

ggagtccatg atcttgagtt attgtctgct ttatccttcg ccgagctggg catgacatca 1860

tttatggctg ttagcatcgt caacactcta aacaaacgca tcgacggcct cacccttcca 1920

cctaatgcat gctacatcca tattgatctt gattctcttg tggacgcgat ttcacttgaa 1980

catggtcatg aaagtatccc tgcagagttg ccttctaacc ctttccccgt tatcgagtcc 2040

catcaacata acgataagga cattgtgata gtcggcaagg cattccgttt acccggctca 2100

ctcaacaata ctgcatctct ctgggaagct ttgttatcga agaacagttc agtcatcagt 2160

gacatcccat ccgatcgctg ggatcacgca agcttttacc cccacgatat atgcttcacg 2220

aaagcaggcc tcgtcgatgt tgcacattac gactacagat tttttggcct cacggcgaca 2280

gaggcgttgt acgtatctcc gacgatgcgc ctcgccttgg aagtgtcatt tgaggccctg 2340

gagaacgcaa atattccact atccaagctg aaggggacgc aaaccgccgt ctatgtcgcc 2400

actaaagacg atggcttcga gacactctta aatgccgagc aaggctacga tgcatacacg 2460

cgattctacg gtacgggtcg tgctccgagt accgcaagcg gccgtataag ctatcttctt 2520

gatattcatg ggccatctgt caccgttgat acagcatgca gcggaggcat tgtatgtatg 2580

gatcaagcca tcactttctt gcaatccgga ggggccgata ccgctattgt ctgttcgagc 2640

aatacgcact gttggccggg atcattcatg ttcctgacgg cgcaaggcat ggtttctcaa 2700

aatggaagat gcgctacatt tactaccgat gcagacggat atgtaccttc ggagggtgca 2760

gtggctttca ttctcaagac gcgcagcgct gcgatacgcg acaacgacaa tatactcgcc 2820

gtgatcagat caacagacgt gtcccataac ggccgttctc aagggctagt tgcaccaaac 2880

gtaaaggcgc agacaaacct acaccggtcg ttgctacgaa aagctgggct gtttcctgat 2940

caaattaact ttatcgaagc tcatgggaca ggtacgtctc taggagacct ttcggaaatc 3000

cagggcatta ataacgccta cacctcaaca cgacctcgtc tggccggtcc ccttatcatt 3060

agcgcatcga aaacagtttt aggacacagt gaaccaacag cagggatggc cggcatcctc 3120

acagccttgc ttgcccttga gaaagagaca gttcctggtc taaatcactt aacggagcac 3180

aaccttaacc cttcgcttga ttgcagcgta gttcctctcc tgattcctca cgagtctatt 3240

cacattggtg gtgcaaagcc acatcgagct gcggttctgt catacggctt cgcgggtacg 3300

ctggccggtg ccatcttaga gggaccacct tctgatgtac caaggccgtc gtcaaataat 3360

atacaagaac accctatgat tttcgtcgtc agtgggaaaa ccgtgcctgc actggaagcg 3420

tacctaggac ggtatttggc atttttgcgg gtcgcaaaaa cacacgactt ccatgacatc 3480

tgctacacta cttgcgtcgg gagggagcac tacaaatacc ggttctcctg cgttgcccga 3540

aacatggcag accttatttc tcaaattgaa catcgactga cagctctttc cacttcgaaa 3600

cagaagcctc gcggctcgct agggttcata ttctcaggac aaggcactta tttccctggt 3660

atggctgcag cacttgccga acaatattcg gggttccgag tgctcgtctc taagtttggg 3720

caggctgccc aagagcggtc gggttatccg atcgataggc tgttgcttga agtttctgat 3780

acattgccag aaacaaacag cgaggtcgat caaatttgca tttttgtcta ccaatattct 3840

gttctgcaat ggctgcagag tctaggcatt caaccgaaag cagtcctcgg tcacagtctg 3900

ggagaaatta ctgcagcagt cgcagctggt gccctttcgt tcgaatctgc gttggacctt 3960

gtggtcaccc gtgctcgtct tctccgtcct agagcaaaag attctgcagg aatggccgca 4020

gtagcagcat ccaaggaaga agtcaaaggg cttatagaaa ccctccaact tgcggactcg 4080

ctgagcgttg cggttcataa cggtccgcgg agtgttgttg tgtcaggcgc atcagccgaa 4140

atcgacgccc tggttgtcgc agctaaagaa cggggcttga aggcctcccg cttaaaggtt 4200

gaccaaggct tccacagccc ttacgttgat tctgcggttc caggtttact cgactggtca 4260

aataagcatc gttcgacctt ccttcctttg aacattcctt tatactcgac tttgactggc 4320

gagcttattc cgaagggacg gaggttcgtc tcggatcact gggtaaacca tgctcgaaaa 4380

cctgtccagt ttgcggcggc agcggcagcc gtggatgaag accgatccat tggtgtgctc 4440

gttgacgttg gaccccaacc cgtcgcgtgg accctccttc aagcaaacaa ccttctcaat 4500

acctctgcag ttgcactatt cgcaaaggct ggaaaggatc aggagatggc gctgcttact 4560

gctttgagct acctcgtcca agagcacaac ctttctccca acttccatga gctttactct 4620

cagcgtcatg gtgctctgaa gaagacagac gttcccacct acccattccg ccgtgtccac 4680

cgctatccga ccttcatacc gtcacgaaat caaagtcctg ctgctgcgac ggtagctatg 4740

ccgccacccc gcttctctgt ccaaaagaat gcggatgtag catcacagtc gaaggaatca 4800

gactgtcgag ctggtttgat cagttgcctt agagccatcc tcgaattaac accggaagag 4860

gagtttgacc tttctgagac tctcaacgct cgtggtatgg attcgatcat gtttgcgcag 4920

ctacggaagc gggttgggga agaattcgac cttgacatac ccatgatcta tttatcagat 4980

gtgttcacga tggaacagat ggttgattac ctcgtcaaac agtccggatc cagacccgca 5040

ttaaaacacg cagaaattcc agttaatcaa ccattagacg aagatctccg gacgaggctc 5100

gtttcatgcc tgaggaacgt gctagaaatc acccccgatg aagaacttga cctatctgaa 5160

actttgaacg ctcgtggtgt tgactcgatc atgttcgctc agctacgaaa acgcgttggg 5220

gaaggatttg gtgtggaaat tccgatgata tatctgtccg acgtgtttac catggaagac 5280

atgatcaatt tcctcgtctc tgagcgctcg 5310

<210> 51

<211> 1770

<212>PRT

<213> Armillaria ostoyae

<400> 51

Met Glu Ala Asp Gly His Tyr Ser Leu Leu Asp Val Phe Leu Ser Val

1 5 10 15

Ala His Asp Ser Glu Lys Ser Lys Arg Asn Val Leu Glu Cys Gly Gln

20 25 30

Asp Thr Trp Thr Tyr Ser Asp Leu Asp Ile Ile Ser Ser Ala Leu Ala

35 40 45

Gln Asp Leu Lys Ala Ile Leu Gly Cys Phe Pro Lys Val Ala Val Val

50 55 60

Ser Glu Asn His Pro Tyr Val Phe Ala Leu Met Leu Ala Val Trp Lys

65 70 75 80

Leu Glu Gly Ile Phe Ile Pro Ile Asp Val His Val Thr Ala Asp Leu

85 90 95

Leu Lys Gly Met Leu Arg Ile Val Ala Pro Thr Cys Leu Val Ile Pro

100 105 110

Glu Thr Asp Ile Phe Asn Gln Arg Val Ala Ser Ala Ile Gly Ile His

115 120 125

Val Leu Pro Phe Asn Val Asn Ala Ser Thr Met Thr Ala Leu Arg Gln

130 135 140

Lys Tyr His Pro Phe Thr Gln Lys Ala Ser Leu Ser Gly Cys Ala Leu

145 150 155 160

Pro Tyr Val Asp Arg Ala Cys Leu Tyr Leu Phe Thr Ser Ser Ala Ser

165 170 175

Ser Thr Ala Asn Leu Lys Cys Val Pro Leu Thr His Thr Leu Ile Leu

180 185 190

Ser Asn Cys Arg Ser Lys Leu Ala Trp Trp Arg Arg Val Arg Pro Glu

195 200 205

Gly Glu Met Asp Glu Ile Arg Val Leu Gly Trp Ala Pro Trp Ser His

210 215 220

Ile Leu Ala Tyr Met Gln Asp Ile Gly Thr Ala Thr Leu Leu Asn Ala

225 230 235 240

Gly Cys Tyr Val Phe Ala Ser Val Pro Ser Thr Tyr Pro Thr Gln Leu

245 250 255

Ala Ala Asn Gly Leu Gln Gly Pro Ile Met Asn Ile Ile Asp Ser Leu

260 265 270

Leu Glu Arg Arg Val Ala Ala Phe Ala Cys Val Pro Phe Ile Leu Ser

275 280 285

Glu Leu Lys Ala Met Cys Glu Thr Ala Ser Ser Pro Asp Asp Lys His

290 295 300

Gln Met Cys Leu Arg Ala Glu Glu Lys Val Arg Leu Val Ser Ala Leu

305 310 315 320

Gln Arg Leu Ile Met Leu Glu Cys Gly Gly Ala Ala Leu Glu Ser Gly

325 330 335

Val Thr Arg Trp Ala Val Glu Asn Gly Ile Ser Val Met Val Gly Ile

340 345 350

Gly Met Thr Glu Thr Val Gly Thr Leu Phe Ala Glu Arg Ala Gln Asp

355 360 365

Ala Arg Ser Asn Gly Tyr Ser Ala Gln Asp Ala Leu Ile Ser Asp Gly

370 375 380

Ile Met Ser Leu Val Gly Ser Asp Asn Glu Glu Ala Thr Leu Glu Gly

385 390 395 400

Glu Leu Val Val Lys Ser Lys Leu Ile Pro His Gly Tyr Ile Lys Tyr

405 410 415

Arg Asp Ser Ser Phe Ser Val Asp Ser Asp Gly Trp Val Thr Phe Lys

420 425 430

Thr Gly Asp Lys Tyr Gln Arg Thr Pro Asp Gly Arg Phe Lys Trp Leu

435 440 445

Gly Arg Lys Thr Asp Phe Ile Gln Met Thr Ser Ser Glu Thr Leu Asp

450 455 460

Pro Arg Pro Ile Glu Glu Ala Leu Cys Ala Asn Pro Ser Ile Ala Asn

465 470 475 480

Ala Cys Val Ile Gly Asp Arg Phe Leu Arg Glu Pro Ala Thr Ser Val

485 490 495

Cys Ala Ile Val Glu Ile Gly Pro Glu Val Asp Met Pro Ser Ser Lys

500 505 510

Ile Asp Arg Glu Ile Ala Asn Thr Leu Thr Pro Ile Asn Arg Gly Leu

515 520 525

Pro Pro Ala Leu Arg Ile Ser Trp Ser Arg Val Leu Ile Ile Arg Pro

530 535 540

Pro Gln Lys Ile Pro Val Thr Arg Lys Gly Asp Val Phe Arg Lys Lys

545 550 555 560

Ile Glu Asp Met Phe Gly Ser Phe Leu Gly Val Gly Val Ser Thr Glu

565 570 575

Val Glu Val Asp His Glu Thr Lys Glu Asp Asp Thr Lys His Val Val

580 585 590

Arg Gln Val Val Ser Asn Leu Leu Gly Val His Asp Leu Glu Leu Leu

595 600 605

Ser Ala Leu Ser Phe Ala Glu Leu Gly Met Thr Ser Phe Met Ala Val

610 615 620

Ser Ile Val Asn Thr Leu Asn Lys Arg Ile Asp Gly Leu Thr Leu Pro

625 630 635 640

Pro Asn Ala Cys Tyr Ile His Ile Asp Leu Asp Ser Leu Val Asp Ala

645 650 655

Ile Ser Leu Glu His Gly His Glu Ser Ile Pro Ala Glu Leu Pro Ser

660 665 670

Asn Pro Phe Pro Val Ile Glu Ser His Gln His Asn Asp Lys Asp Ile

675 680 685

Val Ile Val Gly Lys Ala Phe Arg Leu Pro Gly Ser Leu Asn Asn Thr

690 695 700

Ala Ser Leu Trp Glu Ala Leu Leu Ser Lys Asn Ser Ser Val Ile Ser

705 710 715 720

Asp Ile Pro Ser Asp Arg Trp Asp His Ala Ser Phe Tyr Pro His Asp

725 730 735

Ile Cys Phe Thr Lys Ala Gly Leu Val Asp Val Ala His Tyr Asp Tyr

740 745 750

Arg Phe Phe Gly Leu Thr Ala Thr Glu Ala Leu Tyr Val Ser Pro Thr

755 760 765

Met Arg Leu Ala Leu Glu Val Ser Phe Glu Ala Leu Glu Asn Ala Asn

770 775 780

Ile Pro Leu Ser Lys Leu Lys Gly Thr Gln Thr Ala Val Tyr Val Ala

785 790 795 800

Thr Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Glu Gln Gly Tyr

805 810 815

Asp Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser Thr Ala

820 825 830

Ser Gly Arg Ile Ser Tyr Leu Leu Asp Ile His Gly Pro Ser Val Thr

835 840 845

Val Asp Thr Ala Cys Ser Gly Gly Ile Val Cys Met Asp Gln Ala Ile

850 855 860

Thr Phe Leu Gln Ser Gly Gly Ala Asp Thr Ala Ile Val Cys Ser Ser

865 870 875 880

Asn Thr His Cys Trp Pro Gly Ser Phe Met Phe Leu Thr Ala Gln Gly

885 890 895

Met Val Ser Gln Asn Gly Arg Cys Ala Thr Phe Thr Thr Asp Ala Asp

900 905 910

Gly Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys Thr Arg

915 920 925

Ser Ala Ala Ile Arg Asp Asn Asp Asn Ile Leu Ala Val Ile Arg Ser

930 935 940

Thr Asp Val Ser His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn

945 950 955 960

Val Lys Ala Gln Thr Asn Leu His Arg Ser Leu Leu Arg Lys Ala Gly

965 970 975

Leu Phe Pro Asp Gln Ile Asn Phe Ile Glu Ala His Gly Thr Gly Thr

980 985 990

Ser Leu Gly Asp Leu Ser Glu Ile Gln Gly Ile Asn Asn Ala Tyr Thr

995 1000 1005

Ser Thr Arg Pro Arg Leu Ala Gly Pro Leu Ile Ile Ser Ala Ser

1010 1015 1020

Lys Thr Val Leu Gly His Ser Glu Pro Thr Ala Gly Met Ala Gly

1025 1030 1035

Ile Leu Thr Ala Leu Leu Ala Leu Glu Lys Glu Thr Val Pro Gly

1040 1045 1050

Leu Asn His Leu Thr Glu His Asn Leu Asn Pro Ser Leu Asp Cys

1055 1060 1065

Ser Val Val Pro Leu Leu Ile Pro His Glu Ser Ile His Ile Gly

1070 1075 1080

Gly Ala Lys Pro His Arg Ala Ala Val Leu Ser Tyr Gly Phe Ala

1085 1090 1095

Gly Thr Leu Ala Gly Ala Ile Leu Glu Gly Pro Pro Ser Asp Val

1100 1105 1110

Pro Arg Pro Ser Ser Asn Asn Ile Gln Glu His Pro Met Ile Phe

1115 1120 1125

Val Val Ser Gly Lys Thr Val Pro Ala Leu Glu Ala Tyr Leu Gly

1130 1135 1140

Arg Tyr Leu Ala Phe Leu Arg Val Ala Lys Thr His Asp Phe His

1145 1150 1155

Asp Ile Cys Tyr Thr Thr Cys Val Gly Arg Glu His Tyr Lys Tyr

1160 1165 1170

Arg Phe Ser Cys Val Ala Arg Asn Met Ala Asp Leu Ile Ser Gln

1175 1180 1185

Ile Glu His Arg Leu Thr Ala Leu Ser Thr Ser Lys Gln Lys Pro

1190 1195 1200

Arg Gly Ser Leu Gly Phe Ile Phe Ser Gly Gln Gly Thr Tyr Phe

1205 1210 1215

Pro Gly Met Ala Ala Ala Leu Ala Glu Gln Tyr Ser Gly Phe Arg

1220 1225 1230

Val Leu Val Ser Lys Phe Gly Gln Ala Ala Gln Glu Arg Ser Gly

1235 1240 1245

Tyr Pro Ile Asp Arg Leu Leu Leu Glu Val Ser Asp Thr Leu Pro

1250 1255 1260

Glu Thr Asn Ser Glu Val Asp Gln Ile Cys Ile Phe Val Tyr Gln

1265 1270 1275

Tyr Ser Val Leu Gln Trp Leu Gln Ser Leu Gly Ile Gln Pro Lys

1280 1285 1290

Ala Val Leu Gly His Ser Leu Gly Glu Ile Thr Ala Ala Val Ala

1295 1300 1305

Ala Gly Ala Leu Ser Phe Glu Ser Ala Leu Asp Leu Val Val Thr

1310 1315 1320

Arg Ala Arg Leu Leu Arg Pro Arg Ala Lys Asp Ser Ala Gly Met

1325 1330 1335

Ala Ala Val Ala Ala Ser Lys Glu Glu Val Lys Gly Leu Ile Glu

1340 1345 1350

Thr Leu Gln Leu Ala Asp Ser Leu Ser Val Ala Val His Asn Gly

1355 1360 1365

Pro Arg Ser Val Val Val Ser Gly Ala Ser Ala Glu Ile Asp Ala

1370 1375 1380

Leu Val Val Ala Ala Lys Glu Arg Gly Leu Lys Ala Ser Arg Leu

1385 1390 1395

Lys Val Asp Gln Gly Phe His Ser Pro Tyr Val Asp Ser Ala Val

1400 1405 1410

Pro Gly Leu Leu Asp Trp Ser Asn Lys His Arg Ser Thr Phe Leu

1415 1420 1425

Pro Leu Asn Ile Pro Leu Tyr Ser Thr Leu Thr Gly Glu Leu Ile

1430 1435 1440

Pro Lys Gly Arg Arg Phe Val Ser Asp His Trp Val Asn His Ala

1445 1450 1455

Arg Lys Pro Val Gln Phe Ala Ala Ala Ala Ala Ala Val Asp Glu

1460 1465 1470

Asp Arg Ser Ile Gly Val Leu Val Asp Val Gly Pro Gln Pro Val

1475 1480 1485

Ala Trp Thr Leu Leu Gln Ala Asn Asn Leu Leu Asn Thr Ser Ala

1490 1495 1500

Val Ala Leu Phe Ala Lys Ala Gly Lys Asp Gln Glu Met Ala Leu

1505 1510 1515

Leu Thr Ala Leu Ser Tyr Leu Val Gln Glu His Asn Leu Ser Pro

1520 1525 1530

Asn Phe His Glu Leu Tyr Ser Gln Arg His Gly Ala Leu Lys Lys

1535 1540 1545

Thr Asp Val Pro Thr Tyr Pro Phe Arg Arg Val His Arg Tyr Pro

1550 1555 1560

Thr Phe Ile Pro Ser Arg Asn Gln Ser Pro Ala Ala Ala Thr Val

1565 1570 1575

Ala Met Pro Pro Pro Arg Phe Ser Val Gln Lys Asn Ala Asp Val

1580 1585 1590

Ala Ser Gln Ser Lys Glu Ser Asp Cys Arg Ala Gly Leu Ile Ser

1595 1600 1605

Cys Leu Arg Ala Ile Leu Glu Leu Thr Pro Glu Glu Glu Phe Asp

1610 1615 1620

Leu Ser Glu Thr Leu Asn Ala Arg Gly Met Asp Ser Ile Met Phe

1625 1630 1635

Ala Gln Leu Arg Lys Arg Val Gly Glu Glu Phe Asp Leu Asp Ile

1640 1645 1650

Pro Met Ile Tyr Leu Ser Asp Val Phe Thr Met Glu Gln Met Val

1655 1660 1665

Asp Tyr Leu Val Lys Gln Ser Gly Ser Arg Pro Ala Leu Lys His

1670 1675 1680

Ala Glu Ile Pro Val Asn Gln Pro Leu Asp Glu Asp Leu Arg Thr

1685 1690 1695

Arg Leu Val Ser Cys Leu Arg Asn Val Leu Glu Ile Thr Pro Asp

1700 1705 1710

Glu Glu Leu Asp Leu Ser Glu Thr Leu Asn Ala Arg Gly Val Asp

1715 1720 1725

Ser Ile Met Phe Ala Gln Leu Arg Lys Arg Val Gly Glu Gly Phe

1730 1735 1740

Gly Val Glu Ile Pro Met Ile Tyr Leu Ser Asp Val Phe Thr Met

1745 1750 1755

Glu Asp Met Ile Asn Phe Leu Val Ser Glu Arg Ser

1760 1765 1770

<210> 52

<211> 5322

<212> DNA

<213> Armillaria fuscipes

<400> 52

atgaccatgg aggtcgacag ccactattct ctgctcgatg tctttctcag cattgcacat 60

gattctgaca agtccaaacg caatgtcttg gaatgcggcc tggaggcctg gacatactcg 120

gatttagaca tcatctcgtc ggctctggca caggatctca aagctacctt gggatgtttt 180

cccaaagttg cagtcgtcag cgagaaccat ccctacgtgt ttgctctcat gctggccgtc 240

tggaagctcg aagggatctt catccccata gacgtccacg ttacagctga ccttttaaag 300

ggcatgctac gcattgttgc tcctacttgt ctagtgatcc ctgagagcga tgtttccaac 360

cggcgtgttg cctctgcgat tggtatacgt gttctcccat ttgatgcgaa ttcgtcaacc 420

atgacggcac ttcgacaaaa gtacgaatca ttcactcaga aagcctcgcc atctgagtgc 480

acacttgccc acgccgatcg tacatgcctc tatcttttta catcctctgc atcctctacc 540

gccaacttga aatgtgtgcc tttgacccat actcttatcc tcaataactg ccgtaccaag 600

ctcgcatggt ggcagcgctt tcgtccagaa agcgaaatgg atgggatgcg tgttctaggg 660

tgggcacctt ggtcgcatat ccttgcctac atgcaagata tcggcacagc gacgctcctg 720

aacgccggtt gctatgtctt tgcgtccatt ccatccacat accctacaca actagcagca 780

aatggcttac aaggccccac tatgaatatc atcaacgcac ttcttgaacg acaaattgcc 840

gcatttgctt gcgtgccgtt cattttgagc gaactcaaag ctatgtgcga gatgacttcc 900

tgtacaggaa accaaatgtc cctaagagcc gaggagaaag ttcgcctggt cagggtgctg 960

caggggcttg taatgctcga gtgtggaggc gcggcacttg agtcagatgt tacgcgttgg 1020

gtcgttgaga atgacatacc agtcatggtt ggcattggga tgacggaaac agtcggtacg 1080

ctgttcgcag agcgcgccca agacgtccgt tccagcggat attccgccca agacgctctc 1140

attgctgatg gcattatgtc actggtcggt tctgacaacg aggaagccac tttcgaaggg 1200

gaactagtcg tgaagagcaa gctcatccca cagggataca tagggtaccg tgattcatcg 1260

ttctcggtgg actcagatgg ctgggtaacg ttcaaaactg gagataaata tcagcgcact 1320

ccagatggac gattaaaatg gcttgggaga aagaccgact ttatccagat gacaagcagc 1380

gaaacactgg atcccaggcc cattgagcaa actctttgtg cgaatccata cgtcgcaaaa 1440

gcatgcgtca ttggtgacag attcctgaga gatcccgcga ccagcgtatg tgccataatt 1500

gagatcaggc cggaagtgga catgccttcg tccaagatcg acagagaaat tgcgaatgcc 1560

cttgctccaa tcaatcgcga cctccctcct gctcttcgca tatcatggtc tcgcgtactc 1620

atgatcagac cccctcagaa aatcccagta acgaggaaag gcgatgtgtt ccgtaagaag 1680

attgaagata tgtttgggtc tttcctcggt gtcggtgttt ctactgaggt cggagtcgac 1740

catgaaactg aaaaagatga tacggaacac attgtgagac aggttgttac taatcttctt 1800

ggagtccatg atcccgagct attgtctgct ttatctttcg ctgagcttgg aatgacttca 1860

tttatggctg tcagcatcgt caactctcta aacaagtaca tcgatggcct cacccttcca 1920

cctaatgcat gttacatcca tattgatctt gattctcttg tggaggccat ttcacgtgaa 1980

cggggtcatg gaagtaacgc cacagagttg ccttctcaac ccgtccctgt tgagttacat 2040

caacctaacg ataaggacgt tgtgatagtt ggcaaggcat tccgtttacc tggctcactc 2100

gacagcactg catctctatg ggaagctttg ttatcaaaga acaattcagt cgtcagtgaa 2160

atcccatccg atcgctggga tcacgcaagc ttttaccccc acgacatttg cttcacgaaa 2220

gcaggcctcg tcgatgttgc ccattacgat tacagattct ttggcctcac ggctacagag 2280

gcattgtatg tatctccgac gatgcgtctc gccttggaag tgtcatttga agccctggag 2340

aatgcgaata ttccgttatc caatctgaag ggaacacaaa ccgctgtcta tgtcgccacc 2400

aaagacgatg gtttcgagac acttttaaat gccgagcagg gctatgatgc ctacacacga 2460

ttctatggca cgggccgcgc tccaagcacc gcaagtggcc gtataagcta tctacttgat 2520

attcatgggc catctgtcac cgttgataca gcatgcagcg gaggcattgt gtgtatggat 2580

caagccatca ctttcttgca gtccggaggg gcagatacag ctattgtctg ttcgagcaat 2640

acgcattgtt ggcctggatc atttatgttc ctgacggcgc aaggcatggt ttctccaaat 2700

ggaagatgcg ctacatttag taccgatgca gacggatatg tgccttcaga gggcgcagta 2760

gctttcgttc tcaagacgcg tagcgcagca atacgcgata atgacaatat cctcgccgta 2820

atcaaatcaa cagatgtgtc tcataacggc cgttctcaag ggctggttgc acctaacgtg 2880

aaggcgcaga caaacctgca tcaatcgttg ttacgaaaag ctgggctgtt tcctgatcaa 2940

atcaacttta tcgaagccca tggaacaggt acatctctag gagacctctc agaaatccag 3000

ggcatcaata acgcctacac ctcaacacga cctcgtctag acggtcccct tatcatcagc 3060

gcatcgaaaa cagtgatagg acacagcgaa ccaactgcag ggatggcggg catcctcaca 3120

gccttgcttg ctcttgagaa agaaacagtt cctggtctca atcacttaac ggagcacagc 3180

cttaaccctt cgcttgattg cagcatagtc ccgctcctga ttcctcacga gtctattcac 3240

attggtgggg taaagccaca tcgagctgcg gttctgtcat acggcttcgc gggtacactg 3300

gcgggtgcta tcttagaggg accgccttca gatgcaccaa ggccgtcgtc aaataatgtg 3360

caagatcacc ctatgatttt tgccctcagt gggaaaagcg cgtccgcact ggaagcatac 3420

ctaaggcggt atttggcatt cctgcggatt gcagatccac acgacttcca taacatctgc 3480

tacacttcct gtgtcgggag ggagcactac aaatatcggt tctcctgtgt tgcccgaaac 3540

atggcagacc ttatatctca aattgaacat cgactgacaa ctgtttccat tccgaaaccg 3600

aaacctcgtg gctcaatagg attcacgttc tcaggacaag gcacttattt ccctggcatg 3660

gccgcagcac tcactgaaca atattctgga ttccggacgc tcgtctctaa gcttgggcag 3720

gctgcgcaag agcggtcggg tcatccgatt gacaggctgt tacttgaagt ttccggtaca 3780

tcaccagaaa caaacagtga ggtcgagcaa atttgcacat ttatctacca atatgccgtt 3840

ctgcaatggt tgcagagcct aggcgttcaa ccgaaagcag tcctcggtca cagcctggga 3900

gaaattactg ccgcagtcgc agctggtgcc ctgtcgttcg aatccgcgtt ggaccttgtg 3960

gtgacccgtg ctcgtcttct ccgtcccgaa acaaaagatt ctgctgggat ggtcgcggta 4020

gcaacgtcca aggatgaagt tgaaggactt atagaaacac tccaagttgc ggacgcgcta 4080

agcgttgccg ttcacaacgg ttcacggagt gttgtggttt caggcacatc agcggaagtt 4140

gatgccctgg tcgtcgcagc taaagaacgg ggcttaaagg cttcccgctt aagagtcgac 4200

caaggtttcc acagcccttg cgttgattct gccgttcctg gtttactcga ctggtcaaat 4260

gagcatcgtt ccaccttcct tcctttgaat atgcctttat actcgacttt gaccggcgag 4320

gtcattccca agggacggaa attcgtctgg gatcactggg taaaccatgc tcgaaaacct 4380

gttcagtttg caccggcagc aaaagcggtg gacgaagacc gatccatcgg tgtgctcgtt 4440

gatgtaggac ctcaacctgt cgcttggacc cttttgcaag caaacaacct ttccaacacc 4500

tctacggttg cgctattcgc gaaagccgga aaggatcagg agatggcact gctcactgct 4560

ttgagctacc tcttccaaga gcacaacctt tctcccaagt ttcacgacct ttatactggg 4620

tataatggtg ctctgaagaa gacggacatt cccacgtacc cattccaacg tgtccatcgc 4680

tatcccacct tcataccatc acgaaatcag agtcctgctg tcgcgaaagc agtcgtgcag 4740

ccgccccgct tctctatcca aaggaatcga gaagccacat tacagtcgaa ggaaccagat 4800

caccgagctt gtttagtcac ttgccttaga gccatcctcg aattaacatc agaggaagaa 4860

cttgacctct ctgagaccct caacgctcgt ggcgtggact cgatcatgtt ttcacagcta 4920

cggaagcggg ttggagaaga attcaatctc gagataccca tgatctattt atcagacgta 4980

ttcacgatgg agcagatgat tgactacctc gtcgaacagt ccggatccaa tcccgcatca 5040

aagcaagtag gaactccggt taaccgacta tcaggcgaag aagatcttcg gacggggctc 5100

atctcatgcc tgagggacgt gctagaaatc actcctgatg atgaacttga tcacccaaaa 5160

gacctatctg aaactttaaa tgctcgtggt gttgattcga taatgttcgc tcagctacga 5220

aaacgcgtcg gggaaggatt tggtgtggaa attccgatga tatatctgtc tgatgtgttt 5280

accatggaag acatgattaa tttcctcgtt tctgagcgct ca 5322

<210> 53

<211> 1774

<212>PRT

<213> Armillaria fuscipes

<400> 53

Met Thr Met Glu Val Asp Ser His Tyr Ser Leu Leu Asp Val Phe Leu

1 5 10 15

Ser Ile Ala His Asp Ser Asp Lys Ser Lys Arg Asn Val Leu Glu Cys

20 25 30

Gly Leu Glu Ala Trp Thr Tyr Ser Asp Leu Asp Ile Ile Ser Ser Ala

35 40 45

Leu Ala Gln Asp Leu Lys Ala Thr Leu Gly Cys Phe Pro Lys Val Ala

50 55 60

Val Val Ser Glu Asn His Pro Tyr Val Phe Ala Leu Met Leu Ala Val

65 70 75 80

Trp Lys Leu Glu Gly Ile Phe Ile Pro Ile Asp Val His Val Thr Ala

85 90 95

Asp Leu Leu Lys Gly Met Leu Arg Ile Val Ala Pro Thr Cys Leu Val

100 105 110

Ile Pro Glu Ser Asp Val Ser Asn Arg Arg Val Ala Ser Ala Ile Gly

115 120 125

Ile Arg Val Leu Pro Phe Asp Ala Asn Ser Ser Thr Met Thr Ala Leu

130 135 140

Arg Gln Lys Tyr Glu Ser Phe Thr Gln Lys Ala Ser Pro Ser Glu Cys

145 150 155 160

Thr Leu Ala His Ala Asp Arg Thr Cys Leu Tyr Leu Phe Thr Ser Ser

165 170 175

Ala Ser Ser Thr Ala Asn Leu Lys Cys Val Pro Leu Thr His Thr Leu

180 185 190

Ile Leu Asn Asn Cys Arg Thr Lys Leu Ala Trp Trp Gln Arg Phe Arg

195 200 205

Pro Glu Ser Glu Met Asp Gly Met Arg Val Leu Gly Trp Ala Pro Trp

210 215 220

Ser His Ile Leu Ala Tyr Met Gln Asp Ile Gly Thr Ala Thr Leu Leu

225 230 235 240

Asn Ala Gly Cys Tyr Val Phe Ala Ser Ile Pro Ser Thr Tyr Pro Thr

245 250 255

Gln Leu Ala Ala Asn Gly Leu Gln Gly Pro Thr Met Asn Ile Ile Asn

260 265 270

Ala Leu Leu Glu Arg Gln Ile Ala Ala Phe Ala Cys Val Pro Phe Ile

275 280 285

Leu Ser Glu Leu Lys Ala Met Cys Glu Met Thr Ser Cys Thr Gly Asn

290 295 300

Gln Met Ser Leu Arg Ala Glu Glu Lys Val Arg Leu Val Arg Val Leu

305 310 315 320

Gln Gly Leu Val Met Leu Glu Cys Gly Gly Ala Ala Leu Glu Ser Asp

325 330 335

Val Thr Arg Trp Val Val Glu Asn Asp Ile Pro Val Met Val Gly Ile

340 345 350

Gly Met Thr Glu Thr Val Gly Thr Leu Phe Ala Glu Arg Ala Gln Asp

355 360 365

Val Arg Ser Ser Gly Tyr Ser Ala Gln Asp Ala Leu Ile Ala Asp Gly

370 375 380

Ile Met Ser Leu Val Gly Ser Asp Asn Glu Glu Ala Thr Phe Glu Gly

385 390 395 400

Glu Leu Val Val Lys Ser Lys Leu Ile Pro Gln Gly Tyr Ile Gly Tyr

405 410 415

Arg Asp Ser Ser Phe Ser Val Asp Ser Asp Gly Trp Val Thr Phe Lys

420 425 430

Thr Gly Asp Lys Tyr Gln Arg Thr Pro Asp Gly Arg Leu Lys Trp Leu

435 440 445

Gly Arg Lys Thr Asp Phe Ile Gln Met Thr Ser Ser Glu Thr Leu Asp

450 455 460

Pro Arg Pro Ile Glu Gln Thr Leu Cys Ala Asn Pro Tyr Val Ala Lys

465 470 475 480

Ala Cys Val Ile Gly Asp Arg Phe Leu Arg Asp Pro Ala Thr Ser Val

485 490 495

Cys Ala Ile Ile Glu Ile Arg Pro Glu Val Asp Met Pro Ser Ser Lys

500 505 510

Ile Asp Arg Glu Ile Ala Asn Ala Leu Ala Pro Ile Asn Arg Asp Leu

515 520 525

Pro Pro Ala Leu Arg Ile Ser Trp Ser Arg Val Leu Met Ile Arg Pro

530 535 540

Pro Gln Lys Ile Pro Val Thr Arg Lys Gly Asp Val Phe Arg Lys Lys

545 550 555 560

Ile Glu Asp Met Phe Gly Ser Phe Leu Gly Val Gly Val Ser Thr Glu

565 570 575

Val Gly Val Asp His Glu Thr Glu Lys Asp Asp Thr Glu His Ile Val

580 585 590

Arg Gln Val Val Thr Asn Leu Leu Gly Val His Asp Pro Glu Leu Leu

595 600 605

Ser Ala Leu Ser Phe Ala Glu Leu Gly Met Thr Ser Phe Met Ala Val

610 615 620

Ser Ile Val Asn Ser Leu Asn Lys Tyr Ile Asp Gly Leu Thr Leu Pro

625 630 635 640

Pro Asn Ala Cys Tyr Ile His Ile Asp Leu Asp Ser Leu Val Glu Ala

645 650 655

Ile Ser Arg Glu Arg Gly His Gly Ser Asn Ala Thr Glu Leu Pro Ser

660 665 670

Gln Pro Val Pro Val Glu Leu His Gln Pro Asn Asp Lys Asp Val Val

675 680 685

Ile Val Gly Lys Ala Phe Arg Leu Pro Gly Ser Leu Asp Ser Thr Ala

690 695 700

Ser Leu Trp Glu Ala Leu Leu Ser Lys Asn Asn Ser Val Val Ser Glu

705 710 715 720

Ile Pro Ser Asp Arg Trp Asp His Ala Ser Phe Tyr Pro His Asp Ile

725 730 735

Cys Phe Thr Lys Ala Gly Leu Val Asp Val Ala His Tyr Asp Tyr Arg

740 745 750

Phe Phe Gly Leu Thr Ala Thr Glu Ala Leu Tyr Val Ser Pro Thr Met

755 760 765

Arg Leu Ala Leu Glu Val Ser Phe Glu Ala Leu Glu Asn Ala Asn Ile

770 775 780

Pro Leu Ser Asn Leu Lys Gly Thr Gln Thr Ala Val Tyr Val Ala Thr

785 790 795 800

Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Glu Gln Gly Tyr Asp

805 810 815

Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser Thr Ala Ser

820 825 830

Gly Arg Ile Ser Tyr Leu Leu Asp Ile His Gly Pro Ser Val Thr Val

835 840 845

Asp Thr Ala Cys Ser Gly Gly Ile Val Cys Met Asp Gln Ala Ile Thr

850 855 860

Phe Leu Gln Ser Gly Gly Ala Asp Thr Ala Ile Val Cys Ser Ser Asn

865 870 875 880

Thr His Cys Trp Pro Gly Ser Phe Met Phe Leu Thr Ala Gln Gly Met

885 890 895

Val Ser Pro Asn Gly Arg Cys Ala Thr Phe Ser Thr Asp Ala Asp Gly

900 905 910

Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Val Leu Lys Thr Arg Ser

915 920 925

Ala Ala Ile Arg Asp Asn Asp Asn Ile Leu Ala Val Ile Lys Ser Thr

930 935 940

Asp Val Ser His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn Val

945 950 955 960

Lys Ala Gln Thr Asn Leu His Gln Ser Leu Leu Arg Lys Ala Gly Leu

965 970 975

Phe Pro Asp Gln Ile Asn Phe Ile Glu Ala His Gly Thr Gly Thr Ser

980 985 990

Leu Gly Asp Leu Ser Glu Ile Gln Gly Ile Asn Asn Ala Tyr Thr Ser

995 1000 1005

Thr Arg Pro Arg Leu Asp Gly Pro Leu Ile Ile Ser Ala Ser Lys

1010 1015 1020

Thr Val Ile Gly His Ser Glu Pro Thr Ala Gly Met Ala Gly Ile

1025 1030 1035

Leu Thr Ala Leu Leu Ala Leu Glu Lys Glu Thr Val Pro Gly Leu

1040 1045 1050

Asn His Leu Thr Glu His Ser Leu Asn Pro Ser Leu Asp Cys Ser

1055 1060 1065

Ile Val Pro Leu Leu Ile Pro His Glu Ser Ile His Ile Gly Gly

1070 1075 1080

Val Lys Pro His Arg Ala Ala Val Leu Ser Tyr Gly Phe Ala Gly

1085 1090 1095

Thr Leu Ala Gly Ala Ile Leu Glu Gly Pro Pro Ser Asp Ala Pro

1100 1105 1110

Arg Pro Ser Ser Asn Asn Val Gln Asp His Pro Met Ile Phe Ala

1115 1120 1125

Leu Ser Gly Lys Ser Ala Ser Ala Leu Glu Ala Tyr Leu Arg Arg

1130 1135 1140

Tyr Leu Ala Phe Leu Arg Ile Ala Asp Pro His Asp Phe His Asn

1145 1150 1155

Ile Cys Tyr Thr Ser Cys Val Gly Arg Glu His Tyr Lys Tyr Arg

1160 1165 1170

Phe Ser Cys Val Ala Arg Asn Met Ala Asp Leu Ile Ser Gln Ile

1175 1180 1185

Glu His Arg Leu Thr Thr Val Ser Ile Pro Lys Pro Lys Pro Arg

1190 1195 1200

Gly Ser Ile Gly Phe Thr Phe Ser Gly Gln Gly Thr Tyr Phe Pro

1205 1210 1215

Gly Met Ala Ala Ala Leu Thr Glu Gln Tyr Ser Gly Phe Arg Thr

1220 1225 1230

Leu Val Ser Lys Leu Gly Gln Ala Ala Gln Glu Arg Ser Gly His

1235 1240 1245

Pro Ile Asp Arg Leu Leu Leu Glu Val Ser Gly Thr Ser Pro Glu

1250 1255 1260

Thr Asn Ser Glu Val Glu Gln Ile Cys Thr Phe Ile Tyr Gln Tyr

1265 1270 1275

Ala Val Leu Gln Trp Leu Gln Ser Leu Gly Val Gln Pro Lys Ala

1280 1285 1290

Val Leu Gly His Ser Leu Gly Glu Ile Thr Ala Ala Val Ala Ala

1295 1300 1305

Gly Ala Leu Ser Phe Glu Ser Ala Leu Asp Leu Val Val Thr Arg

1310 1315 1320

Ala Arg Leu Leu Arg Pro Glu Thr Lys Asp Ser Ala Gly Met Val

1325 1330 1335

Ala Val Ala Thr Ser Lys Asp Glu Val Glu Gly Leu Ile Glu Thr

1340 1345 1350

Leu Gln Val Ala Asp Ala Leu Ser Val Ala Val His Asn Gly Ser

1355 1360 1365

Arg Ser Val Val Val Ser Gly Thr Ser Ala Glu Val Asp Ala Leu

1370 1375 1380

Val Val Ala Ala Lys Glu Arg Gly Leu Lys Ala Ser Arg Leu Arg

1385 1390 1395

Val Asp Gln Gly Phe His Ser Pro Cys Val Asp Ser Ala Val Pro

1400 1405 1410

Gly Leu Leu Asp Trp Ser Asn Glu His Arg Ser Thr Phe Leu Pro

1415 1420 1425

Leu Asn Met Pro Leu Tyr Ser Thr Leu Thr Gly Glu Val Ile Pro

1430 1435 1440

Lys Gly Arg Lys Phe Val Trp Asp His Trp Val Asn His Ala Arg

1445 1450 1455

Lys Pro Val Gln Phe Ala Pro Ala Ala Lys Ala Val Asp Glu Asp

1460 1465 1470

Arg Ser Ile Gly Val Leu Val Asp Val Gly Pro Gln Pro Val Ala

1475 1480 1485

Trp Thr Leu Leu Gln Ala Asn Asn Leu Ser Asn Thr Ser Thr Val

1490 1495 1500

Ala Leu Phe Ala Lys Ala Gly Lys Asp Gln Glu Met Ala Leu Leu

1505 1510 1515

Thr Ala Leu Ser Tyr Leu Phe Gln Glu His Asn Leu Ser Pro Lys

1520 1525 1530

Phe His Asp Leu Tyr Thr Gly Tyr Asn Gly Ala Leu Lys Lys Thr

1535 1540 1545

Asp Ile Pro Thr Tyr Pro Phe Gln Arg Val His Arg Tyr Pro Thr

1550 1555 1560

Phe Ile Pro Ser Arg Asn Gln Ser Pro Ala Val Ala Lys Ala Val

1565 1570 1575

Val Gln Pro Pro Arg Phe Ser Ile Gln Arg Asn Arg Glu Ala Thr

1580 1585 1590

Leu Gln Ser Lys Glu Pro Asp His Arg Ala Cys Leu Val Thr Cys

1595 1600 1605

Leu Arg Ala Ile Leu Glu Leu Thr Ser Glu Glu Glu Leu Asp Leu

1610 1615 1620

Ser Glu Thr Leu Asn Ala Arg Gly Val Asp Ser Ile Met Phe Ser

1625 1630 1635

Gln Leu Arg Lys Arg Val Gly Glu Glu Phe Asn Leu Glu Ile Pro

1640 1645 1650

Met Ile Tyr Leu Ser Asp Val Phe Thr Met Glu Gln Met Ile Asp

1655 1660 1665

Tyr Leu Val Glu Gln Ser Gly Ser Asn Pro Ala Ser Lys Gln Val

1670 1675 1680

Gly Thr Pro Val Asn Arg Leu Ser Gly Glu Glu Asp Leu Arg Thr

1685 1690 1695

Gly Leu Ile Ser Cys Leu Arg Asp Val Leu Glu Ile Thr Pro Asp

1700 1705 1710

Asp Glu Leu Asp His Pro Lys Asp Leu Ser Glu Thr Leu Asn Ala

1715 1720 1725

Arg Gly Val Asp Ser Ile Met Phe Ala Gln Leu Arg Lys Arg Val

1730 1735 1740

Gly Glu Gly Phe Gly Val Glu Ile Pro Met Ile Tyr Leu Ser Asp

1745 1750 1755

Val Phe Thr Met Glu Asp Met Ile Asn Phe Leu Val Ser Glu Arg

1760 1765 1770

Ser

<210> 54

<211> 5304

<212> DNA

<213> Armillaria mellea

<400> 54

atggaggcca acggtcacta ctctcttctc gatgtctttc tcagcattgc acacgattct 60

gacaagtcca aacgtaatgt cttggaatgc ggtcaggata cctggacata ctcagatttg 120

gacattatct cgtcggccct ggcacaggat ctccaagcta cgctgggttg ttttcccaaa 180

gttgcagtcg tcagcgagaa ccatccctac atttttgctc tcatgctggc cgtttggaag 240

ctcgaaggga tattcatccc tgtcgacgtc catgttacag ctgaccttct aaagggcatg 300

ttacatatcg tcgctcccac ttgtctggtg atccctgaga ccgatatttc caaccagcgt 360

attgcttccg cgattggtat acatgttctc cccttcagtg cgaatgcgtc gaccatgact 420

gcacttcgac agaaatacga cctatgcatt cagaaagcct cgctatctga gcgcgcactt 480

cctcacgttg atcgcgcttg cctctatctc tttacatcct ctgcgtcctc tactgccaac 540

ttgaaatgcg tacctttgac ccatagtctt atcctcagta actgccgttc caaactcgca 600

tggtggcggc gcgttcgtcc agaggaagaa atggatggga tacgtgttct agggtgggca 660

ccttggtccc atatccttgc ctacatgcaa gatattggca cagcgacgct cctgaacgcc 720

ggttgctacg tctttgcgtc aattccgtcc acatatccta cacaactggt ggtgaatggc 780

ttacaaggcc ccaccatgaa tatcatcgac tcacttctta aacggcgaat cgtcgcattt 840

gcttgtgtcc cgttcatttt gggcgaacta aaagctatgt gcgagacggc ttcgggtcca 900

gatgtcaagc atcatatggg cttgagagct gaggagaagg ttcgccttgt tagggcactg 960

cagcagctta tgatgctcga gtgcggaggc gctgcgctcg agtcagatgt cacgcgttgg 1020

gttgtcgaaa atggcatatc ggtcatggtt ggcatcggga tgacggagac agttggtacg 1080

ctgtttgcag agcgcgcgca agacgcccgt tccaacggct actctgcgca gaacgccctc 1140

attactgatg gcattatgtc actggtcggg cctgacaacg aggaagtcac ctttgaaggg 1200

gaactagtcg tgaagagcaa gctcatccca cacgaataca tcaattaccg tgattcgtcg 1260

ttctcggtgg actcggatgg ctgggtaacg tttaaaacgg gagataaata tcagcgcaca 1320

ccagatggac gattcaaatg gcttggaaga aagaccgact ttattcagat gacaagcagt 1380

gaaacactgg atcccagacc cattgagaaa accctctgtg cgaatccaag tattgtaaat 1440

gcatgcgtca ttggtgacag attcctgagg gagcctgcaa ctagcgtatg cgccattgtc 1500

gagatcaggc cggaagtgga catcccttcg tccaagatcg acagggaaat tgcgaatgcc 1560

cttgctccaa tcaatcgcga cctccctcct gctcttcgca tatcatggtc tcgcgtactc 1620

ataattcgac ctcctcagaa aataccagtt acaaggaaag gcgatgtgtt ccgtaagaag 1680

attgaagatg tatttgggtc tttccttggt gtcggttcta ccgagatcaa agtcgaccat 1740

gaatctaaag aagatgatac ggaacacatt gtgagacagg ttgtcagcaa tcttcttgga 1800

gtccatgatc ctgagctatt gtctgcttta tctttcgctg agctgggaat gacctcgttt 1860

atagctgtta gtattgtcaa cgctctaaac aagcgcatca gcggcctcac ccttccacct 1920

aatgcgtgct acctccatat tgatcttgat tctcttgtga acgccatttc acgtgaacat 1980

ggtcatggaa ctaaccccgc agagttgcct tctaaacttt tccccgtcac cgagtcctat 2040

caacgtaatg ataaggacgt tgtgataatc ggcaaagcat tccgtttacc aggctcactc 2100

gacaatactg catctctctg ggaagctttg ttatcgaaga acaattcagt cgtcagtgac 2160

atcccatccg atcgctggga tcacgcaagc ttttaccccc acgatatatg cttcacgaaa 2220

gcaggcctcg tcgatgttgc acattacgat tacagattct ttgggctcac agcgacagag 2280

gcattgtacg tatctccgac gatgcgcctc gccttggaag tgtcatttga agcccttgag 2340

aacgcgaata ttccgctatc caagctgaag gggacacaaa cccctgtcta tgtcgccact 2400

aaagacgatg gctttgagac actcttaaat gctgagcaag gctacgatgc ttacacgcga 2460

ttctatggta caggtcgcgc tccgagcacc gcaagtggtc gtataagcta tctactcgat 2520

attcatgggc catctgttac cgttgataca gcatgcagcg gaggcattgt atgtatggat 2580

caagccatca ctttcttgca atccggaagg gccgataccg ctattgtctg ttcgagcaat 2640

acgcactgtt ggccgggatc atttatgttc ctgacggcgc agggcatggt ttctccacat 2700

ggaagatgcg ctacatttac taccgatgca gacggttatg taccttcgga gggtgctgtg 2760

gctttcattc tcaagacgcg cagtgctgca atacgcgata atgacaatat actcgccgtg 2820

atcaaatcaa cagatgtgtc ccataacggc cgttctcaag ggctagttgc accaaacgta 2880

aaggcgcaga caagcctaca ccgatcgttg ctacgaaaag ctggactatt tcctgatcaa 2940

atcaatttta tcgaagccca tggaacaggt acatctctag gagacctctc ggaaatccag 3000

ggcatcaaca acgcctacac ctcaacacga cctcgtttgg acggtccct tatcattagc 3060

gcgtcgaaaa cagtgttggg acacagcgaa ccaattgcgg ggatggccgg catcctcaca 3120

gccttgcttt ccctcgagaa agaaacagtt tttggtttaa atcacttaac agagcacaac 3180

cttaaccctt cgcttgattg cagcctagtt cctctcctga ttcctcacga gtctattcac 3240

attggtggtg aaaaaccaca tcgagctgcg gttctgtcat acggtttcgc gggtacgctg 3300

gccggtgcca tcttagaggg accaccttca gatgtaccaa ggccgtcgtc aagcgatatg 3360

caagaacacc ctatggtttt cgtcctcagt gggaaaagcg tgcctgcact ggaaacgtac 3420

ctaagacggt atttggcatt tttgcgcgcc gcaaaaacaa acgacttcca tagcatctgc 3480

tacaccactt gcgtcgggag ggagcattac aaataccggt tctcctgcgt tgcccaaaat 3540

atggcagacc ttgtgtctca aattgaacat cgactgacaa ctctttccaa ttcgaaacag 3600

aaacctcgtg gctcgccagg gtttatgttc tcaggacaag gcacttattt ccctggtatg 3660

gctgcagcgc ttgctgaaca atatttgggg tttcgagtgc tagtctctag gtttgggaag 3720

gctgctcaag agcggtcggg ttatccgatc gataggctgt tgcttgaagt ttctgataca 3780

tcatcagaaa caaacagcga ggctgaccaa atttgcattt ttgtctacca atattccgtt 3840

ctgcaatggc tgcagagtct aggcattcaa ccgaaagcag tcctcggtca cagcctggga 3900

gaaattaccg ccgcagtcgc agctggtgcc ctttcattcg aatctgcgtt ggaccttgta 3960

gtcacccgtg ctcgtcttct ccgtcctgaa acaaaagatt cagcaggaat ggccgcagta 4020

gcagcatcca aggaagaagt tgaagaactt atagaaaacc tccaacttgc gcatgcgtta 4080

agtgttgcgg ttcacaacgg tccacggagt gttgttgtgt caggcgcatc gaccgaaatt 4140

gatgccctgg tcgtcgcagc taaagaacgg ggcttgaagg cttcccgctt aagagttgac 4200

caaggcttcc atagccctta cgttgattct gccgttccgg gtttactcga ctggtcaagt 4260

gaacattgtt cgaccttcct tcctttaaat attcctttat actcgacttt gaccggcgaa 4320

gttattccaa agggacggag gttcgcctgg gatcactggg taaaccatgc tcgaaaacct 4380

gttcagtttg cggcggcggc agcagcggtg gacgaagacc gatccatcgg cgtgctcctt 4440

gatgttggac cccaacccgt tgcgtggacc atccttcaag caagcagcct tttcaacacc 4500

tctgcagttg cgctatttgc gaaggctgga aaggatcagg agatggcgct gcttactact 4560

ttgagctacc tcttccaaga gcacaatctt tgtcccaact ttcacgagct ttactctcag 4620

cgtcatggtg ctcttaagaa gacggacatt cccacctacc cattccaacg tgtccaccgc 4680

tatccaacct tcataccatc acgaaatcaa agtcctgctg tcgcgaaggt agtagtccca 4740

tcgcgctttc ctgtccaaag gaaaggggaa gcaatatcac aatcgaacga atcagattac 4800

cgagctggtt tgatcacttg cctcagaacc atcctcgaat taacatcaga agaagagttt 4860

gacctttctg agaccctcaa cgctcgtggt gtggattcga tcatgttttc acagctacgg 4920

aagcgggttg gggaagaatt cgatctcgat atacccatga tctatttatc agacgtgttc 4980

acgatggaac agatgatcga ctacctcgtc gaacagtccg gatccagacc cgcgccaaag 5040

cacgtagaaa ctccggttaa cgaaccttta ggcaaagatc tccggacggg gctcgtttca 5100

tgcctgagga atgtactaga aatcaccccc gatgaagaac tcgacctatc tgaaactttg 5160

aacgctcgtg gtgtcgactc gatcatgttc gctcagctac gaaaacgcgt cggggaagga 5220

tttggcgtgg aaattccgat gatatatctg tctgacgtgt ttactatgga agacatgatc 5280

aatttcctcg tctctgagca cgcg 5304

<210> 55

<211> 1768

<212>PRT

<213> Armillaria mellea

<400> 55

Met Glu Ala Asn Gly His Tyr Ser Leu Leu Asp Val Phe Leu Ser Ile

1 5 10 15

Ala His Asp Ser Asp Lys Ser Lys Arg Asn Val Leu Glu Cys Gly Gln

20 25 30

Asp Thr Trp Thr Tyr Ser Asp Leu Asp Ile Ile Ser Ser Ala Leu Ala

35 40 45

Gln Asp Leu Gln Ala Thr Leu Gly Cys Phe Pro Lys Val Ala Val Val

50 55 60

Ser Glu Asn His Pro Tyr Ile Phe Ala Leu Met Leu Ala Val Trp Lys

65 70 75 80

Leu Glu Gly Ile Phe Ile Pro Val Asp Val His Val Thr Ala Asp Leu

85 90 95

Leu Lys Gly Met Leu His Ile Val Ala Pro Thr Cys Leu Val Ile Pro

100 105 110

Glu Thr Asp Ile Ser Asn Gln Arg Ile Ala Ser Ala Ile Gly Ile His

115 120 125

Val Leu Pro Phe Ser Ala Asn Ala Ser Thr Met Thr Ala Leu Arg Gln

130 135 140

Lys Tyr Asp Leu Cys Ile Gln Lys Ala Ser Leu Ser Glu Arg Ala Leu

145 150 155 160

Pro His Val Asp Arg Ala Cys Leu Tyr Leu Phe Thr Ser Ser Ala Ser

165 170 175

Ser Thr Ala Asn Leu Lys Cys Val Pro Leu Thr His Ser Leu Ile Leu

180 185 190

Ser Asn Cys Arg Ser Lys Leu Ala Trp Trp Arg Arg Val Arg Pro Glu

195 200 205

Glu Glu Met Asp Gly Ile Arg Val Leu Gly Trp Ala Pro Trp Ser His

210 215 220

Ile Leu Ala Tyr Met Gln Asp Ile Gly Thr Ala Thr Leu Leu Asn Ala

225 230 235 240

Gly Cys Tyr Val Phe Ala Ser Ile Pro Ser Thr Tyr Pro Thr Gln Leu

245 250 255

Val Val Asn Gly Leu Gln Gly Pro Thr Met Asn Ile Ile Asp Ser Leu

260 265 270

Leu Lys Arg Arg Ile Val Ala Phe Ala Cys Val Pro Phe Ile Leu Gly

275 280 285

Glu Leu Lys Ala Met Cys Glu Thr Ala Ser Gly Pro Asp Val Lys His

290 295 300

His Met Gly Leu Arg Ala Glu Glu Lys Val Arg Leu Val Arg Ala Leu

305 310 315 320

Gln Gln Leu Met Met Leu Glu Cys Gly Gly Ala Ala Leu Glu Ser Asp

325 330 335

Val Thr Arg Trp Val Val Glu Asn Gly Ile Ser Val Met Val Gly Ile

340 345 350

Gly Met Thr Glu Thr Val Gly Thr Leu Phe Ala Glu Arg Ala Gln Asp

355 360 365

Ala Arg Ser Asn Gly Tyr Ser Ala Gln Asn Ala Leu Ile Thr Asp Gly

370 375 380

Ile Met Ser Leu Val Gly Pro Asp Asn Glu Glu Val Thr Phe Glu Gly

385 390 395 400

Glu Leu Val Val Lys Ser Lys Leu Ile Pro His Glu Tyr Ile Asn Tyr

405 410 415

Arg Asp Ser Ser Phe Ser Val Asp Ser Asp Gly Trp Val Thr Phe Lys

420 425 430

Thr Gly Asp Lys Tyr Gln Arg Thr Pro Asp Gly Arg Phe Lys Trp Leu

435 440 445

Gly Arg Lys Thr Asp Phe Ile Gln Met Thr Ser Ser Glu Thr Leu Asp

450 455 460

Pro Arg Pro Ile Glu Lys Thr Leu Cys Ala Asn Pro Ser Ile Val Asn

465 470 475 480

Ala Cys Val Ile Gly Asp Arg Phe Leu Arg Glu Pro Ala Thr Ser Val

485 490 495

Cys Ala Ile Val Glu Ile Arg Pro Glu Val Asp Ile Pro Ser Ser Lys

500 505 510

Ile Asp Arg Glu Ile Ala Asn Ala Leu Ala Pro Ile Asn Arg Asp Leu

515 520 525

Pro Pro Ala Leu Arg Ile Ser Trp Ser Arg Val Leu Ile Ile Arg Pro

530 535 540

Pro Gln Lys Ile Pro Val Thr Arg Lys Gly Asp Val Phe Arg Lys Lys

545 550 555 560

Ile Glu Asp Val Phe Gly Ser Phe Leu Gly Val Gly Ser Thr Glu Ile

565 570 575

Lys Val Asp His Glu Ser Lys Glu Asp Asp Thr Glu His Ile Val Arg

580 585 590

Gln Val Val Ser Asn Leu Leu Gly Val His Asp Pro Glu Leu Leu Ser

595 600 605

Ala Leu Ser Phe Ala Glu Leu Gly Met Thr Ser Phe Ile Ala Val Ser

610 615 620

Ile Val Asn Ala Leu Asn Lys Arg Ile Ser Gly Leu Thr Leu Pro Pro

625 630 635 640

Asn Ala Cys Tyr Leu His Ile Asp Leu Asp Ser Leu Val Asn Ala Ile

645 650 655

Ser Arg Glu His Gly His Gly Thr Asn Pro Ala Glu Leu Pro Ser Lys

660 665 670

Leu Phe Pro Val Thr Glu Ser Tyr Gln Arg Asn Asp Lys Asp Val Val

675 680 685

Ile Ile Gly Lys Ala Phe Arg Leu Pro Gly Ser Leu Asp Asn Thr Ala

690 695 700

Ser Leu Trp Glu Ala Leu Leu Ser Lys Asn Asn Ser Val Val Ser Asp

705 710 715 720

Ile Pro Ser Asp Arg Trp Asp His Ala Ser Phe Tyr Pro His Asp Ile

725 730 735

Cys Phe Thr Lys Ala Gly Leu Val Asp Val Ala His Tyr Asp Tyr Arg

740 745 750

Phe Phe Gly Leu Thr Ala Thr Glu Ala Leu Tyr Val Ser Pro Thr Met

755 760 765

Arg Leu Ala Leu Glu Val Ser Phe Glu Ala Leu Glu Asn Ala Asn Ile

770 775 780

Pro Leu Ser Lys Leu Lys Gly Thr Gln Thr Pro Val Tyr Val Ala Thr

785 790 795 800

Lys Asp Asp Gly Phe Glu Thr Leu Leu Asn Ala Glu Gln Gly Tyr Asp

805 810 815

Ala Tyr Thr Arg Phe Tyr Gly Thr Gly Arg Ala Pro Ser Thr Ala Ser

820 825 830

Gly Arg Ile Ser Tyr Leu Leu Asp Ile His Gly Pro Ser Val Thr Val

835 840 845

Asp Thr Ala Cys Ser Gly Gly Ile Val Cys Met Asp Gln Ala Ile Thr

850 855 860

Phe Leu Gln Ser Gly Arg Ala Asp Thr Ala Ile Val Cys Ser Ser Asn

865 870 875 880

Thr His Cys Trp Pro Gly Ser Phe Met Phe Leu Thr Ala Gln Gly Met

885 890 895

Val Ser Pro His Gly Arg Cys Ala Thr Phe Thr Thr Asp Ala Asp Gly

900 905 910

Tyr Val Pro Ser Glu Gly Ala Val Ala Phe Ile Leu Lys Thr Arg Ser

915 920 925

Ala Ala Ile Arg Asp Asn Asp Asn Ile Leu Ala Val Ile Lys Ser Thr

930 935 940

Asp Val Ser His Asn Gly Arg Ser Gln Gly Leu Val Ala Pro Asn Val

945 950 955 960

Lys Ala Gln Thr Ser Leu His Arg Ser Leu Leu Arg Lys Ala Gly Leu

965 970 975

Phe Pro Asp Gln Ile Asn Phe Ile Glu Ala His Gly Thr Gly Thr Ser

980 985 990

Leu Gly Asp Leu Ser Glu Ile Gln Gly Ile Asn Asn Ala Tyr Thr Ser

995 1000 1005

Thr Arg Pro Arg Leu Asp Gly Pro Leu Ile Ile Ser Ala Ser Lys

1010 1015 1020

Thr Val Leu Gly His Ser Glu Pro Ile Ala Gly Met Ala Gly Ile

1025 1030 1035

Leu Thr Ala Leu Leu Ser Leu Glu Lys Glu Thr Val Phe Gly Leu

1040 1045 1050

Asn His Leu Thr Glu His Asn Leu Asn Pro Ser Leu Asp Cys Ser

1055 1060 1065

Leu Val Pro Leu Leu Ile Pro His Glu Ser Ile His Ile Gly Gly

1070 1075 1080

Glu Lys Pro His Arg Ala Ala Val Leu Ser Tyr Gly Phe Ala Gly

1085 1090 1095

Thr Leu Ala Gly Ala Ile Leu Glu Gly Pro Pro Ser Asp Val Pro

1100 1105 1110

Arg Pro Ser Ser Ser Asp Met Gln Glu His Pro Met Val Phe Val

1115 1120 1125

Leu Ser Gly Lys Ser Val Pro Ala Leu Glu Thr Tyr Leu Arg Arg

1130 1135 1140

Tyr Leu Ala Phe Leu Arg Ala Ala Lys Thr Asn Asp Phe His Ser

1145 1150 1155

Ile Cys Tyr Thr Thr Cys Val Gly Arg Glu His Tyr Lys Tyr Arg

1160 1165 1170

Phe Ser Cys Val Ala Gln Asn Met Ala Asp Leu Val Ser Gln Ile

1175 1180 1185

Glu His Arg Leu Thr Thr Leu Ser Asn Ser Lys Gln Lys Pro Arg

1190 1195 1200

Gly Ser Pro Gly Phe Met Phe Ser Gly Gln Gly Thr Tyr Phe Pro

1205 1210 1215

Gly Met Ala Ala Ala Leu Ala Glu Gln Tyr Leu Gly Phe Arg Val

1220 1225 1230

Leu Val Ser Arg Phe Gly Lys Ala Ala Gln Glu Arg Ser Gly Tyr

1235 1240 1245

Pro Ile Asp Arg Leu Leu Leu Glu Val Ser Asp Thr Ser Ser Glu

1250 1255 1260

Thr Asn Ser Glu Ala Asp Gln Ile Cys Ile Phe Val Tyr Gln Tyr

1265 1270 1275

Ser Val Leu Gln Trp Leu Gln Ser Leu Gly Ile Gln Pro Lys Ala

1280 1285 1290

Val Leu Gly His Ser Leu Gly Glu Ile Thr Ala Ala Val Ala Ala

1295 1300 1305

Gly Ala Leu Ser Phe Glu Ser Ala Leu Asp Leu Val Val Thr Arg

1310 1315 1320

Ala Arg Leu Leu Arg Pro Glu Thr Lys Asp Ser Ala Gly Met Ala

1325 1330 1335

Ala Val Ala Ala Ser Lys Glu Glu Val Glu Glu Leu Ile Glu Asn

1340 1345 1350

Leu Gln Leu Ala His Ala Leu Ser Val Ala Val His Asn Gly Pro

1355 1360 1365

Arg Ser Val Val Val Ser Gly Ala Ser Thr Glu Ile Asp Ala Leu

1370 1375 1380

Val Val Ala Ala Lys Glu Arg Gly Leu Lys Ala Ser Arg Leu Arg

1385 1390 1395

Val Asp Gln Gly Phe His Ser Pro Tyr Val Asp Ser Ala Val Pro

1400 1405 1410

Gly Leu Leu Asp Trp Ser Ser Glu His Cys Ser Thr Phe Leu Pro

1415 1420 1425

Leu Asn Ile Pro Leu Tyr Ser Thr Leu Thr Gly Glu Val Ile Pro

1430 1435 1440

Lys Gly Arg Arg Phe Ala Trp Asp His Trp Val Asn His Ala Arg

1445 1450 1455

Lys Pro Val Gln Phe Ala Ala Ala Ala Ala Ala Val Asp Glu Asp

1460 1465 1470

Arg Ser Ile Gly Val Leu Leu Asp Val Gly Pro Gln Pro Val Ala

1475 1480 1485

Trp Thr Ile Leu Gln Ala Ser Ser Leu Phe Asn Thr Ser Ala Val

1490 1495 1500

Ala Leu Phe Ala Lys Ala Gly Lys Asp Gln Glu Met Ala Leu Leu

1505 1510 1515

Thr Thr Leu Ser Tyr Leu Phe Gln Glu His Asn Leu Cys Pro Asn

1520 1525 1530

Phe His Glu Leu Tyr Ser Gln Arg His Gly Ala Leu Lys Lys Thr

1535 1540 1545

Asp Ile Pro Thr Tyr Pro Phe Gln Arg Val His Arg Tyr Pro Thr

1550 1555 1560

Phe Ile Pro Ser Arg Asn Gln Ser Pro Ala Val Ala Lys Val Val

1565 1570 1575

Val Pro Ser Arg Phe Pro Val Gln Arg Lys Gly Glu Ala Ile Ser

1580 1585 1590

Gln Ser Asn Glu Ser Asp Tyr Arg Ala Gly Leu Ile Thr Cys Leu

1595 1600 1605

Arg Thr Ile Leu Glu Leu Thr Ser Glu Glu Glu Phe Asp Leu Ser

1610 1615 1620

Glu Thr Leu Asn Ala Arg Gly Val Asp Ser Ile Met Phe Ser Gln

1625 1630 1635

Leu Arg Lys Arg Val Gly Glu Glu Phe Asp Leu Asp Ile Pro Met

1640 1645 1650

Ile Tyr Leu Ser Asp Val Phe Thr Met Glu Gln Met Ile Asp Tyr

1655 1660 1665

Leu Val Glu Gln Ser Gly Ser Arg Pro Ala Pro Lys His Val Glu

1670 1675 1680

Thr Pro Val Asn Glu Pro Leu Gly Lys Asp Leu Arg Thr Gly Leu

1685 1690 1695

Val Ser Cys Leu Arg Asn Val Leu Glu Ile Thr Pro Asp Glu Glu

1700 1705 1710

Leu Asp Leu Ser Glu Thr Leu Asn Ala Arg Gly Val Asp Ser Ile

1715 1720 1725

Met Phe Ala Gln Leu Arg Lys Arg Val Gly Glu Gly Phe Gly Val

1730 1735 1740

Glu Ile Pro Met Ile Tyr Leu Ser Asp Val Phe Thr Met Glu Asp

1745 1750 1755

Met Ile Asn Phe Leu Val Ser Glu His Ala

1760 1765

<210> 56

<211> 40

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 1 hispidin synthase

<220>

<221> MISC_FEATURE

<223> X - any amino acid

<220>

<221> MISC_FEATURE

<222> (1)..(40)

<223> X - any amino acid

<400> 56

Val Ala Xaa Xaa Xaa Glu Asn His Pro Xaa Xaa Xaa Ala Leu Xaa Xaa

1 5 10 15

Ala Val Trp Lys Xaa Xaa Gly Xaa Phe Xaa Pro Xaa Asp Xaa His Xaa

20 25 30

Xaa Xaa Xaa Xaa Xaa Xaa Gly Met

35 40

<210> 57

<211> 36

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 2 hispidin synthases

<220>

<221> MISC_FEATURE

<222> (1)..(36)

<223> X - any amino acid

<400> 57

Leu Gly Trp Ala Pro Xaa Ser His Xaa Leu Xaa Xaa Met Gln Asp Ile

1 5 10 15

Gly Xaa Xaa Xaa Xaa Leu Xaa Ala Gly Cys Tyr Val Phe Xaa Xaa Xaa

20 25 30

Pro Xaa Xaa Tyr

35

<210> 58

<211> 33

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 3 hispidin synthases

<220>

<221> MISC_FEATURE

<222> (6)..(33)

<223> X - any amino acid

<400> 58

Glu Cys Gly Gly Ala Xaa Leu Xaa Xaa Xaa Xaa Xaa Xaa Trp Xaa Xaa

1 5 10 15

Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Gly Xaa Gly Met Thr Glu Thr Xaa

20 25 30

Gly

<210> 59

<211> 8

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 4 hispidin synthases

<400> 59

Phe Ala Glu Leu Gly Met Thr Ser

15

<210> 60

<211> 25

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 5 hispidin synthases

<220>

<221> MISC_FEATURE

<222> (2)..(24)

<223> X - any amino acid

<400> 60

Ile Xaa Xaa Xaa Arg Trp Asp His Xaa Ser Phe Tyr Pro Xaa Xaa Ile

1 5 10 15

Xaa Xaa Xaa Xaa Ala Gly Leu Xaa Asp

20 25

<210> 61

<211> 65

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 6 hispidin synthases

<220>

<221> MISC_FEATURE

<222> (2)..(64)

<223> X - any amino acid

<400> 61

Asp Xaa Xaa Phe Phe Gly Xaa Xaa Xaa Xaa Glu Ala Xaa Xaa Xaa Ser

1 5 10 15

Pro Thr Met Arg Xaa Ala Leu Glu Val Xaa Xaa Glu Ala Leu Glu Xaa

20 25 30

Ala Asn Ile Pro Xaa Xaa Xaa Xaa Lys Gly Xaa Xaa Xaa Xaa Val Xaa

35 40 45

Xaa Ala Xaa Xaa Asp Asp Gly Phe Glu Thr Leu Leu Xaa Ala Xaa Xaa

50 55 60

Gly

65

<210> 62

<211> 50

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 7 hispidin synthases

<220>

<221> MISC_FEATURE

<222> (5)..(47)

<223> X - any amino acid

<400> 62

Ala Asp Gly Tyr Xaa Pro Ser Glu Gly Ala Val Xaa Phe Xaa Leu Lys

1 5 10 15

Thr Xaa Xaa Ala Ala Xaa Arg Asp Xaa Xaa Xaa Ile Xaa Ala Xaa Ile

20 25 30

Xaa Xaa Xaa Xaa Xaa Xaa His Asn Gly Arg Ser Gln Gly Leu Xaa Ala

35 40 45

Pro Asn

50

<210> 63

<211> 9

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence of 8 hispidin synthases

<220>

<221> MISC_FEATURE

<222> (8)..(8)

<223> X - any amino acid

<400> 63

Gly His Ser Leu Gly Glu Ile Xaa Ala

15

<210> 64

<211> 867

<212> DNA

<213> Neonothopanus nambi

<400> 64

atggcgccaa tttcttcaac ttggtctcgt ctcattcgat ttgtggctat tgaaacgtcc 60

ctcgtgcata tcggtgaacc gatacgcc accatggacg tcggtctggc gagacgagaa 120

ggcaagacga tccaagcata cgagattatt ggatcaggct cggctctaga cctctcagcc 180

caagtatcga agaatgtgct gactgtaagg gaactcctga tgccgctttc aagagaggaa 240

attaaaactg tacgatgctt ggggttgaac taccctgttc atgccaccga agcgaacgtt 300

gctgttccaa aattcccgaa tttgttctac aaaccagtga cctcgctcat tggccccgat 360

ggactcatta ccatcccttc cgttgtccaa cccccgaagg agcatcagtc cgattatgaa 420

gcggaacttg tcattgtcat cgggaaagca gcaaagaatg tatcggagga tgaggctttg 480

gattatgtat tgggatacac tgccgcgaac gatatttcgt ttaggaaaca ccagctagca 540

gtctcacaat ggtctttctc gaaaggattt ggtagccttc tactcactat ccgtatggca 600

caaacccact cgggtaacat taatcgcttc tccagagacc agattttcaa tgtcaagaag 660

acaatttcct tcctgtcaca aggcactaca ctggaaccag gttctatcat tttgactggt 720

acacctgacg gagtgggctt tgtgcgcaat ccaccacttt accttaaaga tggagatgaa 780

gtaatgacct ggattggaag tggaatcgga acattagcca atacagtgca agaagagaag 840

acttgcttcg ctagtggcgg acacgag 867

<210> 65

<211> 289

<212>PRT

<213> Neonothopanus nambi

<400> 65

Met Ala Pro Ile Ser Ser Thr Trp Ser Arg Leu Ile Arg Phe Val Ala

1 5 10 15

Ile Glu Thr Ser Leu Val His Ile Gly Glu Pro Ile Asp Ala Thr Met

20 25 30

Asp Val Gly Leu Ala Arg Arg Glu Gly Lys Thr Ile Gln Ala Tyr Glu

35 40 45

Ile Ile Gly Ser Gly Ser Ala Leu Asp Leu Ser Ala Gln Val Ser Lys

50 55 60

Asn Val Leu Thr Val Arg Glu Leu Leu Met Pro Leu Ser Arg Glu Glu

65 70 75 80

Ile Lys Thr Val Arg Cys Leu Gly Leu Asn Tyr Pro Val His Ala Thr

85 90 95

Glu Ala Asn Val Ala Val Pro Lys Phe Pro Asn Leu Phe Tyr Lys Pro

100 105 110

Val Thr Ser Leu Ile Gly Pro Asp Gly Leu Ile Thr Ile Pro Ser Val

115 120 125

Val Gln Pro Pro Lys Glu His Gln Ser Asp Tyr Glu Ala Glu Leu Val

130 135 140

Ile Val Ile Gly Lys Ala Ala Lys Asn Val Ser Glu Asp Glu Ala Leu

145 150 155 160

Asp Tyr Val Leu Gly Tyr Thr Ala Ala Asn Asp Ile Ser Phe Arg Lys

165 170 175

His Gln Leu Ala Val Ser Gln Trp Ser Phe Ser Lys Gly Phe Gly Ser

180 185 190

Leu Leu Leu Thr Ile Arg Met Ala Gln Thr His Ser Gly Asn Ile Asn

195 200 205

Arg Phe Ser Arg Asp Gln Ile Phe Asn Val Lys Lys Thr Ile Ser Phe

210 215 220

Leu Ser Gln Gly Thr Thr Leu Glu Pro Gly Ser Ile Ile Leu Thr Gly

225 230 235 240

Thr Pro Asp Gly Val Gly Phe Val Arg Asn Pro Pro Leu Tyr Leu Lys

245 250 255

Asp Gly Asp Glu Val Met Thr Trp Ile Gly Ser Gly Ile Gly Thr Leu

260 265 270

Ala Asn Thr Val Gln Glu Glu Lys Thr Cys Phe Ala Ser Gly Gly His

275 280 285

Glu

<210> 66

<211> 963

<212> DNA

<213> Neonothopanus gardneri

<400> 66

atggcgccga ttttgacagt gagttccagt tcaatggtgc tgaatagctc aaaacaggct 60

atattgaata tttctgaata tcaccagccc tggactcgtc tcattcgatt tgtagccgtt 120

gagacgtcac tcgtgcatat tggtgaaccc atcgaggtga ctttggacgt cgggcaggca 180

aaatgtgaag gcaagacgat caaagcgtac gagattattg gatcagggtc ggccttggac 240

ctctcagctc aggtatcgaa gaatgtgcta accgtaaagg aactcttgat gccgctttcg 300

agagaagagg tcaagactgt gcggtgcttg ggactgaact attttactca tgcttccacc 360

gggcgcccgc tgtcgactag attaccgact ttgttctata agccagtgac ttcactcatc 420

ggacctgagg cgttcattaa tattccttcc gctgttcaac caccgaagga gcatcagtcc 480

gattatgaag cggagttggt aattattatt gggagagcgg cgaaggatgt acggaagag 540

gaggctttga attatgtttt gggatacacc gccgccaacg acatttcatt taggaaatat 600

caatttgcag tttcccagtg gtgtttttcg aaaggtttcg aatacaaa cccaatcggt 660

ccgtgcatcg tttccgcatc ttccattccg aacccgcaag acatccagat ccaatgcaaa 720

ctgaacggga atgtcgttca gaatggaaac accagtgatc aaatttttaa tatcaagaaa 780

acagtcgctt ttttgtcgca aggaacaaca cttgagtcag gatcaatcat cctgaccggt 840

acgcctggcg gagtgggatt tgtgcgcgat ccaccgcttt accttaaaga tggagatgaa 900

gtagtgactt ggattggaag tggggttgga agtttagtca acgtagtaaa agaagagaag 960

act 963

<210> 67

<211> 321

<212>PRT

<213> Neonothopanus gardneri

<400> 67

Met Ala Pro Ile Leu Thr Val Ser Ser Ser Ser Met Val Leu Asn Ser

1 5 10 15

Ser Lys Gln Ala Ile Leu Asn Ile Ser Glu Tyr His Gln Pro Trp Thr

20 25 30

Arg Leu Ile Arg Phe Val Ala Val Glu Thr Ser Leu Val His Ile Gly

35 40 45

Glu Pro Ile Glu Val Thr Leu Asp Val Gly Gln Ala Lys Cys Glu Gly

50 55 60

Lys Thr Ile Lys Ala Tyr Glu Ile Ile Gly Ser Gly Ser Ala Leu Asp

65 70 75 80

Leu Ser Ala Gln Val Ser Lys Asn Val Leu Thr Val Lys Glu Leu Leu

85 90 95

Met Pro Leu Ser Arg Glu Glu Val Lys Thr Val Arg Cys Leu Gly Leu

100 105 110

Asn Tyr Phe Thr His Ala Ser Thr Gly Arg Pro Leu Ser Thr Arg Leu

115 120 125

Pro Thr Leu Phe Tyr Lys Pro Val Thr Ser Leu Ile Gly Pro Glu Ala

130 135 140

Phe Ile Asn Ile Pro Ser Ala Val Gln Pro Pro Lys Glu His Gln Ser

145 150 155 160

Asp Tyr Glu Ala Glu Leu Val Ile Ile Ile Gly Arg Ala Ala Lys Asp

165 170 175

Val Pro Glu Glu Glu Ala Leu Asn Tyr Val Leu Gly Tyr Thr Ala Ala

180 185 190

Asn Asp Ile Ser Phe Arg Lys Tyr Gln Phe Ala Val Ser Gln Trp Cys

195 200 205

Phe Ser Lys Gly Phe Asp Asn Thr Asn Pro Ile Gly Pro Cys Ile Val

210 215 220

Ser Ala Ser Ser Ile Pro Asn Pro Gln Asp Ile Gln Ile Gln Cys Lys

225 230 235 240

Leu Asn Gly Asn Val Val Gln Asn Gly Asn Thr Ser Asp Gln Ile Phe

245 250 255

Asn Ile Lys Lys Thr Val Ala Phe Leu Ser Gln Gly Thr Thr Leu Glu

260 265 270

Ser Gly Ser Ile Ile Leu Thr Gly Thr Pro Gly Gly Val Gly Phe Val

275 280 285

Arg Asp Pro Pro Leu Tyr Leu Lys Asp Gly Asp Glu Val Val Thr Trp

290 295 300

Ile Gly Ser Gly Val Gly Ser Leu Val Asn Val Val Lys Glu Glu Lys

305 310 315 320

Thr

<210> 68

<211> 906

<212> DNA

<213> Armillaria gallica

<400> 68

atggcgccta ttatcactca atggtccaga cttatccgct ttgtcgccgt cgaaacctct 60

cgtgtacata ttggacagcc cgtagattct aacctggatg ttggtctagc ggcgtaccag 120

ggaatgctaa tcaaggctta cgaaatactt ggttctgctc tcgatccatc cgcccaaatg 180

actagcaaga tcctcactgt caaacaacta ttaaccccgc tgtctggcga agatgtcaag 240

gtcgttagat gcttgggtct caactaccca gcacatgcga atgaagggaa agtagaagca 300

cctaagtttc ctaacctttt ctataagcca gtgacgtcgc ttatcgggcc cctcgcacct 360

gtgatcattc ctgcagtcgc acaaccttct gcaatacatc aatccgatta tgaggctgaa 420

tttactgtcg tcataggcag ggcagctaag aatatcactg aagcggaagc tttagactat 480

gttctcggct acaccggcgg caatgacgtg tcttttcgtc agcatcaatt tgcggtctct 540

caatggtgtt tctccaaaag ttttgacaac acaaatccct ttgggccatg cttagttgcc 600

gcatcgtcta ttcccgaccc tcaaactgtg gccattaagt ttacactgaa tggtcaaact 660

gtccaagacg gaactactgc cgatcaactt ttcagcgtca aaaagaccat agcttatctt 720

tctcaaggca cgacgttaca gccgggctcc ataattatga ctggtactcc cagtggcgtt 780

ggattcgtcc gaaacccacc tctctacctc aaagatggag accatatgtt gacttggata 840

agcggtggaa tcggtacgct tgcaaacagc gtcgtggagg agaagactcc gactcctggt 900

ttagat 906

<210> 69

<211> 302

<212>PRT

<213> Armillaria gallica

<400> 69

Met Ala Pro Ile Ile Thr Gln Trp Ser Arg Leu Ile Arg Phe Val Ala

1 5 10 15

Val Glu Thr Ser Arg Val His Ile Gly Gln Pro Val Asp Ser Asn Leu

20 25 30

Asp Val Gly Leu Ala Ala Tyr Gln Gly Met Leu Ile Lys Ala Tyr Glu

35 40 45

Ile Leu Gly Ser Ala Leu Asp Pro Ser Ala Gln Met Thr Ser Lys Ile

50 55 60

Leu Thr Val Lys Gln Leu Leu Thr Pro Leu Ser Gly Glu Asp Val Lys

65 70 75 80

Val Val Arg Cys Leu Gly Leu Asn Tyr Pro Ala His Ala Asn Glu Gly

85 90 95

Lys Val Glu Ala Pro Lys Phe Pro Asn Leu Phe Tyr Lys Pro Val Thr

100 105 110

Ser Leu Ile Gly Pro Leu Ala Pro Val Ile Ile Pro Ala Val Ala Gln

115 120 125

Pro Ser Ala Ile His Gln Ser Asp Tyr Glu Ala Glu Phe Thr Val Val

130 135 140

Ile Gly Arg Ala Ala Lys Asn Ile Thr Glu Ala Glu Ala Leu Asp Tyr

145 150 155 160

Val Leu Gly Tyr Thr Gly Gly Asn Asp Val Ser Phe Arg Gln His Gln

165 170 175

Phe Ala Val Ser Gln Trp Cys Phe Ser Lys Ser Phe Asp Asn Thr Asn

180 185 190

Pro Phe Gly Pro Cys Leu Val Ala Ala Ser Ser Ile Pro Asp Pro Gln

195 200 205

Thr Val Ala Ile Lys Phe Thr Leu Asn Gly Gln Thr Val Gln Asp Gly

210 215 220

Thr Thr Ala Asp Gln Leu Phe Ser Val Lys Lys Thr Ile Ala Tyr Leu

225 230 235 240

Ser Gln Gly Thr Thr Leu Gln Pro Gly Ser Ile Ile Met Thr Gly Thr

245 250 255

Pro Ser Gly Val Gly Phe Val Arg Asn Pro Pro Leu Tyr Leu Lys Asp

260 265 270

Gly Asp His Met Leu Thr Trp Ile Ser Gly Gly Ile Gly Thr Leu Ala

275 280 285

Asn Ser Val Val Glu Glu Lys Thr Pro Thr Pro Gly Leu Asp

290 295 300

<210> 70

<211> 906

<212> DNA

<213> Armillaria ostoyae

<400> 70

atggcaccta ttattactca atggtccaga cttattcgct ttgttgccgt cgagacctct 60

cgtgtacata ttggacagcc catagattct accctggata ttggtctagc ggcgtaccag 120

ggaatgctaa tcaaggctta tgaaatactt ggttctgctc tcgatccatc cgcccaaatg 180

accagcaaga tcctcaccgt taaacagcta ttaactccgc tgtctggcga agatgtcaag 240

gtcgtccgat gcttgggtct taactatcca gctcatgcga atgaagggaa agtagaagcg 300

cctaagtttc ctaacctttt ctataagcca gtgacatcgc ttatcgggcc cctcgcacct 360

gtgatcattc ctgcagtcgc acagccttct gcaatacatc aatccgatta tgaggctgaa 420

tttactgtcg tcataggcag ggcagctaag aatgtcactg aagcggaagc tttagactat 480

gttctcgggt acaccggcgg caatgatgtg tcttttcgtc agcatcaatt tgcggtctct 540

caatggtgtt tctctaaaag ttttgacaat acaaatccct tcggtccatg cttagttgcc 600

gcatcgtcta ttcctgatcc tcaaactgtg gccattaagt ttacattgaa tggtgacacc 660

gtccaagacg gaactactgc tgatcaactt ttcagcgtca aaaagaccat cgcttatctt 720

tctcagggca cgacgttaca gccgggctcc ataattatga ctggcactcc cagtggtgtt 780

gggttcgtcc aaaacccacc tctctacctc aaagatgggg atcaaatgtt gacttggata 840

agcggtggaa tcggtacgct tgcaaacaac gtcgtagagg agaagactcc gactcctcgt 900

ttagac 906

<210> 71

<211> 302

<212>PRT

<213> Armillaria ostoyae

<400> 71

Met Ala Pro Ile Ile Thr Gln Trp Ser Arg Leu Ile Arg Phe Val Ala

1 5 10 15

Val Glu Thr Ser Arg Val His Ile Gly Gln Pro Ile Asp Ser Thr Leu

20 25 30

Asp Ile Gly Leu Ala Ala Tyr Gln Gly Met Leu Ile Lys Ala Tyr Glu

35 40 45

Ile Leu Gly Ser Ala Leu Asp Pro Ser Ala Gln Met Thr Ser Lys Ile

50 55 60

Leu Thr Val Lys Gln Leu Leu Thr Pro Leu Ser Gly Glu Asp Val Lys

65 70 75 80

Val Val Arg Cys Leu Gly Leu Asn Tyr Pro Ala His Ala Asn Glu Gly

85 90 95

Lys Val Glu Ala Pro Lys Phe Pro Asn Leu Phe Tyr Lys Pro Val Thr

100 105 110

Ser Leu Ile Gly Pro Leu Ala Pro Val Ile Ile Pro Ala Val Ala Gln

115 120 125

Pro Ser Ala Ile His Gln Ser Asp Tyr Glu Ala Glu Phe Thr Val Val

130 135 140

Ile Gly Arg Ala Ala Lys Asn Val Thr Glu Ala Glu Ala Leu Asp Tyr

145 150 155 160

Val Leu Gly Tyr Thr Gly Gly Asn Asp Val Ser Phe Arg Gln His Gln

165 170 175

Phe Ala Val Ser Gln Trp Cys Phe Ser Lys Ser Phe Asp Asn Thr Asn

180 185 190

Pro Phe Gly Pro Cys Leu Val Ala Ala Ser Ser Ile Pro Asp Pro Gln

195 200 205

Thr Val Ala Ile Lys Phe Thr Leu Asn Gly Asp Thr Val Gln Asp Gly

210 215 220

Thr Thr Ala Asp Gln Leu Phe Ser Val Lys Lys Thr Ile Ala Tyr Leu

225 230 235 240

Ser Gln Gly Thr Thr Leu Gln Pro Gly Ser Ile Ile Met Thr Gly Thr

245 250 255

Pro Ser Gly Val Gly Phe Val Gln Asn Pro Pro Leu Tyr Leu Lys Asp

260 265 270

Gly Asp Gln Met Leu Thr Trp Ile Ser Gly Gly Ile Gly Thr Leu Ala

275 280 285

Asn Asn Val Val Glu Glu Lys Thr Pro Thr Pro Arg Leu Asp

290 295 300

<210> 72

<211> 891

<212> DNA

<213> Armillaria mellea

<400> 72

atggcgccta ttatcactca gtggtccaga cttattcgct ttgttgccgt cgagacttct 60

cgtgtacatt ttggacagcc cgtagattct accctggatg ttggtctagc ggcgtaccag 120

ggtgtgttga tcaaggctta tgaaatactt ggttctgctc ttgatccatc cgcccaaatg 180

accagcaaga tcctcaccgt gaaacagcta ttaactccgc tgtctggcga ggatgtcaaa 240

gtcgtccgat gcttgggtct taactatcca gcacatgcga atgaagggaa agtagaagca 300

cccaagtttc ctaacctgtt ctataagcca gtgacatcgc ttatcgggcc cctcgcgcct 360

gtgatcattc ctgcagtcgc acagccttct gcaatacatc aatctgatta tgaggctgaa 420

tttactgttg tcctaggcag ggcagctaag aatgtcactg aagctgaagc cttggactat 480

gttctcggtt acaccggcgg caatgatgtg tcttttcggc agcatcaatt tgctgtctct 540

caatggtgtt tctccaaaag ttttgacaat acaaatccct tcggtccatg cttagttgcc 600

gcgtcgtcta ttcctgatcc tcaaactgtg gccattaagt ttacattgaa tggcaacacc 660

gtccaagatg gaactactgc tgatcaactt ttcagcgtca gaaagaccat cgcttatctt 720

tctcaaggca cgacgttaca gcctggctcc ataattatga ccggtactcc cagtggcgtt 780

gggttcgtcc gaaacccacc tctctacctc aaagatgggg atcaaatgtt gacttggatt 840

agcggtggaa tcggtacgct tgcaaacagc gtcatagagg agaagacccc a 891

<210> 73

<211> 297

<212>PRT

<213> Armillaria mellea

<400> 73

Met Ala Pro Ile Ile Thr Gln Trp Ser Arg Leu Ile Arg Phe Val Ala

1 5 10 15

Val Glu Thr Ser Arg Val His Phe Gly Gln Pro Val Asp Ser Thr Leu

20 25 30

Asp Val Gly Leu Ala Ala Tyr Gln Gly Val Leu Ile Lys Ala Tyr Glu

35 40 45

Ile Leu Gly Ser Ala Leu Asp Pro Ser Ala Gln Met Thr Ser Lys Ile

50 55 60

Leu Thr Val Lys Gln Leu Leu Thr Pro Leu Ser Gly Glu Asp Val Lys

65 70 75 80

Val Val Arg Cys Leu Gly Leu Asn Tyr Pro Ala His Ala Asn Glu Gly

85 90 95

Lys Val Glu Ala Pro Lys Phe Pro Asn Leu Phe Tyr Lys Pro Val Thr

100 105 110

Ser Leu Ile Gly Pro Leu Ala Pro Val Ile Ile Pro Ala Val Ala Gln

115 120 125

Pro Ser Ala Ile His Gln Ser Asp Tyr Glu Ala Glu Phe Thr Val Val

130 135 140

Leu Gly Arg Ala Ala Lys Asn Val Thr Glu Ala Glu Ala Leu Asp Tyr

145 150 155 160

Val Leu Gly Tyr Thr Gly Gly Asn Asp Val Ser Phe Arg Gln His Gln

165 170 175

Phe Ala Val Ser Gln Trp Cys Phe Ser Lys Ser Phe Asp Asn Thr Asn

180 185 190

Pro Phe Gly Pro Cys Leu Val Ala Ala Ser Ser Ile Pro Asp Pro Gln

195 200 205

Thr Val Ala Ile Lys Phe Thr Leu Asn Gly Asn Thr Val Gln Asp Gly

210 215 220

Thr Thr Ala Asp Gln Leu Phe Ser Val Arg Lys Thr Ile Ala Tyr Leu

225 230 235 240

Ser Gln Gly Thr Thr Leu Gln Pro Gly Ser Ile Ile Met Thr Gly Thr

245 250 255

Pro Ser Gly Val Gly Phe Val Arg Asn Pro Pro Leu Tyr Leu Lys Asp

260 265 270

Gly Asp Gln Met Leu Thr Trp Ile Ser Gly Gly Ile Gly Thr Leu Ala

275 280 285

Asn Ser Val Ile Glu Glu Lys Thr Pro

290 295

<210> 74

<211> 891

<212> DNA

<213> Armillaria fuscipes

<400> 74

atggcaccta ttatcactca atggtccaga cttattcgct ttgtctccat tgagacttct 60

ggtgttcata ttggacaacc cgtagatcct acccttgacg tcggtctagc ggcgtcccag 120

ggagtggcaa tcaaggttta tgaaataatt ggttctgcgc ttgatccatc cgcccaagtg 180

accagcaaaa tccttaccgt caaacagcta ttaactccgc tgtctggtga ggatgtcaag 240

gtcgtccggt gcttgggtct gaactatcca gcacatgcca atgaagggaa agtagaagca 300

cccaagtttc ctaacctttt ctataagcca gtgacatcgc ttattgggcc cctcgcgcct 360

gtgatcattc ctgcagtcgc acagccggcc gcaatacatc aatctgatta tgaggctgaa 420

tttactgttg tcattggcag ggcagccaag aatgtcacgg aagctgaagc cctggactat 480

gttcttggct acaccggcgg taatgatgtg tcttttcgga agcatcaatt tgcagtctct 540

caatggtgtt tctccaaaag ttttgacaat acaaatccct ttggaccatg cttagttgcc 600

gcttcgtcta tccctgatcc tcagaatgtg gccattaagt tcacgttgaa tggtaacacc 660

gttcaagatg gaactactgc tgaccaaatt ttcagcgtta gaaagactat cgcttatctt 720

tctcaaggca cgacgttaca gccaggctcc atcattatga ctggcactcc caatggcgtt 780

gggtttgtcc gagacccacc tctctacctc aaagacgggg atcaaatgct gacttggatt 840

agcggtggaa ttggtacgct tgcgaatggc gtcgtagaag agaagacccg g 891

<210> 75

<211> 297

<212>PRT

<213> Armillaria fuscipes

<400> 75

Met Ala Pro Ile Ile Thr Gln Trp Ser Arg Leu Ile Arg Phe Val Ser

1 5 10 15

Ile Glu Thr Ser Gly Val His Ile Gly Gln Pro Val Asp Pro Thr Leu

20 25 30

Asp Val Gly Leu Ala Ala Ser Gln Gly Val Ala Ile Lys Val Tyr Glu

35 40 45

Ile Ile Gly Ser Ala Leu Asp Pro Ser Ala Gln Val Thr Ser Lys Ile

50 55 60

Leu Thr Val Lys Gln Leu Leu Thr Pro Leu Ser Gly Glu Asp Val Lys

65 70 75 80

Val Val Arg Cys Leu Gly Leu Asn Tyr Pro Ala His Ala Asn Glu Gly

85 90 95

Lys Val Glu Ala Pro Lys Phe Pro Asn Leu Phe Tyr Lys Pro Val Thr

100 105 110

Ser Leu Ile Gly Pro Leu Ala Pro Val Ile Ile Pro Ala Val Ala Gln

115 120 125

Pro Ala Ala Ile His Gln Ser Asp Tyr Glu Ala Glu Phe Thr Val Val

130 135 140

Ile Gly Arg Ala Ala Lys Asn Val Thr Glu Ala Glu Ala Leu Asp Tyr

145 150 155 160

Val Leu Gly Tyr Thr Gly Gly Asn Asp Val Ser Phe Arg Lys His Gln

165 170 175

Phe Ala Val Ser Gln Trp Cys Phe Ser Lys Ser Phe Asp Asn Thr Asn

180 185 190

Pro Phe Gly Pro Cys Leu Val Ala Ala Ser Ser Ile Pro Asp Pro Gln

195 200 205

Asn Val Ala Ile Lys Phe Thr Leu Asn Gly Asn Thr Val Gln Asp Gly

210 215 220

Thr Thr Ala Asp Gln Ile Phe Ser Val Arg Lys Thr Ile Ala Tyr Leu

225 230 235 240

Ser Gln Gly Thr Thr Leu Gln Pro Gly Ser Ile Ile Met Thr Gly Thr

245 250 255

Pro Asn Gly Val Gly Phe Val Arg Asp Pro Pro Leu Tyr Leu Lys Asp

260 265 270

Gly Asp Gln Met Leu Thr Trp Ile Ser Gly Gly Ile Gly Thr Leu Ala

275 280 285

Asn Gly Val Val Glu Glu Lys Thr Arg

290 295

<210> 76

<211> 42

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 1

caffeoylpyruvate hydrolase

<220>

<221> MISC_FEATURE

<222> (2)..(39)

<223> X - any amino acid

<400> 76

Trp Xaa Arg Leu Ile Arg Phe Val Xaa Xaa Glu Thr Ser Xaa Val His

1 5 10 15

Xaa Gly Xaa Pro Xaa Xaa Xaa Xaa Xaa Asp Xaa Gly Xaa Ala Xaa Xaa

20 25 30

Xaa Gly Xaa Xaa Ile Xaa Xaa Tyr Glu Ile

35 40

<210> 77

<211> 137

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 2

caffeoylpyruvate hydrolase

<220>

<221> MISC_FEATURE

<222> (5)..(136)

<223> X - any amino acid

<400> 77

Ser Ala Leu Asp Xaa Ser Ala Gln Xaa Xaa Xaa Xaa Xaa Leu Thr Val

1 5 10 15

Xaa Xaa Leu Leu Xaa Pro Leu Ser Xaa Glu Xaa Xaa Lys Xaa Val Arg

20 25 30

Cys Leu Gly Leu Asn Tyr Xaa Xaa His Ala Xaa Xaa Xaa Xaa Xaa Xaa

35 40 45

Xaa Xaa Xaa Xaa Pro Xaa Leu Phe Tyr Lys Pro Val Thr Ser Leu Ile

50 55 60

Gly Pro Xaa Xaa Xaa Xaa Xaa Ile Pro Xaa Xaa Xaa Gln Pro Xaa Xaa

65 70 75 80

Xaa His Gln Ser Asp Tyr Glu Ala Glu Xaa Xaa Xaa Xaa Xaa Gly Xaa

85 90 95

Ala Ala Lys Xaa Xaa Xaa Glu Xaa Glu Ala Leu Xaa Tyr Val Leu Gly

100 105 110

Tyr Thr Xaa Xaa Asn Asp Xaa Ser Phe Arg Xaa Xaa Gln Xaa Ala Val

115 120 125

Ser Gln Trp Xaa Phe Ser Lys Xaa Phe

130 135

<210> 78

<211> 69

<212>PRT

<213> Artificial sequence

<220>

<223> Consensus amino acid sequence 3

caffeoylpyruvate hydrolase

<220>

<221> MISC_FEATURE

<222> (3)..(65)

<223> X - any amino acid

<400> 78

Asp Gln Xaa Phe Xaa Xaa Xaa Lys Thr Xaa Xaa Xaa Leu Ser Gln Gly

1 5 10 15

Thr Thr Leu Xaa Xaa Gly Ser Ile Ile Xaa Thr Gly Thr Pro Xaa Gly

20 25 30

Val Gly Phe Val Xaa Xaa Pro Pro Leu Tyr Leu Lys Asp Gly Asp Xaa

35 40 45

Xaa Xaa Thr Trp Ile Xaa Xaa Gly Xaa Gly Xaa Leu Xaa Asn Xaa Val

50 55 60

Xaa Glu Glu Lys Thr

65

<210> 79

<211> 801

<212> DNA

<213> Neonothopanus nambi

<400> 79

atgcgcatta acattagcct ctcgtctctc ttcgaacgtc tctccaaact tagcagtcgc 60

agcatagcga ttacatgtgg agttgttctc gcctccgcaa tcgcctttcc catcatccgc 120

agagactacc agactttcct agaagtggga ccctcgtacg ctccgcagaa ctttagagga 180

tacatcatcg tctgtgtcct ctcgctattc cgccaagagc agaaagggct cgccatctat 240

gatcgtcttc ccgagaaacg caggtggttg gccgaccttc cctttcgtga aggaaccaga 300

cccagcatta ccagccatat cattcagcga cagcgcactc aactggtcga tcaggagttt 360

gccaccaggg agctcataga caaggtcatc cctcgcgtgc aagcacgaca caccgacaaa 420

acgttcctca gcacatcaaa gttcgagttt catgcgaagg ccatatttct cttgccttct 480

atcccaatca acgaccctct gaatatccct agccacgaca ctgtccgccg aacgaagcgc 540

gagattgcac atatgcatga ttatcatgat tgcacacttc atcttgctct cgctgcgcag 600

gatggaaagg aggtgctgaa gaaaggttgg ggacaacgac atcctttggc tggtcctgga 660

gttcctggtc caccaacgga atggactttt ctttatgcgc ctcgcaacga agaagaggct 720

cgagtagtgg agatgatcgt tgaggcttcc atagggtata tgacgaacga tcctgcagga 780

aagattgtag aaaacgccaa g 801

<210> 80

<211> 267

<212>PRT

<213> Neonothopanus nambi

<400> 80

Met Arg Ile Asn Ile Ser Leu Ser Ser Leu Phe Glu Arg Leu Ser Lys

1 5 10 15

Leu Ser Ser Arg Ser Ile Ala Ile Thr Cys Gly Val Val Leu Ala Ser

20 25 30

Ala Ile Ala Phe Pro Ile Ile Arg Arg Asp Tyr Gln Thr Phe Leu Glu

35 40 45

Val Gly Pro Ser Tyr Ala Pro Gln Asn Phe Arg Gly Tyr Ile Ile Val

50 55 60

Cys Val Leu Ser Leu Phe Arg Gln Glu Gln Lys Gly Leu Ala Ile Tyr

65 70 75 80

Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Ala Asp Leu Pro Phe Arg

85 90 95

Glu Gly Thr Arg Pro Ser Ile Thr Ser His Ile Ile Gln Arg Gln Arg

100 105 110

Thr Gln Leu Val Asp Gln Glu Phe Ala Thr Arg Glu Leu Ile Asp Lys

115 120 125

Val Ile Pro Arg Val Gln Ala Arg His Thr Asp Lys Thr Phe Leu Ser

130 135 140

Thr Ser Lys Phe Glu Phe His Ala Lys Ala Ile Phe Leu Leu Pro Ser

145 150 155 160

Ile Pro Ile Asn Asp Pro Leu Asn Ile Pro Ser His Asp Thr Val Arg

165 170 175

Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp Cys Thr

180 185 190

Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys Glu Val Leu Lys Lys

195 200 205

Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Val Pro Gly Pro

210 215 220

Pro Thr Glu Trp Thr Phe Leu Tyr Ala Pro Arg Asn Glu Glu Glu Ala

225 230 235 240

Arg Val Val Glu Met Ile Val Glu Ala Ser Ile Gly Tyr Met Thr Asn

245 250 255

Asp Pro Ala Gly Lys Ile Val Glu Asn Ala Lys

260 265

<210> 81

<211> 783

<212> DNA

<213> Omphalotus olearius

<400> 81

atgctcccag ctttcatcta caaaccaagg ctagtgatca cttgtgtatt cgttctggcc 60

tccgcactcg catttccctt catacgcaaa gattaccaga ctttcctgga ggtgggaccc 120

tcgtacgccc cgcagaacct ccaaggatac atcatcgtct gtgtactctc tctgttccgg 180

caagaacaga aagacgtagc gatttatgat cgccttcctg agaaaaggag gtggttagga 240

gacctcccgt ttcgcgaggg gccaagaccg agtatcacta gccatatcat ccagcgacag 300

cgcacccaat tggctgacgc cgagttcgct accaaagagc tgataggcaa aatcatccct 360

cgcgtccaag cccgacacac caacacaaca ttcctcagca catctaaatt cgaattccac 420

gcccaggcca tcttcctttt gccctctatc ccaatcaacg accctcaaaa cattccaagc 480

cacgataccg ttcgtcgcac gaaacgcgag atcgcgcata tgcatgatta tcacgactgt 540

acgttgcatc tcgcacttgc tgctcaagat gggaaggagg ttttagagaa aggatggggt 600

cagcgacatc ctcttgctgg acctggtgtt cctggcccgc cgacggagtg gacgtttctt 660

tatgcaccgc gcagcgaaga ggaggttcgg gttgtggaga tgattgttga ggcatcagtt 720

gtgtatatga cgaatgatcc tgcggataaa atcgtagaag ctactgtgca gggtactgaa 780

gaa 783

<210> 82

<211> 261

<212>PRT

<213> Omphalotus olearius

<400> 82

Met Leu Pro Ala Phe Ile Tyr Lys Pro Arg Leu Val Ile Thr Cys Val

1 5 10 15

Phe Val Leu Ala Ser Ala Leu Ala Phe Pro Phe Ile Arg Lys Asp Tyr

20 25 30

Gln Thr Phe Leu Glu Val Gly Pro Ser Tyr Ala Pro Gln Asn Leu Gln

35 40 45

Gly Tyr Ile Ile Val Cys Val Leu Ser Leu Phe Arg Gln Glu Gln Lys

50 55 60

Asp Val Ala Ile Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Gly

65 70 75 80

Asp Leu Pro Phe Arg Glu Gly Pro Arg Pro Ser Ile Thr Ser His Ile

85 90 95

Ile Gln Arg Gln Arg Thr Gln Leu Ala Asp Ala Glu Phe Ala Thr Lys

100 105 110

Glu Leu Ile Gly Lys Ile Ile Pro Arg Val Gln Ala Arg His Thr Asn

115 120 125

Thr Thr Phe Leu Ser Thr Ser Lys Phe Glu Phe His Ala Gln Ala Ile

130 135 140

Phe Leu Leu Pro Ser Ile Pro Ile Asn Asp Pro Gln Asn Ile Pro Ser

145 150 155 160

His Asp Thr Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp

165 170 175

Tyr His Asp Cys Thr Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys

180 185 190

Glu Val Leu Glu Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro

195 200 205

Gly Val Pro Gly Pro Pro Thr Glu Trp Thr Phe Leu Tyr Ala Pro Arg

210 215 220

Ser Glu Glu Glu Val Arg Val Val Glu Met Ile Val Glu Ala Ser Val

225 230 235 240

Val Tyr Met Thr Asn Asp Pro Ala Asp Lys Ile Val Glu Ala Thr Val

245 250 255

Gln Gly Thr Glu Glu

260

<210> 83

<211> 798

<212> DNA

<213> Armillaria gallica

<400> 83

atgtccttca tcgacagcat gaaacttgac ctcgtcggac acctctttgg catcaggaat 60

cgcggcttag ccgccgcttg ttgtgctcta gcagtcgcct ctactatcgc cttcccttac 120

attcgtaggg actaccagac atttttatct ggcggtccct cttacgctcc ccagaatatc 180

agaggatatt tcatcgtctg cgttctggcc ttgttccgtc aggagcaaaa gggccttgcg 240

atatatgatc gccttcccga gaagcgcagg tggctgcctg acttgcctcc tcgcaatggc 300

ccgcggccga tcacgaccag ccatataatc caaagacagc gcaaccaggc gccggacccc 360

aagttcgccc tcgaggaact caaggccacg gttattccac gggtgcaggc tcgccatact 420

gacctcaccc atctcagcct atccaaattc gagttccatg ctgaagcaat tttcctgctc 480

ccctctgtac ccatcgatga tccaaaaaat gttccaagtc acgacacggt gcgcaggacg 540

aaaagggaga tcgcgcatat gcacgactac catgacttca cgctgcatct tgcactggcc 600

gcccaagacg ggaaggaagt cgtgtcgaag ggatgggggc agcgacaccc cctagcaggc 660

cctggcgttc ctggtccacc tacggagtgg acatttattt atgcgccacg taacgaagag 720

gaactggcag tggtggaaat gattatcgag gcatcaatag gctatatgac caatgaccct 780

gctggagtag ttatcgca 798

<210> 84

<211> 266

<212>PRT

<213> Armillaria gallica

<400> 84

Met Ser Phe Ile Asp Ser Met Lys Leu Asp Leu Val Gly His Leu Phe

1 5 10 15

Gly Ile Arg Asn Arg Gly Leu Ala Ala Ala Cys Cys Ala Leu Ala Val

20 25 30

Ala Ser Thr Ile Ala Phe Pro Tyr Ile Arg Arg Asp Tyr Gln Thr Phe

35 40 45

Leu Ser Gly Gly Pro Ser Tyr Ala Pro Gln Asn Ile Arg Gly Tyr Phe

50 55 60

Ile Val Cys Val Leu Ala Leu Phe Arg Gln Glu Gln Lys Gly Leu Ala

65 70 75 80

Ile Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Pro Asp Leu Pro

85 90 95

Pro Arg Asn Gly Pro Arg Pro Ile Thr Thr Ser His Ile Ile Gln Arg

100 105 110

Gln Arg Asn Gln Ala Pro Asp Pro Lys Phe Ala Leu Glu Glu Leu Lys

115 120 125

Ala Thr Val Ile Pro Arg Val Gln Ala Arg His Thr Asp Leu Thr His

130 135 140

Leu Ser Leu Ser Lys Phe Glu Phe His Ala Glu Ala Ile Phe Leu Leu

145 150 155 160

Pro Ser Val Pro Ile Asp Asp Pro Lys Asn Val Pro Ser His Asp Thr

165 170 175

Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp

180 185 190

Phe Thr Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys Glu Val Val

195 200 205

Ser Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Val Pro

210 215 220

Gly Pro Pro Thr Glu Trp Thr Phe Ile Tyr Ala Pro Arg Asn Glu Glu

225 230 235 240

Glu Leu Ala Val Val Glu Met Ile Ile Glu Ala Ser Ile Gly Tyr Met

245 250 255

Thr Asn Asp Pro Ala Gly Val Val Ile Ala

260 265

<210> 85

<211> 798

<212> DNA

<213> Armillaria ostoyae

<400> 85

atgtccttca tcgacagcat gaaacttgac ttcgtcggac acctctttgg catcaggaat 60

cgcggcttag ccaccgcttg ttgtgctgtg gcagtcgctt ctgccatcgc cttcccttac 120

attcgtaggg actaccagac attcttatct ggcggtccct cttacgctcc ccagaacatc 180

aaaggatatc tcatcgtctg cgtcctggcc ttgttccgtc aggagcaaaa gggccttgcg 240

atatatgacc gccttcccga gaagcgcagg tggctacctg acttgcctcc tcgcaatggc 300

ccgcggccca tcacgaccag ccatataatc caaagacagc gcaaccaggc gccagactcc 360

aagttcgccc tcgaggaact caaggctacg gtcattccac gggtgcaggc tcgccacact 420

gacctcaccc atctcagcct atccaagttc gagttccatg ctgaagcaat cttcctgctc 480

ccctctgtac ccatcgatga tccaaaaaat gttccaagtc atgacacggt gcgcaggacg 540

aagaggga tcgcgcatat gcacgactac catgacttta cgttgcatct tgcactggcc 600

gcccaagacg ggaaggaagt cgtggcgaag ggatgggggc agcgacaccc gctggcaggc 660

cctggcgttc ctggtccacc tacggagtgg acgtttattt atgcgccacg taacgaagag 720

gaactggcag tggtggaaat gattatcgag gcatcaatag gctatatgac caatgaccct 780

gctggaacag ttatcgta 798

<210> 86

<211> 266

<212>PRT

<213> Armillaria ostoyae

<400> 86

Met Ser Phe Ile Asp Ser Met Lys Leu Asp Phe Val Gly His Leu Phe

1 5 10 15

Gly Ile Arg Asn Arg Gly Leu Ala Thr Ala Cys Cys Ala Val Ala Val

20 25 30

Ala Ser Ala Ile Ala Phe Pro Tyr Ile Arg Arg Asp Tyr Gln Thr Phe

35 40 45

Leu Ser Gly Gly Pro Ser Tyr Ala Pro Gln Asn Ile Lys Gly Tyr Leu

50 55 60

Ile Val Cys Val Leu Ala Leu Phe Arg Gln Glu Gln Lys Gly Leu Ala

65 70 75 80

Ile Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Pro Asp Leu Pro

85 90 95

Pro Arg Asn Gly Pro Arg Pro Ile Thr Thr Ser His Ile Ile Gln Arg

100 105 110

Gln Arg Asn Gln Ala Pro Asp Ser Lys Phe Ala Leu Glu Glu Leu Lys

115 120 125

Ala Thr Val Ile Pro Arg Val Gln Ala Arg His Thr Asp Leu Thr His

130 135 140

Leu Ser Leu Ser Lys Phe Glu Phe His Ala Glu Ala Ile Phe Leu Leu

145 150 155 160

Pro Ser Val Pro Ile Asp Asp Pro Lys Asn Val Pro Ser His Asp Thr

165 170 175

Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp

180 185 190

Phe Thr Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys Glu Val Val

195 200 205

Ala Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Val Pro

210 215 220

Gly Pro Pro Thr Glu Trp Thr Phe Ile Tyr Ala Pro Arg Asn Glu Glu

225 230 235 240

Glu Leu Ala Val Val Glu Met Ile Ile Glu Ala Ser Ile Gly Tyr Met

245 250 255

Thr Asn Asp Pro Ala Gly Thr Val Ile Val

260 265

<210> 87

<211> 798

<212> DNA

<213> Armillaria mellea

<400> 87

atgtccttct tcgacagcgt gaaacttgac ctcgtcggac gcctctttgg catcaggaat 60

cgcggcttag ctgttacttg ttgtgctgtg gcagtcgcct ctatcatcgc gttcccttac 120

attcgtaggg actaccagac atttttatct gggggtccct cctacgctcc ccagaacatc 180

agaggatacc tcattgtctg cgtcctggcc ttgttccgtc aggagcaaaa aggccttgcg 240

atatacgacc gccttcccga gaagcgcagg tggctacctg acttgcctcc tcgcgatggc 300

ccacggccca tcacgaccag ccatataatc caaagacagc gcaaccaggc gccggacctc 360

aagttcgccc tcgaggaact caaggccacg gtcattccac gggtgcaggc tcgccacact 420

gacctcaccc atctcagcct atccaagttc gagttccatg ctgaagcaat cttcctgctc 480

ccctctgtac ccatcgatga tccaaagaat gtgccaagtc acgacacggt gcgcaggacg 540

aagagggaaa ttgcgcatat gcacgactac catgactaca cgctgcatct tgcgttggcc 600

gcccaagacg ggaaggaagt cgtatcaaag ggatgggggc agcgacaccc gctggcaggc 660

cctggcgttc ctggtccacc gacggagtgg acgtttattt atgcgccacg taacgaagag 720

gagctggcag tggtggaaat gattatcgag gcatcgatag gctatatgac caatgaccct 780

gcaggaaaaa ctatcgca 798

<210> 88

<211> 266

<212>PRT

<213> Armillaria mellea

<400> 88

Met Ser Phe Phe Asp Ser Val Lys Leu Asp Leu Val Gly Arg Leu Phe

1 5 10 15

Gly Ile Arg Asn Arg Gly Leu Ala Val Thr Cys Cys Ala Val Ala Val

20 25 30

Ala Ser Ile Ile Ala Phe Pro Tyr Ile Arg Arg Asp Tyr Gln Thr Phe

35 40 45

Leu Ser Gly Gly Pro Ser Tyr Ala Pro Gln Asn Ile Arg Gly Tyr Leu

50 55 60

Ile Val Cys Val Leu Ala Leu Phe Arg Gln Glu Gln Lys Gly Leu Ala

65 70 75 80

Ile Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Pro Asp Leu Pro

85 90 95

Pro Arg Asp Gly Pro Arg Pro Ile Thr Thr Ser His Ile Ile Gln Arg

100 105 110

Gln Arg Asn Gln Ala Pro Asp Leu Lys Phe Ala Leu Glu Glu Leu Lys

115 120 125

Ala Thr Val Ile Pro Arg Val Gln Ala Arg His Thr Asp Leu Thr His

130 135 140

Leu Ser Leu Ser Lys Phe Glu Phe His Ala Glu Ala Ile Phe Leu Leu

145 150 155 160

Pro Ser Val Pro Ile Asp Asp Pro Lys Asn Val Pro Ser His Asp Thr

165 170 175

Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp

180 185 190

Tyr Thr Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys Glu Val Val

195 200 205

Ser Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Val Pro

210 215 220

Gly Pro Pro Thr Glu Trp Thr Phe Ile Tyr Ala Pro Arg Asn Glu Glu

225 230 235 240

Glu Leu Ala Val Val Glu Met Ile Ile Glu Ala Ser Ile Gly Tyr Met

245 250 255

Thr Asn Asp Pro Ala Gly Lys Thr Ile Ala

260 265

<210> 89

<211> 798

<212> DNA

<213> Armillaria fuscipes

<400> 89

atgaccttct tggacagtat caaacttgac ctcgttgggc gcctctttgg catcaggaat 60

cacggcgtag ccgctgcctg ttgtgctgca gcagttgcct ctgccatcgt gttcccttat 120

attcgtaggg actaccagac atttctatct ggcggccctt cctacgctcc ccagaacatc 180

agaggataca tcattgtctg cgtcctagcc ttattccgcc aggagcaaaa aggccttgcg 240

atatatgacc gccttcccga gaagcgcagg tggttagctg acttgcctcc tcgcaatggc 300

ccacggccca tcacaaccag tcatataatt caaagacagc gcaaccaggc gccagacccc 360

aagttcgccc tcgaagaact caaggccaca gtcattccac gggtacaggc tcgccacact 420

gacctcaccc atctcagcct gtccaaattc gagtttcacg ctgaagcaat cttcctgctc 480

ccctctgtac ccatcgacga cccaaagaat atcccaagcc atgacacagt gcgcaggacg 540

aaaagggaga tcgcgcatat gcacgactat catgatttca cgctgcatct tgcactggct 600

gcccaagacg ggaaggaagt cgtatcaaag ggatgggggc agcggcaccc gctggcaggc 660

cctggtgtcc ctggtccacc aacggagtgg acgtttattt acgcgccacg gaacgaagag 720

gagctggcag tagtggaaat gataattgag gcatcaatag gctacatgac caatgaccct 780

gcaggatcag ttattcca 798

<210> 90

<211> 266

<212>PRT

<213> Armillaria fuscipes

<400> 90

Met Thr Phe Leu Asp Ser Ile Lys Leu Asp Leu Val Gly Arg Leu Phe

1 5 10 15

Gly Ile Arg Asn His Gly Val Ala Ala Ala Cys Cys Ala Ala Ala Val

20 25 30

Ala Ser Ala Ile Val Phe Pro Tyr Ile Arg Arg Asp Tyr Gln Thr Phe

35 40 45

Leu Ser Gly Gly Pro Ser Tyr Ala Pro Gln Asn Ile Arg Gly Tyr Ile

50 55 60

Ile Val Cys Val Leu Ala Leu Phe Arg Gln Glu Gln Lys Gly Leu Ala

65 70 75 80

Ile Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Ala Asp Leu Pro

85 90 95

Pro Arg Asn Gly Pro Arg Pro Ile Thr Thr Ser His Ile Ile Gln Arg

100 105 110

Gln Arg Asn Gln Ala Pro Asp Pro Lys Phe Ala Leu Glu Glu Leu Lys

115 120 125

Ala Thr Val Ile Pro Arg Val Gln Ala Arg His Thr Asp Leu Thr His

130 135 140

Leu Ser Leu Ser Lys Phe Glu Phe His Ala Glu Ala Ile Phe Leu Leu

145 150 155 160

Pro Ser Val Pro Ile Asp Asp Pro Lys Asn Ile Pro Ser His Asp Thr

165 170 175

Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp

180 185 190

Phe Thr Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys Glu Val Val

195 200 205

Ser Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Val Pro

210 215 220

Gly Pro Pro Thr Glu Trp Thr Phe Ile Tyr Ala Pro Arg Asn Glu Glu

225 230 235 240

Glu Leu Ala Val Val Glu Met Ile Ile Glu Ala Ser Ile Gly Tyr Met

245 250 255

Thr Asn Asp Pro Ala Gly Ser Val Ile Pro

260 265

<210> 91

<211> 738

<212> DNA

<213> Mycena citricolor

<400> 91

atggcttatc agctcacttg gattcagact ctcgtgctgg gtgcccttgt ggcaatggca 60

gtagcgttcc ccttcatcaa gaaagactac gagacgttcc tgaagggcgg cccctcctat 120

gcgccccaaa acgttcgcgg atacatcatc gtgctcgtgc tcgcgctctt ccgccaagag 180

cagctcgggc tggagatcta cgaccgcatg cccgagaaac gtcgctggct cgcgaatctc 240

cctcagcgcg agggcccccg ccccaagacc acaagtcaca tcatccagcg gcagctcagc 300

cagcacacgg accccgcatt cggcgccgcg tacctcaaag acaccgtcat tccgcgcgtc 360

caggcgcggc acgcagccaa cacgcacatc gcgcgctcga cgttcgagtt ccacgccgcc 420

gcgatcttcc tgaacgcgga cgtgccgctg cccgagggcc tgcccgcaag cgagacggtg 480

cggcggacca agggcgagat cgcgcacatg cacgactacc acgacttcac gctgcacctc 540

gcgctcgcgg cggcggatgg gaaggaggtg gtcggcaagg gctgggggca gcgccatccg 600

ctggcgggac ccggtgtgcc gggtccgccg aacgagtgga cctttgtgta tgcgccgagg 660

aatgaagagg agatgggcgt ggtcgagcag atcgtagagg cggcgattgg gtacatgtcg 720

aacgtgcctg cgctggaa 738

<210> 92

<211> 246

<212>PRT

<213> Mycena citricolor

<400> 92

Met Ala Tyr Gln Leu Thr Trp Ile Gln Thr Leu Val Leu Gly Ala Leu

1 5 10 15

Val Ala Met Ala Val Ala Phe Pro Phe Ile Lys Lys Asp Tyr Glu Thr

20 25 30

Phe Leu Lys Gly Gly Pro Ser Tyr Ala Pro Gln Asn Val Arg Gly Tyr

35 40 45

Ile Ile Val Leu Val Leu Ala Leu Phe Arg Gln Glu Gln Leu Gly Leu

50 55 60

Glu Ile Tyr Asp Arg Met Pro Glu Lys Arg Arg Trp Leu Ala Asn Leu

65 70 75 80

Pro Gln Arg Glu Gly Pro Arg Pro Lys Thr Thr Ser His Ile Ile Gln

85 90 95

Arg Gln Leu Ser Gln His Thr Asp Pro Ala Phe Gly Ala Ala Tyr Leu

100 105 110

Lys Asp Thr Val Ile Pro Arg Val Gln Ala Arg His Ala Ala Asn Thr

115 120 125

His Ile Ala Arg Ser Thr Phe Glu Phe His Ala Ala Ala Ile Phe Leu

130 135 140

Asn Ala Asp Val Pro Leu Pro Glu Gly Leu Pro Ala Ser Glu Thr Val

145 150 155 160

Arg Arg Thr Lys Gly Glu Ile Ala His Met His Asp Tyr His Asp Phe

165 170 175

Thr Leu His Leu Ala Leu Ala Ala Ala Asp Gly Lys Glu Val Val Gly

180 185 190

Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Val Pro Gly

195 200 205

Pro Pro Asn Glu Trp Thr Phe Val Tyr Ala Pro Arg Asn Glu Glu Glu

210 215 220

Met Gly Val Val Glu Gln Ile Val Glu Ala Ala Ile Gly Tyr Met Ser

225 230 235 240

Asn Val Pro Ala Leu Glu

245

<210> 93

<211> 747

<212> DNA

<213> Panellus stipticus

<400> 93

atgaacatca acctgaaagc tctgatcgga gtctgtgccg tgctcatcac cgctgcagtg 60

ttccccttcg ttcgtaaaga ctatcacacc tttcttgaag gtggaccatc ctacgcgccg 120

cagaatttgc aaggctatat catcgtgttg gtgctctcac tctttcgagg ggagaggacg 180

ggattggaaa tatacgaccg cttgcccgaa aaacgccgct ggctcgagga gctgcctgtt 240

cgcgaaggcc cgcgcccaaa gacaaccagc cacatcattc agagacaatt gaatcagcac 300

gttgacccgg acttcggaat gaactctttg aaaggctccg tcatccggcg ccttcaatcc 360

cgccaccagg acataactca actcgcactc tcgaaattcg aattccacgc cgaggccata 420

tttctgcgcc ccgatatcgc gatcaacgat cccaaacacg tcccgagcca cgacacggtg 480

cgccgcacaa agcgcgagat agctcacatg cacgactacc atgattacac gtgtcatttg 540

gcgctcgcag cgcaggatgg gaagcaagtg attgcaaaag ggtggggcca gagacatccg 600

ctcgcgggac cgggcatgcc ggggccgccg acggagtgga catttttgta tgcgccgagg 660

aatgaggcgg aggttcaagt gttggagacg attatcgaag cgtcaatcgg gtacatgtcg 720

aacgcaccag ccttgggtgg gagcgag 747

<210> 94

<211> 249

<212>PRT

<213> Panellus stipticus

<400> 94

Met Asn Ile Asn Leu Lys Ala Leu Ile Gly Val Cys Ala Val Leu Ile

1 5 10 15

Thr Ala Ala Val Phe Pro Phe Val Arg Lys Asp Tyr His Thr Phe Leu

20 25 30

Glu Gly Gly Pro Ser Tyr Ala Pro Gln Asn Leu Gln Gly Tyr Ile Ile

35 40 45

Val Leu Val Leu Ser Leu Phe Arg Gly Glu Glu Thr Gly Leu Glu Ile

50 55 60

Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Glu Glu Leu Pro Val

65 70 75 80

Arg Glu Gly Pro Arg Pro Lys Thr Thr Ser His Ile Ile Gln Arg Gln

85 90 95

Leu Asn Gln His Val Asp Pro Asp Phe Gly Met Asn Ser Leu Lys Gly

100 105 110

Ser Val Ile Arg Arg Leu Gln Ser Arg His Gln Asp Ile Thr Gln Leu

115 120 125

Ala Leu Ser Lys Phe Glu Phe His Ala Glu Ala Ile Phe Leu Arg Pro

130 135 140

Asp Ile Ala Ile Asn Asp Pro Lys His Val Pro Ser His Asp Thr Val

145 150 155 160

Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp Tyr

165 170 175

Thr Cys His Leu Ala Leu Ala Ala Gln Asp Gly Lys Gln Val Ile Ala

180 185 190

Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Met Pro Gly

195 200 205

Pro Pro Thr Glu Trp Thr Phe Leu Tyr Ala Pro Arg Asn Glu Ala Glu

210 215 220

Val Gln Val Leu Glu Thr Ile Ile Glu Ala Ser Ile Gly Tyr Met Ser

225 230 235 240

Asn Ala Pro Ala Leu Gly Gly Ser Glu

245

<210> 95

<211> 792

<212> DNA

<213> Neonothopanus gardneri

<400> 95

atgaatcttc cgtctttcgt ccaacgtctc tccacagcaa gcagtcgcag tatagcgatt 60

acttgcgtag ttgtccttgc ctctgcaatc gcctttccct tcatccgcag agactaccag 120

accttcctgg aagtgggacc ctcgtacgcc ccgcagaact ttagaggata catcatcgtc 180

tgtgtcctct cgttgttccg ccaagaacaa aaaggactcg aaatctacga tcggctccca 240

gagaaacgaa ggtggttgtc cgaccttccc tttcgtgacg ggcccagacc cagcatcaca 300

agccatatca ttcaacgaca gcgtacccaa ctagttgatc cggacttcgc tacccaggag 360

ctcataggca aagtcatccc tcgtgtgcaa gcacgacaca ccgacaaaac attcctcagc 420

acctccaaat tcgaatttca cgcaaaagcc atattcctcc tgccttccat cccaatcaac 480

gaccctctga acgttccaag ccacgacact gtccgacgaa cgaagcgcga gatcgcgcat 540

atgcatgatt atcatgattg cactcttcac atcgctctcg ctgctcagga cggaaaggag 600

gttttgaaga agggatgggg gcaacgacac ccactcgctg gacctggagt gcccggccca 660

ccgacggagt ggacgtttct ctatgcgcct cgaaacgaag aagaggttcg agttgtggag 720

atgattattg aggctgccat aggttacatg acgaatgatc cggcaggaaa agttgtagaa 780

gccactggaa ag 792

<210> 96

<211> 264

<212>PRT

<213> Neonothopanus gardneri

<400> 96

Met Asn Leu Pro Ser Phe Val Gln Arg Leu Ser Thr Ala Ser Ser Arg

1 5 10 15

Ser Ile Ala Ile Thr Cys Val Val Val Leu Ala Ser Ala Ile Ala Phe

20 25 30

Pro Phe Ile Arg Arg Asp Tyr Gln Thr Phe Leu Glu Val Gly Pro Ser

35 40 45

Tyr Ala Pro Gln Asn Phe Arg Gly Tyr Ile Ile Val Cys Val Leu Ser

50 55 60

Leu Phe Arg Gln Glu Gln Lys Gly Leu Glu Ile Tyr Asp Arg Leu Pro

65 70 75 80

Glu Lys Arg Arg Trp Leu Ser Asp Leu Pro Phe Arg Asp Gly Pro Arg

85 90 95

Pro Ser Ile Thr Ser His Ile Ile Gln Arg Gln Arg Thr Gln Leu Val

100 105 110

Asp Pro Asp Phe Ala Thr Gln Glu Leu Ile Gly Lys Val Ile Pro Arg

115 120 125

Val Gln Ala Arg His Thr Asp Lys Thr Phe Leu Ser Thr Ser Lys Phe

130 135 140

Glu Phe His Ala Lys Ala Ile Phe Leu Leu Pro Ser Ile Pro Ile Asn

145 150 155 160

Asp Pro Leu Asn Val Pro Ser His Asp Thr Val Arg Arg Thr Lys Arg

165 170 175

Glu Ile Ala His Met His Asp Tyr His Asp Cys Thr Leu His Ile Ala

180 185 190

Leu Ala Ala Gln Asp Gly Lys Glu Val Leu Lys Lys Gly Trp Gly Gln

195 200 205

Arg His Pro Leu Ala Gly Pro Gly Val Pro Gly Pro Pro Thr Glu Trp

210 215 220

Thr Phe Leu Tyr Ala Pro Arg Asn Glu Glu Glu Val Arg Val Val Glu

225 230 235 240

Met Ile Ile Glu Ala Ala Ile Gly Tyr Met Thr Asn Asp Pro Ala Gly

245 250 255

Lys Val Val Glu Ala Thr Gly Lys

260

<210> 97

<211> 753

<212> DNA

<213> Mycena chlorophos

<400> 97

atggtccaac tcacccgaac ctccggattc atcgccgctg cggccattgt tgctgccatc 60

gccttcccgt tcattcgtcg agactaccag acgttccttc gtggtgggcc gtcctatgcc 120

ccacagaaca tccgcggcta tatcatcgtc ctggttctgt ccctcttccg cggcgaggag 180

aagggtcttg caatctacga gccccttcct gagaagcgca catggctgcc ggagcttccg 240

cggcgcgcgg gagaccggcc caagacgacg agccacatca tccaacggca gctcgaccag 300

taccccgacc cggactttgt cctcaaagcc ctgaaagcga cggtcatccc gcgtgtccaa 360

gcccggcaca cagacaagac tcacctcgcg ctgtccaagt tcgagttcca tgctgaggcc 420

atcttcgtgc gcccggaaat cgccatcgac gacccgaagc atatccccag ccacgacacg 480

gtgcgacgga cgaagcgcga gattgcgcac atgcacgact atcacgactg cacgctgcat 540

ttggcgctag cggcgcagga cgcgaagcag gtgctgcaga agggctgggg ccagcgccat 600

ccgctggcag ggcctgggat gcccggggccg cccacggagt ggacgttctt gtatgccccg 660

aggaccgagg aggaagtgaa ggttgtggag accattgtcg aggcctctat cgcgtacatg 720

acgaacgcgg agaagccggt cgagctggtg cag 753

<210> 98

<211> 251

<212>PRT

<213> Mycena chlorophos

<400> 98

Met Val Gln Leu Thr Arg Thr Ser Gly Phe Ile Ala Ala Ala Ala Ile

1 5 10 15

Val Ala Ala Ile Ala Phe Pro Phe Ile Arg Arg Asp Tyr Gln Thr Phe

20 25 30

Leu Arg Gly Gly Pro Ser Tyr Ala Pro Gln Asn Ile Arg Gly Tyr Ile

35 40 45

Ile Val Leu Val Leu Ser Leu Phe Arg Gly Glu Glu Lys Gly Leu Ala

50 55 60

Ile Tyr Glu Pro Leu Pro Glu Lys Arg Thr Trp Leu Pro Glu Leu Pro

65 70 75 80

Arg Arg Ala Gly Asp Arg Pro Lys Thr Thr Ser His Ile Ile Gln Arg

85 90 95

Gln Leu Asp Gln Tyr Pro Asp Pro Asp Phe Val Leu Lys Ala Leu Lys

100 105 110

Ala Thr Val Ile Pro Arg Val Gln Ala Arg His Thr Asp Lys Thr His

115 120 125

Leu Ala Leu Ser Lys Phe Glu Phe His Ala Glu Ala Ile Phe Val Arg

130 135 140

Pro Glu Ile Ala Ile Asp Asp Pro Lys His Ile Pro Ser His Asp Thr

145 150 155 160

Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp Tyr His Asp

165 170 175

Cys Thr Leu His Leu Ala Leu Ala Ala Gln Asp Ala Lys Gln Val Leu

180 185 190

Gln Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro Gly Met Pro

195 200 205

Gly Pro Pro Thr Glu Trp Thr Phe Leu Tyr Ala Pro Arg Thr Glu Glu

210 215 220

Glu Val Lys Val Val Glu Thr Ile Val Glu Ala Ser Ile Ala Tyr Met

225 230 235 240

Thr Asn Ala Glu Lys Pro Val Glu Leu Val Gln

245 250

<210> 99

<211> 801

<212> DNA

<213> Artificial sequence

<220>

<223> Optimized for expression in mammals

(humanized) nucleic acid encoding protein SEQ ID

No:80

<400> 99

atgcgcatta acatctccct ttcatctctt ttcgagcgat tgagcaaact gagttccagg 60

agtattgcaa tcacttgtgg ggttgtcctc gcgagcgcca tcgcattccc catcatccgg 120

agagattatc agacgtttct tgaggtgggc cctagctatg caccacagaa cttccgagga 180

tatatcatcg tgtgtgtact gtcactgttt aggcaagaac aaaagggatt ggctatctat 240

gataggttgc ctgagaaacg gcggtggctc gctgatctcc catttagaga ggggacacga 300

ccgagcatca cttcacacat catacaaaga cagcgaacgc agctcgttga ccaagagttc 360

gcaactaggg aactgattga taaggtgata cccagagtac aggcgcgaca caccgataaa 420

acttttcttt ccacctctaa attcgagttc catgccaaag ctattttctt gttgccttcc 480

ataccgatta atgatcctct gaatattcca tcccacgaca cagttcgacg gacgaaacgc 540

gaaattgcgc acatgcacga ctatcacgat tgcactttgc acctggcact ggctgctcaa 600

gacggaaaag aagttctgaa aaagggttgg gggcaaagac atccgctggc gggacccggt 660

gtacctgggc cgcctacgga atggacattt ttgtacgcac cgaggaacga agaggaggcc 720

agggtcgttg agatgattgt tgaggctagt attgggtaca tgacgaatga tccggctggt 780

aaaattgttg aaaatgcaaa g 801

<210> 100

<211> 1266

<212> DNA

<213> Artificial sequence

<220>

<223> Optimized for expression in mammals

(humanized) nucleic acid encoding protein SEQ ID

No:2

<400> 100

atggcgtcat tcgagaacag tcttagcgtg ctgatcgtcg gcgctgggct cggcggtctt 60

gctgcggcaa tcgccctcag gcgacaagga cacgttgtta aaatctatga cagctcatca 120

tttaaggcag agttgggcgc aggcctcgcg gtccccccaa acactttgag atcactgcaa 180

caactgggtt gtaatactga gaaccttaac ggcgtggata acctctgctt cactgcaatg 240

ggttacgatg gcagtgtggg tatgatgaac aatatgaccg attataggga ggcgtacggc 300

actagctgga taatggtcca tcgggttgat ctccacaatg agcttatgcg cgtagcgttg 360

gatccgggcg gattgggacc cccagctacc ttgcacttga atcaccgcgt gactttttgt 420

gatgtcgacg catgcacagt aaccttcacc aatgggacga ctcagtcagc ggatctcatc 480

gtcggcgccg acggtatacg atccactatc cgcagattcg tcctggagga agatgtcaca 540

gttccggcat ccggaatcgt tggtttccgc tggctcgtcc aggctgatgc tttggatcct 600

taccctgaac ttgactggat tgttaaaaag ccccctctcg gcgctaggtt gataagtacg 660

cctcaaaacc cgcagtctgg ggtaggtctc gcggatcgca gaacgatcat tatatacgcg 720

tgtcgaggag gtactatggt aaacgtactt gccgtccatg acgatgagag ggatcagaat 780

acggcagatt ggtccgtgcc agctagcaag gatgatcttt tcagagtttt tcacgactat 840

catcctcgat ttcggcggct gttggagttg gcgcaagaca tcaatctgtg gcagatgcgg 900

gtggtccccg ttctgaagaa atgggtgaac aaaagagtct gtctcttggg ggatgcagcg 960

catgcgtccc tccctacctt ggggcagggt ttcggcatgg ggttggagga cgccgtagcc 1020

cttgggactt tgcttccaaa ggggacgaca gcatcccaaa tagaaacaag acttgccgta 1080

tatgagcagc tccgaaaaga tcgcgccgag ttcgtcgcgg ctgagtccta cgaagaacaa 1140

tatgtaccag aaatgagggg actttacctg cgatccaaag aattgcgcga ccgggtaatg 1200

ggctatgaca taaaggtgga gtccgaaaag gtcttggaga cactgttgcg gtcaagcaat 1260

tccgcc 1266

<210> 101

<211> 2112

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleic acid. encoding fusion protein

hispidin hydroxylase and luciferase

<400> 101

atggcatcgt ttgagaattc tctaagcgtt ttgattgtcg gggccggact tggtgggctt 60

gctgctgcca tcgcgctgcg tcgccaaggg catgtcgtga aaatatacga ctcctctagc 120

ttcaaagccg aacttggtgc gggactcgct gtgccgccta acaccttgcg cagtctacag 180

caacttggtt gcaataccga gaacctcaat ggtgtggata atctttgctt cactgcgatg 240

gggtatgacg ggagtgtagg gatgatgaac aacatgactg actatcgaga ggcatacggt 300

acttcttgga tcatggtcca ccgcgttgac ttgcataacg agctgatgcg cgtagcactt 360

gatccaggtg ggctcggacc tcctgcgaca ctccatctta atcatcgtgt cacattctgc 420

gatgtcgacg cttgcaccgt gacattcacc aacgggacca ctcaatcagc tgatctcatc 480

gttggtgcag acggtatacg ctctaccatt cggcggtttg tcttagaaga agacgtgact 540

gtgcctgcgt caggaatcgt cgggtttcga tggcttgtac aagctgacgc gctggaccca 600

tatcctgaac tcgactggat tgttaaaaag cctcctctag gcgcgcgact gatctccact 660

cctcagaatc cacagtctgg tgttggcttg gctgacaggc gcactatcat catctacgca 720

tgtcgtggcg gcaccatggt caatgtcctt gcagtgcatg atgacgaacg tgaccagaac 780

accgcagatt ggagtgtacc ggcttccaaa gacgatctat ttcgtgtttt ccacgattac 840

catccacgct ttcggcggct tttagagctt gcgcaggata ttaatctctg gcaaatgcgt 900

gttgtacctg ttttgaaaaa atgggttaac aagcgggttt gcttgttagg agatgctgcg 960

cacgcttctt taccgacgtt gggtcaaggt tttggtatgg gcctggaaga tgccgtagca 1020

cttggtacac tccttccaaa gggtaccact gcatctcaga tcgagactcg acttgcggtg 1080

tacgaacagc tacgtaagga tcgtgcggaa tttgttgcgg ctgaatcata tgaagagcaa 1140

tatgttcctg aaatgcgggg actttatctg aggtcaaagg aactgcgtga tagagtcatg 1200

ggttatgata tcaaagtgga gagcgagaag gttctcgaga cgctcctaag aagttctaat 1260

tctgcctccg gaactggggg caacgcctct gacggtggtg ggtctggtgg tatgcgcatt 1320

aacattagcc tctcgtctct cttcgaacgt ctctccaaac ttagcagtcg cagcatagcg 1380

attacatgtg gagttgttct cgcctccgca atcgcctttc ccatcatccg cagagactac 1440

cagactttcc tagaagtggg accctcgtac gctccgcaga actttagagg atacatcatc 1500

gtctgtgtcc tctcgctatt ccgccaagag cagaaagggc tcgccatcta tgatcgtctt 1560

cccgagaaac gcaggtggtt ggccgacctt ccctttcgtg aaggaaccag acccagcatt 1620

accagccata tcattcagcg acagcgcact caactggtcg atcaggagtt tgccaccagg 1680

gagctcatag acaaggtcat ccctcgcgtg caagcacgac acaccgacaa aacgttcctc 1740

agcacatcaa agttcgagtt tcatgcgaag gccatatttc tcttgccttc tatcccaatc 1800

aacgaccctc tgaatatccc tagccacgac actngtccgcc gaacgaagcg cgagattgca 1860

catatgcatg attatcatga ttgcacactt catcttgctc tcgctgcgca ggatggaaag 1920

gaggtgctga agaaaggttg gggacaacga catcctttgg ctggtcctgg agttcctggt 1980

ccaccaacgg aatggacttt tctttatgcg cctcgcaacg aagaagaggc tcgagtagtg 2040

gagatgatcg ttgaggcttc catagggtat atgacgaacg atcctgcagg aaagattgta 2100

gaaaacgcca ag 2112

<210> 102

<211> 704

<212>PRT

<213> Artificial sequence

<220>

<223> Amino acid sequence of fusion protein

hispidin hydroxylase and luciferase

<400> 102

Met Ala Ser Phe Glu Asn Ser Leu Ser Val Leu Ile Val Gly Ala Gly

1 5 10 15

Leu Gly Gly Leu Ala Ala Ala Ile Ala Leu Arg Arg Gln Gly His Val

20 25 30

Val Lys Ile Tyr Asp Ser Ser Ser Phe Lys Ala Glu Leu Gly Ala Gly

35 40 45

Leu Ala Val Pro Pro Asn Thr Leu Arg Ser Leu Gln Gln Leu Gly Cys

50 55 60

Asn Thr Glu Asn Leu Asn Gly Val Asp Asn Leu Cys Phe Thr Ala Met

65 70 75 80

Gly Tyr Asp Gly Ser Val Gly Met Met Asn Asn Met Thr Asp Tyr Arg

85 90 95

Glu Ala Tyr Gly Thr Ser Trp Ile Met Val His Arg Val Asp Leu His

100 105 110

Asn Glu Leu Met Arg Val Ala Leu Asp Pro Gly Gly Leu Gly Pro Pro

115 120 125

Ala Thr Leu His Leu Asn His Arg Val Thr Phe Cys Asp Val Asp Ala

130 135 140

Cys Thr Val Thr Phe Thr Asn Gly Thr Thr Gln Ser Ala Asp Leu Ile

145 150 155 160

Val Gly Ala Asp Gly Ile Arg Ser Thr Ile Arg Arg Phe Val Leu Glu

165 170 175

Glu Asp Val Thr Val Pro Ala Ser Gly Ile Val Gly Phe Arg Trp Leu

180 185 190

Val Gln Ala Asp Ala Leu Asp Pro Tyr Pro Glu Leu Asp Trp Ile Val

195 200 205

Lys Lys Pro Pro Leu Gly Ala Arg Leu Ile Ser Thr Pro Gln Asn Pro

210 215 220

Gln Ser Gly Val Gly Leu Ala Asp Arg Arg Thr Ile Ile Ile Tyr Ala

225 230 235 240

Cys Arg Gly Gly Thr Met Val Asn Val Leu Ala Val His Asp Asp Glu

245 250 255

Arg Asp Gln Asn Thr Ala Asp Trp Ser Val Pro Ala Ser Lys Asp Asp

260 265 270

Leu Phe Arg Val Phe His Asp Tyr His Pro Arg Phe Arg Arg Leu Leu

275 280 285

Glu Leu Ala Gln Asp Ile Asn Leu Trp Gln Met Arg Val Val Pro Val

290 295 300

Leu Lys Lys Trp Val Asn Lys Arg Val Cys Leu Leu Gly Asp Ala Ala

305 310 315 320

His Ala Ser Leu Pro Thr Leu Gly Gln Gly Phe Gly Met Gly Leu Glu

325 330 335

Asp Ala Val Ala Leu Gly Thr Leu Leu Pro Lys Gly Thr Thr Ala Ser

340 345 350

Gln Ile Glu Thr Arg Leu Ala Val Tyr Glu Gln Leu Arg Lys Asp Arg

355 360 365

Ala Glu Phe Val Ala Ala Glu Ser Tyr Glu Glu Gln Tyr Val Pro Glu

370 375 380

Met Arg Gly Leu Tyr Leu Arg Ser Lys Glu Leu Arg Asp Arg Val Met

385 390 395 400

Gly Tyr Asp Ile Lys Val Glu Ser Glu Lys Val Leu Glu Thr Leu Leu

405 410 415

Arg Ser Ser Asn Ser Ala Ser Gly Thr Gly Gly Asn Ala Ser Asp Gly

420 425 430

Gly Gly Ser Gly Gly Met Arg Ile Asn Ile Ser Leu Ser Ser Leu Phe

435 440 445

Glu Arg Leu Ser Lys Leu Ser Ser Arg Ser Ile Ala Ile Thr Cys Gly

450 455 460

Val Val Leu Ala Ser Ala Ile Ala Phe Pro Ile Ile Arg Arg Asp Tyr

465 470 475 480

Gln Thr Phe Leu Glu Val Gly Pro Ser Tyr Ala Pro Gln Asn Phe Arg

485 490 495

Gly Tyr Ile Ile Val Cys Val Leu Ser Leu Phe Arg Gln Glu Gln Lys

500 505 510

Gly Leu Ala Ile Tyr Asp Arg Leu Pro Glu Lys Arg Arg Trp Leu Ala

515 520 525

Asp Leu Pro Phe Arg Glu Gly Thr Arg Pro Ser Ile Thr Ser His Ile

530 535 540

Ile Gln Arg Gln Arg Thr Gln Leu Val Asp Gln Glu Phe Ala Thr Arg

545 550 555 560

Glu Leu Ile Asp Lys Val Ile Pro Arg Val Gln Ala Arg His Thr Asp

565 570 575

Lys Thr Phe Leu Ser Thr Ser Lys Phe Glu Phe His Ala Lys Ala Ile

580 585 590

Phe Leu Leu Pro Ser Ile Pro Ile Asn Asp Pro Leu Asn Ile Pro Ser

595 600 605

His Asp Thr Val Arg Arg Thr Lys Arg Glu Ile Ala His Met His Asp

610 615 620

Tyr His Asp Cys Thr Leu His Leu Ala Leu Ala Ala Gln Asp Gly Lys

625 630 635 640

Glu Val Leu Lys Lys Gly Trp Gly Gln Arg His Pro Leu Ala Gly Pro

645 650 655

Gly Val Pro Gly Pro Pro Thr Glu Trp Thr Phe Leu Tyr Ala Pro Arg

660 665 670

Asn Glu Glu Glu Ala Arg Val Val Glu Met Ile Val Glu Ala Ser Ile

675 680 685

Gly Tyr Met Thr Asn Asp Pro Ala Gly Lys Ile Val Glu Asn Ala Lys

690 695 700

<210> 103

<211> 1266

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleic acid optimized for expression in plants,

encoding protein SEQ ID No:2

<400> 103

atggcatctt ttgagaattc tctgagcgtt ttgattgtcg gggccggact tggtgggctt 60

gctgctgcca tcgccctgcg tcgacaaggg catgtcgtga aaatatacga ctcctctagc 120

ttcaaagccg aacttggtgc tggactcgct gtgccgccta acaccttgcg tagtctccag 180

caacttggtt gcaataccga gaacctcaat ggtgtggata atctttgctt cactgctatg 240

gggtatgacg ggagtgtagg gatgatgaac aacatgactg actatcgaga ggcatacggt 300

acttcttgga tcatggtcca cagagttgac ttgcataacg agctgatgag ggtagcactt 360

gatccaggtg ggctcggacc tcctgcaaca ctccatctta atcatcgtgt cacattctgc 420

gatgtcgacg cttgcaccgt gacattcacc aacgggacca ctcaatcagc tgatctcatc 480

gttggtgcag acggtataag atctaccatt cgaaggtttg tcctggaaga agacgtgact 540

gtgcctgcat caggaatcgt cgggtttcga tggcttgtac aagctgacgc tctggaccca 600

tatcctgaac tcgactggat tgttaaaaag cctcctcttg gcgcacgact gatctccact 660

cctcagaatc cacagtctgg tgttggcttg gctgacagga ggactatcat catctacgca 720

tgtcgtggcg gcaccatggt caatgtcctt gcagtgcatg atgacgaacg tgaccagaac 780

accgcagatt ggagtgtacc ggcttccaaa gacgatctct ttcgtgtttt ccacgattac 840

catccacgat ttaggaggct tctcgagctt gctcaggata ttaatctctg gcaaatgcgt 900

gttgtacctg ttttgaaaaa atgggttaac aagagggttt gcttgctcgg agatgctgca 960

cacgcttctc tgccgacgtt gggtcaaggt tttggtatgg gtctggaaga tgccgtagca 1020

cttggtacac tccttccaaa gggtaccact gcatctcaga tcgagactcg acttgctgtg 1080

tacgaacagt tgcgtaagga tcgtgccgaa tttgttgccg ctgaatcata tgaagagcaa 1140

tatgttcctg aaatgcgagg actttatctg aggtcaaagg aactgcgtga tagagtcatg 1200

ggttatgata tcaaagtgga gagcgagaag gttctcgaga cgctccttag aagttctaat 1260

tctgcc 1266

<210> 104

<211> 1032

<212> DNA

<213> Aspergillus nidulans

<400> 104

atggtgcagg acacgtcatc tgcttctact tcccccattt tgacacgttg gtacatcgac 60

acaagaccct taacagcatc cacggcagca cttcctcttc tggaaacgtt gcagcctgca 120

gatcagatta gtgtacagaa gtattatcat ttaaaggata aacatatgag tcttgcttct 180

aacttattga agtatctgtt tgttcatcgt aattgtcgta ttccctggag ttctatcgtg 240

atctcccgta cacctgatcc tcaccgtcgt ccatgctaca ttcctccctc aggctctcag 300

gaagattcct tcaaagacgg atacactggt atcaatgttg agttcaatgt atcacaccaa 360

gccagtatgg tcgccattgc tggaaccgca tttactccca attctggtgg cgattctaaa 420

ctgaagcctg aggtgggaat cgacatcaca tgtgtaaacg agcgtcaggg ccgtaacggt 480

gaggagcgtt cattggaatc tcttcgtcaa tatatcgaca tcttctctga agtattctcc 540

acggctgaga tggcaaacat ccgtcgtctt gatggcgtta gtagttcttc tttaagtgct 600

gatcgtcttg ttgactatgg ttaccgtctt ttttatacat actgggccct gaaggaagcc 660

tatatcaaga tgactggtga agcattgctt gcaccatggt tacgtgagct ggagttctcc 720

aatgttgttg cccctgccgc tgttgctgaa tccggtgact ctgccggcga cttcggtgag 780

ccatacacag gagtccgtac gactttatat aaaaatttag ttgaggatgt gcgtatcgag 840

gtagccgccc ttggtggcga ttatcttttt gcaaccgcag ctcgtggagg aggaattgga 900

gccagttcaa ggcccggagg cggccctgat ggtagtggta tccgttcaca ggacccttgg 960

cgtccatta agaaacttga tatcgagcgt gacattcagc catgtgcaac aggtgtatgc 1020

aactgcctta gt 1032

<210> 105

<211> 344

<212>PRT

<213> Aspergillus nidulans

<400> 105

Met Val Gln Asp Thr Ser Ser Ala Ser Thr Ser Pro Ile Leu Thr Arg

1 5 10 15

Trp Tyr Ile Asp Thr Arg Pro Leu Thr Ala Ser Thr Ala Ala Leu Pro

20 25 30

Leu Leu Glu Thr Leu Gln Pro Ala Asp Gln Ile Ser Val Gln Lys Tyr

35 40 45

Tyr His Leu Lys Asp Lys His Met Ser Leu Ala Ser Asn Leu Leu Lys

50 55 60

Tyr Leu Phe Val His Arg Asn Cys Arg Ile Pro Trp Ser Ser Ile Val

65 70 75 80

Ile Ser Arg Thr Pro Asp Pro His Arg Arg Pro Cys Tyr Ile Pro Pro

85 90 95

Ser Gly Ser Gln Glu Asp Ser Phe Lys Asp Gly Tyr Thr Gly Ile Asn

100 105 110

Val Glu Phe Asn Val Ser His Gln Ala Ser Met Val Ala Ile Ala Gly

115 120 125

Thr Ala Phe Thr Pro Asn Ser Gly Gly Asp Ser Lys Leu Lys Pro Glu

130 135 140

Val Gly Ile Asp Ile Thr Cys Val Asn Glu Arg Gln Gly Arg Asn Gly

145 150 155 160

Glu Glu Arg Ser Leu Glu Ser Leu Arg Gln Tyr Ile Asp Ile Phe Ser

165 170 175

Glu Val Phe Ser Thr Ala Glu Met Ala Asn Ile Arg Arg Leu Asp Gly

180 185 190

Val Ser Ser Ser Ser Leu Ser Ala Asp Arg Leu Val Asp Tyr Gly Tyr

195 200 205

Arg Leu Phe Tyr Thr Tyr Trp Ala Leu Lys Glu Ala Tyr Ile Lys Met

210 215 220

Thr Gly Glu Ala Leu Leu Ala Pro Trp Leu Arg Glu Leu Glu Phe Ser

225 230 235 240

Asn Val Val Ala Pro Ala Ala Val Ala Glu Ser Gly Asp Ser Ala Gly

245 250 255

Asp Phe Gly Glu Pro Tyr Thr Gly Val Arg Thr Thr Leu Tyr Lys Asn

260 265 270

Leu Val Glu Asp Val Arg Ile Glu Val Ala Ala Leu Gly Gly Asp Tyr

275 280 285

Leu Phe Ala Thr Ala Ala Arg Gly Gly Gly Ile Gly Ala Ser Ser Arg

290 295 300

Pro Gly Gly Gly Pro Asp Gly Ser Gly Ile Arg Ser Gln Asp Pro Trp

305 310 315 320

Arg Pro Phe Lys Lys Leu Asp Ile Glu Arg Asp Ile Gln Pro Cys Ala

325 330 335

Thr Gly Val Cys Asn Cys Leu Ser

340

<210> 106

<211> 1593

<212> DNA

<213> Rhodobacter capsulatus

<400> 106

atgactcttc aatcccaaac tgctaaggat tgcttggcac tggatggtgc actgactttg 60

gtgcagtgcg aggcaatcgc cacgcatcgt agtcgtattt ccgttacgcc agccttgcgt 120

gaacgttgtg ctcgtgccca cgcacgtttg gagcacgcaa tcgcagagca gcgtcatatt 180

tacggtatca ccactggatt tggacccctg gctaaccgtc ttattggagc cgatcagggc 240

gctgagctgc aacagaatct gatttaccat cttgcaacgg gcgttggacc caagctgtcc 300

tgggcagagg cccgtgcact tatgttggca cgtcttaatt ccattttgca gggcgctagt 360

ggtgcttctc ctgaaacgat cgaccgtatc gtcgcagtac ttaacgctgg ctttgctcct 420

gaagttcccg cacaaggtac tgttggcgca agtggcgact tgactcccct tgcccacatg 480

gtactggccc tgcaaggccg tggacgtatg atcgatccta gtggacgtgt gcaagaggct 540

ggagccgtca tggatcgtct ttgcggaggc cctcttacgt tggccgcacg tgacggcctt 600

gctctggtca acggtacaag tgctatgact gctatcgctg ctcttactgg agtcgaggcc 660

gctcgtgcta tcgatgctgc acttcgtcac tccgccgttt tgatggaagt cctgtctgga 720

catgcagaag cctggcaccc cgcctttgca gagttgagac cccacccagg ccagttgcgt 780

gctactgaac gtttggccca ggccctggat ggcgcaggcc gtgtatgccg tactctgacg 840

gctgcccgtc gtcttactgc agccgacctg cgtccagagg accaccccgc acaagacgct 900

tattctctgc gtgtcgttcc tcaactggtt ggtgccgtat gggatactct tgactggcac 960

gaccgtgtcg tgacctgtga gcttaattca gtaaccgata accccatttt tccagaggt 1020

tgcgccgttc ctgcacttca cggaggaaac ttcatgggag tccacgtcgc actggcttca 1080

gacgctttga atgcagccct ggttaccctt gccggtcttg tagagcgtca aattgcccgt 1140

cttaccgacg agaaacttaa taaaggtctg cctgctttcc ttcacggtgg acaggcaggc 1200

cttcagagtg gcttcatggg cgctcaggtt acagcaactg ccctgttggc tgagatgcgt 1260

gcaaatgcaa ctccagtctc agtacaatcc ttgtccacaa acggtgcaaa ccaagacgtc 1320

gtatctatgg gaactatcgc cgcacgtcgt gctcgtgcac aattgttgcc actgtcacag 1380

attcaggcta tcttggcttt ggcattggcc caggctatgg atcttcttga cgatcccgag 1440

ggtcaagccg gatggtcttt gacagcccgt gatttgcgtg accgtatccg tgccgtttct 1500

cccggactgc gtgccgaccg tccactggca ggccatatcg aagctgttgc tcagggactt 1560

cgtcacccat ctgctgcagc cgatccacca gcc 1593

<210> 107

<211> 531

<212>PRT

<213> Rhodobacter capsulatus

<400> 107

Met Thr Leu Gln Ser Gln Thr Ala Lys Asp Cys Leu Ala Leu Asp Gly

1 5 10 15

Ala Leu Thr Leu Val Gln Cys Glu Ala Ile Ala Thr His Arg Ser Arg

20 25 30

Ile Ser Val Thr Pro Ala Leu Arg Glu Arg Cys Ala Arg Ala His Ala

35 40 45

Arg Leu Glu His Ala Ile Ala Glu Gln Arg His Ile Tyr Gly Ile Thr

50 55 60

Thr Gly Phe Gly Pro Leu Ala Asn Arg Leu Ile Gly Ala Asp Gln Gly

65 70 75 80

Ala Glu Leu Gln Gln Asn Leu Ile Tyr His Leu Ala Thr Gly Val Gly

85 90 95

Pro Lys Leu Ser Trp Ala Glu Ala Arg Ala Leu Met Leu Ala Arg Leu

100 105 110

Asn Ser Ile Leu Gln Gly Ala Ser Gly Ala Ser Pro Glu Thr Ile Asp

115 120 125

Arg Ile Val Ala Val Leu Asn Ala Gly Phe Ala Pro Glu Val Pro Ala

130 135 140

Gln Gly Thr Val Gly Ala Ser Gly Asp Leu Thr Pro Leu Ala His Met

145 150 155 160

Val Leu Ala Leu Gln Gly Arg Gly Arg Met Ile Asp Pro Ser Gly Arg

165 170 175

Val Gln Glu Ala Gly Ala Val Met Asp Arg Leu Cys Gly Gly Pro Leu

180 185 190

Thr Leu Ala Ala Arg Asp Gly Leu Ala Leu Val Asn Gly Thr Ser Ala

195 200 205

Met Thr Ala Ile Ala Ala Leu Thr Gly Val Glu Ala Ala Arg Ala Ile

210 215 220

Asp Ala Ala Leu Arg His Ser Ala Val Leu Met Glu Val Leu Ser Gly

225 230 235 240

His Ala Glu Ala Trp His Pro Ala Phe Ala Glu Leu Arg Pro His Pro

245 250 255

Gly Gln Leu Arg Ala Thr Glu Arg Leu Ala Gln Ala Leu Asp Gly Ala

260 265 270

Gly Arg Val Cys Arg Thr Leu Thr Ala Ala Arg Arg Leu Thr Ala Ala

275 280 285

Asp Leu Arg Pro Glu Asp His Pro Ala Gln Asp Ala Tyr Ser Leu Arg

290 295 300

Val Val Pro Gln Leu Val Gly Ala Val Trp Asp Thr Leu Asp Trp His

305 310 315 320

Asp Arg Val Val Thr Cys Glu Leu Asn Ser Val Thr Asp Asn Pro Ile

325 330 335

Phe Pro Glu Gly Cys Ala Val Pro Ala Leu His Gly Gly Asn Phe Met

340 345 350

Gly Val His Val Ala Leu Ala Ser Asp Ala Leu Asn Ala Ala Leu Val

355 360 365

Thr Leu Ala Gly Leu Val Glu Arg Gln Ile Ala Arg Leu Thr Asp Glu

370 375 380

Lys Leu Asn Lys Gly Leu Pro Ala Phe Leu His Gly Gly Gln Ala Gly

385 390 395 400

Leu Gln Ser Gly Phe Met Gly Ala Gln Val Thr Ala Thr Ala Leu Leu

405 410 415

Ala Glu Met Arg Ala Asn Ala Thr Pro Val Ser Val Gln Ser Leu Ser

420 425 430

Thr Asn Gly Ala Asn Gln Asp Val Val Ser Met Gly Thr Ile Ala Ala

435 440 445

Arg Arg Ala Arg Ala Gln Leu Leu Pro Leu Ser Gln Ile Gln Ala Ile

450 455 460

Leu Ala Leu Ala Leu Ala Gln Ala Met Asp Leu Leu Asp Asp Pro Glu

465 470 475 480

Gly Gln Ala Gly Trp Ser Leu Thr Ala Arg Asp Leu Arg Asp Arg Ile

485 490 495

Arg Ala Val Ser Pro Gly Leu Arg Ala Asp Arg Pro Leu Ala Gly His

500 505 510

Ile Glu Ala Val Ala Gln Gly Leu Arg His Pro Ser Ala Ala Ala Asp

515 520 525

Pro Pro Ala

530

<210> 108

<211> 1560

<212> DNA

<213> Escherichia coli

<400> 108

atgaaacctg aagattttcg tgcttccacc cagcggccct ttactggaga agaatatctg 60

aagtctcttc aagacggccg tgagatctat atttacggcg agcgtgtaaa ggatgttaca 120

acgcaccctg cttttcgtaa tgctgctgcc tcagttgccc aactttatga cgccctgcat 180

aaacccgaaa tgcaggactc actttgttgg aatacggaca cggggtccgg tggttacaca 240

cacaagttct ttcgtgtcgc caaatctgcc gatgacttgc gtcaacaacg tgatgctatc 300

gctgaatggt cacgtctgag ttacggctgg atgggccgta cgcccgacta taaggccgcc 360

tttggttgcg cattgggtgc aaatcccggc ttttatggac agtttgaaca aaatgcacgt 420

aactggtata ctcgtatcca ggaaaccggc ttgtacttta atcatgctat tgttaaccca 480

cctatcgatc gtcatttgcc aaccgataag gtcaaggatg tatatattaa acttgaaaag 540

gagacggacg ctggtatcat tgtgtctggc gctaaagtgg tggctacgaa cagtgcattg 600

acgcattaca atatgattgg ttttggttcc gcccaagtga tgggcgagaa ccccgacttt 660

gccttgatgt ttgttgcccc tatggacgct gacggagtaa aacttatttc acgtgcctca 720

tatgaaatgg tagctggtgc cactggttct ccttatgatt atcctctttc atcccgtttt 780

gacgaaaacg acgccatcct ggtcatggac aatgttctta ttccctggga gaacgtgctg 840

atctaccgtg acttcgatcg ttgccgtcgt tggactatgg aaggaggttt cgctcgtatg 900

tatccactgc aggcctgtgt tcgtttggct gtgaagttgg actttatcac agcccttctt 960

aaaaagtcac ttgagtgtac tggaacgttg gagttccgtg gtgtacaggc agatctgggc 1020

gaagtagtag cctggcgtaa tactttctgg gcactgtctg actctatgtg ctccgaggca 1080

acaccatggg tgaacggcgc atacctgccc gatcacgccg ctcttcaaac atatcgtgtg 1140

cttgctccca tggcttatgc taaaatcaaa aatattatcg agcgtaatgt aacttctggt 1200

ctgatttacc tgccatccag tgctcgtgac cttaacaatc ctcaaatcga tcagtatctt 1260

gcaaaatatg tgcgtggctc aaacggaatg gaccatgtgc agcgtatcaa aatcctgaag 1320

ttgatgtggg atgccatcgg ttctgaattt ggcggacgtc atgaactgta tgagattaac 1380

tactctggat ctcaagacga aatcagactt cagtgtttgc gtcaggccca aaattctgga 1440

aatatggata aaatgatggc tatggtagac cgttgtctga gtgagtacga tcaagatggt 1500

tggacagttc ctcatcttca taataatgac gatattaata tgcttgataa gttgcttaaa 1560

<210> 109

<211> 520

<212>PRT

<213> Escherichia coli

<400> 109

Met Lys Pro Glu Asp Phe Arg Ala Ser Thr Gln Arg Pro Phe Thr Gly

1 5 10 15

Glu Glu Tyr Leu Lys Ser Leu Gln Asp Gly Arg Glu Ile Tyr Ile Tyr

20 25 30

Gly Glu Arg Val Lys Asp Val Thr Thr His Pro Ala Phe Arg Asn Ala

35 40 45

Ala Ala Ser Val Ala Gln Leu Tyr Asp Ala Leu His Lys Pro Glu Met

50 55 60

Gln Asp Ser Leu Cys Trp Asn Thr Asp Thr Gly Ser Gly Gly Tyr Thr

65 70 75 80

His Lys Phe Phe Arg Val Ala Lys Ser Ala Asp Asp Leu Arg Gln Gln

85 90 95

Arg Asp Ala Ile Ala Glu Trp Ser Arg Leu Ser Tyr Gly Trp Met Gly

100 105 110

Arg Thr Pro Asp Tyr Lys Ala Ala Phe Gly Cys Ala Leu Gly Ala Asn

115 120 125

Pro Gly Phe Tyr Gly Gln Phe Glu Gln Asn Ala Arg Asn Trp Tyr Thr

130 135 140

Arg Ile Gln Glu Thr Gly Leu Tyr Phe Asn His Ala Ile Val Asn Pro

145 150 155 160

Pro Ile Asp Arg His Leu Pro Thr Asp Lys Val Lys Asp Val Tyr Ile

165 170 175

Lys Leu Glu Lys Glu Thr Asp Ala Gly Ile Ile Val Ser Gly Ala Lys

180 185 190

Val Val Ala Thr Asn Ser Ala Leu Thr His Tyr Asn Met Ile Gly Phe

195 200 205

Gly Ser Ala Gln Val Met Gly Glu Asn Pro Asp Phe Ala Leu Met Phe

210 215 220

Val Ala Pro Met Asp Ala Asp Gly Val Lys Leu Ile Ser Arg Ala Ser

225 230 235 240

Tyr Glu Met Val Ala Gly Ala Thr Gly Ser Pro Tyr Asp Tyr Pro Leu

245 250 255

Ser Ser Arg Phe Asp Glu Asn Asp Ala Ile Leu Val Met Asp Asn Val

260 265 270

Leu Ile Pro Trp Glu Asn Val Leu Ile Tyr Arg Asp Phe Asp Arg Cys

275 280 285

Arg Arg Trp Thr Met Glu Gly Gly Phe Ala Arg Met Tyr Pro Leu Gln

290 295 300

Ala Cys Val Arg Leu Ala Val Lys Leu Asp Phe Ile Thr Ala Leu Leu

305 310 315 320

Lys Lys Ser Leu Glu Cys Thr Gly Thr Leu Glu Phe Arg Gly Val Gln

325 330 335

Ala Asp Leu Gly Glu Val Val Ala Trp Arg Asn Thr Phe Trp Ala Leu

340 345 350

Ser Asp Ser Met Cys Ser Glu Ala Thr Pro Trp Val Asn Gly Ala Tyr

355 360 365

Leu Pro Asp His Ala Ala Leu Gln Thr Tyr Arg Val Leu Ala Pro Met

370 375 380

Ala Tyr Ala Lys Ile Lys Asn Ile Ile Glu Arg Asn Val Thr Ser Gly

385 390 395 400

Leu Ile Tyr Leu Pro Ser Ser Ala Arg Asp Leu Asn Asn Pro Gln Ile

405 410 415

Asp Gln Tyr Leu Ala Lys Tyr Val Arg Gly Ser Asn Gly Met Asp His

420 425 430

Val Gln Arg Ile Lys Ile Leu Lys Leu Met Trp Asp Ala Ile Gly Ser

435 440 445

Glu Phe Gly Gly Arg His Glu Leu Tyr Glu Ile Asn Tyr Ser Gly Ser

450 455 460

Gln Asp Glu Ile Arg Leu Gln Cys Leu Arg Gln Ala Gln Asn Ser Gly

465 470 475 480

Asn Met Asp Lys Met Met Ala Met Val Asp Arg Cys Leu Ser Glu Tyr

485 490 495

Asp Gln Asp Gly Trp Thr Val Pro His Leu His Asn Asn Asp Asp Ile

500 505 510

Asn Met Leu Asp Lys Leu Leu Lys

515 520

<210> 110

<211> 510

<212> DNA

<213> Escherichia coli

<400> 110

atgcagcttg atgagcagcg tttgcgtttt cgtgacgcta tggcatcact tagtgccgct 60

gtgaacatca tcacgactga gggagacgcc ggtcagtgcg gaatcacagc aaccgcagta 120

tgtagtgtta cggacactcc cccatctctg atggtctgta tcaatgccaa tagtgctatg 180

aatccagtct ttcagggtaa tggtaaactt tgtgtcaacg tcctgaatca cgagcaagag 240

ttgatggctc gtcattttgc aggaatgact ggtatggcta tggaagagcg tttttccttg 300

tcttgttggc aaaaaggacc actggcccag cctgtattga aaggttcttt ggcttccctg 360

gagggtgaga tccgtgacgt tcaagccatt ggaactcatc tggtatattt ggtagaaatc 420

aagaatatta tcctgtcagc cgagggccat ggtctgatct actttaaacg tcgttttcac 480

cccgtgatgc tggagatgga ggcagccatt 510

<210> 111

<211> 170

<212>PRT

<213> Escherichia coli

<400> 111

Met Gln Leu Asp Glu Gln Arg Leu Arg Phe Arg Asp Ala Met Ala Ser

1 5 10 15

Leu Ser Ala Ala Val Asn Ile Ile Thr Thr Glu Gly Asp Ala Gly Gln

20 25 30

Cys Gly Ile Thr Ala Thr Ala Val Cys Ser Val Thr Asp Thr Pro Pro

35 40 45

Ser Leu Met Val Cys Ile Asn Ala Asn Ser Ala Met Asn Pro Val Phe

50 55 60

Gln Gly Asn Gly Lys Leu Cys Val Asn Val Leu Asn His Glu Gln Glu

65 70 75 80

Leu Met Ala Arg His Phe Ala Gly Met Thr Gly Met Ala Met Glu Glu

85 90 95

Arg Phe Ser Leu Ser Cys Trp Gln Lys Gly Pro Leu Ala Gln Pro Val

100 105 110

Leu Lys Gly Ser Leu Ala Ser Leu Glu Gly Glu Ile Arg Asp Val Gln

115 120 125

Ala Ile Gly Thr His Leu Val Tyr Leu Val Glu Ile Lys Asn Ile Ile

130 135 140

Leu Ser Ala Glu Gly His Gly Leu Ile Tyr Phe Lys Arg Arg Phe His

145 150 155 160

Pro Val Met Leu Glu Met Glu Ala Ala Ile

165 170

<210> 112

<211> 801

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleic acid optimized for expression in plants,

encoding protein SEQ ID No:80

<400> 112

atgaggataa atatctcttt gtctagtctc tttgagagac tgagcaaatt gtcatcccga 60

tccatcgcaa ttacctgcgg cgtagtcttg gcaagtgcaa tagcattccc aattatccga 120

agagactatc aaacgttcct tgaagtcggt ccgagctatg ccccacagaa cttccgaggc 180

tatatcatcg tttgtgtctt gtcacttttt aggcaagagc agaaagggtt ggcaatctat 240

gacaggcttc cagaaaagag gcgttggctt gccgacttgc cgtttcgtga agggacgagg 300

ccatccataa cctcccacat tatacagcgt cagcgtactc agctggtaga tcaagagttt 360

gcaaccagag aacttatcga taaggtgatc ccacgtgtgc aagcaagaca cacagacaaa 420

acttttctca gtacgtcaaa atttgagttt catgcaaagg ccatcttcct tctcccctct 480

atccctatta atgatcctct taacattccc tcacacgata cggtaagaag aaccaagcgt 540

gagatcgctc acatgcatga ttaccatgat tgcactctgc acttggctct tgctgctcag 600

gatggtaagg aagttttgaa gaaggggtgg ggccagcgtc acccactggc cggaccagga 660

gtcccaggcc ctcctactga gtggaccttc ctttacgcac caaggaacga agaggaggcc 720

agagttgtcg agatgatagt cgaagctagt attggctaca tgactaacga tcctgctggt 780

aagattgtcg aaaatgccaa g 801

<210> 113

<211> 5034

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleic acid optimized for expression in plants,

encoding protein SEQ ID No:35

<400> 113

atgaattcca gcaagaatcc tccttccact cttcttgatg tttttctgga tactgccagg 60

aaccttgata ccgctttgag gaatgtcttg gaatgcggcg aacacagatg gtcctacaga 120

gagcttgata ctgtttcatc tgctctcgcc cagcatctta ggtacactgt cggtcttagt 180

cctactgtcg ccgtcatcag tgaaaaccat ccttatattc tcgctttgat gctggctgta 240

tggaaacttg gaggcacctt cgctcctatt gatgtccatt ctcctgccga attggtagct 300

ggcatgctga acatagtctc tccttcttgc ttggttattc cgagctcaga tgtaactaat 360

caaactcttg cctgcgatct taatatcccc gtcgttgcat ttcacccaca tcaatccact 420

attcctgagc tgaacaagaa gtacctcacc gattctcaaa tttctccgga tcttcctttt 480

tcagatccaa acagacctgc tctgtacctc ttcacttcat ccgccacttc tcgaagtaat 540

ctcaaatgcg tgcctctcac tcacaccttt atcctccgaa acagcctctc taagcgtgca 600

tggtgcaagc gtatgcgtcc agagacagac tttgacggca tacgtgttct tggatgggcc 660

ccgtggtctc acgtcctcgc acacatgcaa gacatcggac cactcaccct gcttaatgcc 720

ggatgctacg tttttgcaac tactccatcc acgtacccta cggaattgaa ggacgacagg 780

gacctgatat cttgcgccgc aaatgctatc atgtacaagg gcgtcaagtc atttgcttgt 840

cttccctttg tactcggagg gctgaaggca ctctgcgagt ctgagccatc cgtgaaggcc 900

catcttcagg tcgaggagag agctcaactc ctgaagtctc tgcaacacat ggaaattctt 960

gagtgtggag gtgccatgct cgaagcaagt gttgcctctt gggctattga gaactgcatt 1020

cccatttcta tcggtattgg tatgacggag actggtggag ccctctttgc aggccccgtt 1080

caggccatca aaaccgggtt ttcttcagag gataaattca ttgaagatgc tacttacttg 1140

ctcgttaagg atgatcatga gagtcatgct gaggaggata ttaacgagg tgaactcgtt 1200

gtgaaaagta aaatgctccc acgaggctac cttggctata gtgatccttc cttctcagtc 1260

gacgatgctg gctgggttac atttagaaca ggagacagat acagcgttac acctgacgga 1320

aagttttcct ggctgggccg aaacactgat ttcattcaga tgaccagtgg tgagacgctg 1380

gatccccgac caattgagag cagtctctgc gaaagttctc ttatttctag agcatgcgtt 1440

atcggagata aatttctcaa cgggcctgct gctgctgttt gtgctatcat tgagcttgag 1500

cccacagctg tggaaaaagg acaagctcac agccgtgaga tagcaagagt tttcgcacct 1560

attaatcgag acttgccgcc tcctcttagg attgcatgga gtcacgtttt ggttctccag 1620

cccagtgaga agataccgat gacgaagaag ggtaccatct tcaggaagaa aattgagcag 1680

gtgtttggct ctgccttggg tggcagctct ggagataact ctcaagccac tgctgatgct 1740

ggcgttgttc gacgagacga gctttcaaac actgtcaagc acataattag ccgtgttctc 1800

ggagtttccg atgacgaact cctttggacg ttgtcatttg ccgagctcgg aatgacgtca 1860

gcattggcca ctcgaatcgc caacgagttg aacgaagttt tggttggagt taatctccct 1920

atcaacgctt gctatataca tgtcgacctt ccttctctga gcaatgccgt ctatgccaaa 1980

cttgcacacc tcaagctgcc agatcgtact cccgaaccca ggcaagcccc tgtcgaaaac 2040

tctggtggga aggagatcgt tgtcgttggc caggcctttc gtcttcctgg ctcaataaac 2100

gatgtcgcct ctcttcgaga cgcattcctg gccagacaag catcatccat tatcactgaa 2160

ataccatccg ataggtggga ccacgccagc ttctatccca aggatatacg tttcaacaag 2220

gctggccttg tggatatagc caattatgat catagctttt tcggactgac ggcaaccgaa 2280

gcactctatc tgtccccaac tatgcgtttg gcactcgaag tttcctttga agccttggag 2340

aatgctaata tcccggtgtc acaactcaag ggttctcaaa cagctgttta tgttgctact 2400

acagatgacg gatttgagac ccttttgaat gccgaggccg gctatgatgc ttatacaaga 2460

ttctatggca ctggtcgagc agcaagtaca gccagcgggc gtataagctg tcttcttgat 2520

gtccatggac cctctattac tgttgatacg gcatgcagtg gaggggctgt ttgtattgac 2580

caagcaatcg actatctgca atcaagcagt gcagcagaca ccgctatcat atgtgctagt 2640

aacacgcact gctggccagg cagcttcagg tttctttccg cacaagggat ggtatcccca 2700

ggaggacgat gcgcaacatt tacaactgat gctgatggct acgtgccctc tgaggcgct 2760

gtcgccttca tattgaaaac ccgagaagca gctatgcgtg acaaggacac tatcctcgcc 2820

acaatcaaag ctacacagat atcacacaat ggccgatctc aaggtcttgt ggcaccgaat 2880

gtcaacagcc aagctgacct tcatagaagc ttgcttcaaa aagctggcct tagcccggct 2940

gatatccgtt tcattgaagc tcatgggaca ggaacgtcac tgggagacct ctcagaaatt 3000

caagctataa atgatgctta tacctcctct cagccgcgta cgaccggccc actcatagtc 3060

agcgcttcca aaacggtcat tggtcatacc gaaccagctg gccccttggt cggtatgctg 3120

agtgtcttga actctttcaa agaaggcgcc gtccctggtc tcgcccatct taccgcagac 3180

aatttgaatc ccagtctgga ctgttcttct gtgccacttc tcattcccta tcaacctgtt 3240

cacctggctg cacccaagcc tcaccgagct gctgtaaggt catacggctt ttcaggtacc 3300

ctgggcggca tcgttctcga ggctcctgac gaagaaagac tggaagaaga gctgccaaat 3360

gacaagccca tgttgttcgt cgtcagcgca aagacacata cagcacttat cgaatacctg 3420

gggcgttatc tcgagttcct cttgcaggca aacccccaag atttttgtga catttgttat 3480

acaagctgcg ttgggcgtga gcactataga tatcgatatg cttgtgtagc aaatgatatg 3540

gaggacctca taggccaact ccagaaacgt ttgggcagca aggtgccgcc aaagccgtca 3600

tacaaacgtg gtgctttggc ctttgccttt tctggtcagg gtacacaatt ccgagggatg 3660

gccacagagc ttgcaaaagc atactccggc ttccgaaaga tcgtgtccga tctcgcaaag 3720

agagctagcg agttgtcagg tcatgccatt gaccgttttc ttcttgcata tgacataggc 3780

gctgaaaatg tagctcctga tagtgaggca gaccagattt gcatctttgt gtatcagtgt 3840

tctgtccttc gttggctgca gactatgggg attagaccca gtgcagtgat aggccatagc 3900

ctcggggaga tctcagcttc tgtggcagca ggagcacttt ctcttgactc cgctttggat 3960

cttgtcatct cacgagctcg tcttttgagg tcttcagcaa gtgctcctgc aggaatggca 4020

gctatgtctg cctctcaaga cgaggttgtg gagttgattg ggaaactcga cctcgacaag 4080

gctaattctc tcagcgtttc agtcataaat ggtccccaaa atactgtcgt gtccggctct 4140

tcagctgcta ttgaaagcat agtggctctt gccaaaggga gaaagatcaa agcctctgcc 4200

ctgaatatca atcaagcttt tcatagtcca tacgtcgaca gtgccgtccc tggtctccgt 4260

gcttggtcag aaaagcatat ctcctcagct agaccattgc aaattccgct gtattcaacg 4320

ttgttgggag cacaaatctc tgagggagag atgttgaatc cagatcactg ggtcgaccat 4380

gcacgaaagc ctgtacagtt cgcacaagca gccacaacca tgaaagaatc cttcaccgga 4440

gtcatcatag atatcggccc tcaagtagtg gcttggtcac ttctgctcag taacgggctc 4500

acgtccgtga ctgctctcgc tgcaaaaaga gggagaagtc aacaggtggc tttcctcagc 4560

gccttggccg atttgtatca agattacggt gttgttcctg attttgtcgg gctttatgct 4620

cagcaggaag atgcttcaag gttgaagaag acggatatct tgacgtatcc gttccagcgt 4680

ggcgaagaga ctctttctag tggttctagc actccgacat tggaaaacac ggatttggat 4740

tccggtaagg aactccttat gggaccgact agaggttgc tccgtgctga cgacttgcgt 4800

gacagtatcg tttcttctgt gaaggatgtt ctggaactca agtcaaatga agacctcgat 4860

ttgtctgaaa gtctgaatgc acttggtatg gacagcatca tgttcgctca gctccgaaag 4920

cgtattgggg aaggactcgg attgaatgtt ccgatggttt ttctgagcga cgccttttct 4980

attggtgaga tggttagtaa tcttgtggaa caggcagagg cttctgagga caat 5034

<210> 114

<211> 867

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleic acid optimized for expression in plants,

encoding protein SEQ ID No:65

<400> 114

atggctccaa tttcttcaac ttggtctcgt ctcattcgat ttgtggctat tgaaacgtcc 60

ctcgtgcata tcggtgaacc gatacgcc accatggacg tcggtctggc cagacgagaa 120

ggcaagacga tccaagcata cgagattatt ggatcaggca gtgctcttga cctctcagcc 180

caagtaagta agaatgtgct gactgtaagg gaactcctga tgccgctttc aagagaggaa 240

attaaaactg tacgatgctt ggggttgaac taccctgttc atgccaccga agcaaacgtt 300

gctgttccaa aattcccgaa tttgttctac aaaccagtga cctccctcat tggccccgat 360

ggactcatta ccatcccttc cgttgtccaa cccccgaagg agcatcagtc cgattatgaa 420

gcagaacttg tcattgtcat cgggaaagca gcaaagaatg taagtgagga tgaggctttg 480

gattatgtat tgggatacac tgccgctaac gatatttcct ttaggaaaca ccagcttgca 540

gtctcacaat ggtctttctc taaaggattt ggtagccttt tgctcactat ccgtatggca 600

caaacccact ctggtaacat taatagattc tccagagacc agattttcaa tgtcaagaag 660

acaatttcct tcctgtcaca aggcactaca ctggaaccag gttctatcat tttgactggt 720

acacctgacg gagtgggctt tgtgcgaaat ccaccacttt accttaaaga tggagatgaa 780

gtaatgacct ggattggaag tggaatcgga acattggcca atacagtgca agaagagaag 840

acttgcttcg ctagtggcgg acacgag 867

<210> 115

<211> 9

<212>PRT

<213> Artificial sequence

<220>

<223> Amino acid linker sequence

<400> 115

Gly Gly Ser Gly Gly Ser Gly Gly Ser

15

<210> 116

<211> 1569

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleic acid optimized for expression in plants,

encoding phenylalanine ammonium lyase from Streptomyces maritimus (SEQ

ID No: 117)

<400> 116

atgacattcg tgatagagct ggacatgaat gtcactcttg atcagttaga ggatgctgca 60

agacaaagaa cacccgtgga attgtctgca cctgtacgtt caagggtccg tgcttcacgt 120

gatgttttag ttaaattcgt acaagatgag cgtgtgattt acggtgtaaa tacttcaatg 180

ggcggattcg tggatcacct agtacctgtg agtcaggccc gtcaattgca agagaattta 240

atcaatgcag ttgccacaaa tgttggagca tatttagatg atacaacggc caggacgatt 300

atgttgagta ggattgtatc cctggcaagg ggcaattctg ccataactcc cgccaacctt 360

gacaaattag tggcagtgtt gaacgcagga atcgtcccat gcatccctga aaaaggttca 420

ttaggtactt caggtgactt aggccctctg gcagctatcg ccttagtatg cgctggccag 480

tggaaggcac gttacaatgg acaaataatg ccaggccgtc aggctctatc agaggccggt 540

gtcgaaccta tggaactttc atataaggac ggcctagcac ttataaacgg tacgtctgga 600

atggtaggcc ttggaactat ggttttacaa gccgctagga ggttagtgga caggtatttg 660

caagtaagtg ccttatctgt agaaggcttg gcaggcatga ctaaaccttt tgaccctcgt 720

gtacacggag ttaagcccca tagaggtcaa agacaagtag cttcccgtct atgggaagga 780

ttggccgatt ctcaccttgc cgtgaacgag cttgatactg aacaaaccct tgcaggtgaa 840

atgggcacgg tggcaaaggc tggaagttta gccatagagg atgcctactc cattcgttgc 900

acccctcaga tcctgggtcc agtcgtagac gtattggata ggatcggtgc aacattacaa 960

gacgagctga attcttcaaa cgataacccc attgtccttc ctgaggaagc agaagtattc 1020

cataatggac actttcatgg acagtatgtc gccatggcaa tggatcacct aaatatggct 1080

cttgctactg ttacgaatct tgccaataga agagtcgaca gatttctaga taaatcaaat 1140

tccaatggac taccagcctt tctttgccgt gaagacccag gccttaggct tggtttgatg 1200

ggcggccagt tcatgactgc atcaatcacc gctgaaaccc gtacactgac aatcccaatg 1260

tctgtacaat cacttacttc cacggcagat tttcaagaca tagtatcctt tggcttcgta 1320

gccgcaagga gagctagaga ggtgttgaca aacgctgctt atgtggtagc ctttgaacta 1380

ctgtgtgcct gtcaggctgt ggacatcaga ggagcagaca agctatcttc tttcacccgt 1440

cctctatatg aaagaaccag gaaaattgtt ccctttttcg acagagatga aacaattact 1500

gactacgttg agaagttagc tgctgattta atagcaggcg agcctgtaga tgccgctgtg 1560

gctgcacat 1569

<210> 117

<211> 523

<212>PRT

<213> Streptomyces maritimus

<400> 117

Met Thr Phe Val Ile Glu Leu Asp Met Asn Val Thr Leu Asp Gln Leu

1 5 10 15

Glu Asp Ala Ala Arg Gln Arg Thr Pro Val Glu Leu Ser Ala Pro Val

20 25 30

Arg Ser Arg Val Arg Ala Ser Arg Asp Val Leu Val Lys Phe Val Gln

35 40 45

Asp Glu Arg Val Ile Tyr Gly Val Asn Thr Ser Met Gly Gly Phe Val

50 55 60

Asp His Leu Val Pro Val Ser Gln Ala Arg Gln Leu Gln Glu Asn Leu

65 70 75 80

Ile Asn Ala Val Ala Thr Asn Val Gly Ala Tyr Leu Asp Asp Thr Thr

85 90 95

Ala Arg Thr Ile Met Leu Ser Arg Ile Val Ser Leu Ala Arg Gly Asn

100 105 110

Ser Ala Ile Thr Pro Ala Asn Leu Asp Lys Leu Val Ala Val Leu Asn

115 120 125

Ala Gly Ile Val Pro Cys Ile Pro Glu Lys Gly Ser Leu Gly Thr Ser

130 135 140

Gly Asp Leu Gly Pro Leu Ala Ala Ile Ala Leu Val Cys Ala Gly Gln

145 150 155 160

Trp Lys Ala Arg Tyr Asn Gly Gln Ile Met Pro Gly Arg Gln Ala Leu

165 170 175

Ser Glu Ala Gly Val Glu Pro Met Glu Leu Ser Tyr Lys Asp Gly Leu

180 185 190

Ala Leu Ile Asn Gly Thr Ser Gly Met Val Gly Leu Gly Thr Met Val

195 200 205

Leu Gln Ala Ala Arg Arg Leu Val Asp Arg Tyr Leu Gln Val Ser Ala

210 215 220

Leu Ser Val Glu Gly Leu Ala Gly Met Thr Lys Pro Phe Asp Pro Arg

225 230 235 240

Val His Gly Val Lys Pro His Arg Gly Gln Arg Gln Val Ala Ser Arg

245 250 255

Leu Trp Glu Gly Leu Ala Asp Ser His Leu Ala Val Asn Glu Leu Asp

260 265 270

Thr Glu Gln Thr Leu Ala Gly Glu Met Gly Thr Val Ala Lys Ala Gly

275 280 285

Ser Leu Ala Ile Glu Asp Ala Tyr Ser Ile Arg Cys Thr Pro Gln Ile

290 295 300

Leu Gly Pro Val Val Asp Val Leu Asp Arg Ile Gly Ala Thr Leu Gln

305 310 315 320

Asp Glu Leu Asn Ser Ser Asn Asp Asn Pro Ile Val Leu Pro Glu Glu

325 330 335

Ala Glu Val Phe His Asn Gly His Phe His Gly Gln Tyr Val Ala Met

340 345 350

Ala Met Asp His Leu Asn Met Ala Leu Ala Thr Val Thr Asn Leu Ala

355 360 365

Asn Arg Arg Val Asp Arg Phe Leu Asp Lys Ser Asn Ser Asn Gly Leu

370 375 380

Pro Ala Phe Leu Cys Arg Glu Asp Pro Gly Leu Arg Leu Gly Leu Met

385 390 395 400

Gly Gly Gln Phe Met Thr Ala Ser Ile Thr Ala Glu Thr Arg Thr Leu

405 410 415

Thr Ile Pro Met Ser Val Gln Ser Leu Thr Ser Thr Ala Asp Phe Gln

420 425 430

Asp Ile Val Ser Phe Gly Phe Val Ala Ala Arg Arg Ala Arg Glu Val

435 440 445

Leu Thr Asn Ala Ala Tyr Val Val Ala Phe Glu Leu Leu Cys Ala Cys

450 455 460

Gln Ala Val Asp Ile Arg Gly Ala Asp Lys Leu Ser Ser Phe Thr Arg

465 470 475 480

Pro Leu Tyr Glu Arg Thr Arg Lys Ile Val Pro Phe Phe Asp Arg Asp

485 490 495

Glu Thr Ile Thr Asp Tyr Val Glu Lys Leu Ala Ala Asp Leu Ile Ala

500 505 510

Gly Glu Pro Val Asp Ala Ala Val Ala Ala His

515 520

<210> 118

<211> 1173

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Aquilaria sinensis,

optimized for expression in Nicotiana benthamiana

<400> 118

atgggatctc aagatgttgc tggaggagct cttaagggag ttaatccagg aaaggctact 60

attcttgctc ttggaaaggc ttttccatat caacttgtta tgcaagaatt tcttgttgat 120

ggatatttta agaatacttc ttgtaaggat caagaactta agcaaaagct tgctagactt 180

tgtaagacta ctactgttaa gactagatat gttgttatgt ctgaagaaat tcttaataag 240

tatccagaac ttgctgttga aggaattcca actcttaagc aaagacttga tattggaaat 300

gaagctctta ctgaaatggc tattgaagct tctcaagctt gtattaagaa gtggggaaga 360

ccagcttctg aaattactca tcttgtttat gtttcttctt ctgaagctag acttccagga 420

ggagatcttt atcttgctca aggacttgga ctttctccaa gaactaagag agttgttctt 480

tattttatgg gatgttctgg aggagttgct ggacttagag ttgctaagga tattgctgaa 540

aataatccag gatctagagt tcttcttgct acttctgaaa ctactattgt tggatttaag 600

cccacatctg ctcatagacc atatgatctt gttggagttg ctctttttgg agatggagct 660

ggagctatgg ttattggatc tgatccactt ccadgaactg aatctccact ttttgaactt 720

catactgcta ttcaaaattt tcttccaaat actgaaaaga ctattgatgg aagacttact 780

gaagaaggaa tttcttttaa gcttgctaga gaacttccac aaattgttga agatcatatt 840

gaaggatttt gtggacaact tactggagtt attggacttt ctcataagca atataataag 900

atgttttggg ctgttcatcc aggaggacca gctattctta atagagttga aaagagactt 960

gatcttcatc caaataagct tgatgcttct agaagagctc ttgaagatta tggaaatgct 1020

tcttctaatt ctattgttta tgttcttgat tatatgattg aagaaactct taagatgaag 1080

actgaatctc ttgaaccatc tgaatgggga cttattcttg cttttggacc aggagttact 1140

tttgaaggaa ttcttgctag aaatcttgct gtt 1173

<210> 119

<211> 391

<212>PRT

<213> Aquilaria sinensis

<400> 119

Met Gly Ser Gln Asp Val Ala Gly Gly Ala Leu Lys Gly Val Asn Pro

1 5 10 15

Gly Lys Ala Thr Ile Leu Ala Leu Gly Lys Ala Phe Pro Tyr Gln Leu

20 25 30

Val Met Gln Glu Phe Leu Val Asp Gly Tyr Phe Lys Asn Thr Ser Cys

35 40 45

Lys Asp Gln Glu Leu Lys Gln Lys Leu Ala Arg Leu Cys Lys Thr Thr

50 55 60

Thr Val Lys Thr Arg Tyr Val Val Met Ser Glu Glu Ile Leu Asn Lys

65 70 75 80

Tyr Pro Glu Leu Ala Val Glu Gly Ile Pro Thr Leu Lys Gln Arg Leu

85 90 95

Asp Ile Gly Asn Glu Ala Leu Thr Glu Met Ala Ile Glu Ala Ser Gln

100 105 110

Ala Cys Ile Lys Lys Trp Gly Arg Pro Ala Ser Glu Ile Thr His Leu

115 120 125

Val Tyr Val Ser Ser Ser Glu Ala Arg Leu Pro Gly Gly Asp Leu Tyr

130 135 140

Leu Ala Gln Gly Leu Gly Leu Ser Pro Arg Thr Lys Arg Val Val Leu

145 150 155 160

Tyr Phe Met Gly Cys Ser Gly Gly Val Ala Gly Leu Arg Val Ala Lys

165 170 175

Asp Ile Ala Glu Asn Asn Pro Gly Ser Arg Val Leu Leu Ala Thr Ser

180 185 190

Glu Thr Thr Ile Val Gly Phe Lys Pro Pro Ser Ala His Arg Pro Tyr

195 200 205

Asp Leu Val Gly Val Ala Leu Phe Gly Asp Gly Ala Gly Ala Met Val

210 215 220

Ile Gly Ser Asp Pro Leu Pro Gly Thr Glu Ser Pro Leu Phe Glu Leu

225 230 235 240

His Thr Ala Ile Gln Asn Phe Leu Pro Asn Thr Glu Lys Thr Ile Asp

245 250 255

Gly Arg Leu Thr Glu Glu Gly Ile Ser Phe Lys Leu Ala Arg Glu Leu

260 265 270

Pro Gln Ile Val Glu Asp His Ile Glu Gly Phe Cys Gly Gln Leu Thr

275 280 285

Gly Val Ile Gly Leu Ser His Lys Gln Tyr Asn Lys Met Phe Trp Ala

290 295 300

Val His Pro Gly Gly Pro Ala Ile Leu Asn Arg Val Glu Lys Arg Leu

305 310 315 320

Asp Leu His Pro Asn Lys Leu Asp Ala Ser Arg Arg Ala Leu Glu Asp

325 330 335

Tyr Gly Asn Ala Ser Ser Asn Ser Ile Val Tyr Val Leu Asp Tyr Met

340 345 350

Ile Glu Glu Thr Leu Lys Met Lys Thr Glu Ser Leu Glu Pro Ser Glu

355 360 365

Trp Gly Leu Ile Leu Ala Phe Gly Pro Gly Val Thr Phe Glu Gly Ile

370 375 380

Leu Ala Arg Asn Leu Ala Val

385 390

<210> 120

<211> 1164

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS2 from Aquilaria sinensis,

optimized for expression in Nicotiana benthamiana

<400> 120

atgtctcaag ctattgctga taatgcttat agacatcatc ttaagagagc tccaactcca 60

ggaaaggcta ctgttcttgc tcttggaaag gcttttccaa agcaagttat tccacaagaa 120

aatcttgttg aaggatatat tagagatact aagtgtgaag atgtttctat taaggaaaag 180

cttgaaagac tttgtaagac tactactgtt aagactagat atactgttat gtctaaggaa 240

attcttgata attatccaga acttgttact gaaggatctc caactattag acaaagactt 300

gaaattgcta atccagctgt tgttgaaatg gctaaggaag cttctcttgc ttgtattaag 360

caatggggaa gaccagctgg agatattact catattgttt atgtttcttc ttctgaaatt 420

agacttccag gaggagatct ttatcttgct aatgaacttg gacttaagaa tgatattaat 480

agaattatgc tttattttct tggatgttat ggaggagtta ctggacttag agttgctaag 540

gatattgctg aaaataatcc aggatctaga attcttctta ctacttctga aactactatt 600

cttggattta gaccaccaaa taagtctaga ccatatgatc ttgttggagc tgctcttttt 660

ggagatggag ctgctgctgt tattattgga gctaatccag aaattggaag agaatctcca 720

tttatggaac ttaattttgc tcttcaacaa tttcttccag gaactcatgg agttattgat 780

ggaagacttt ctgaagaagg aattaatttt aagcttggaa gagatcttcc acaaaagatt 840

gaagataata ttgaagattt ttgtagaaag cttatgatta aggctgatgg agatcttaag 900

gaatttaatg aacttttttg ggctgttcat cccaggagac cagctattct taatagactt 960

gaatctattc ttgatcttaa gaatggaaag cttgaatgtt ctagaagagc tcttatggat 1020

tatggaaatg tttcttctaa tactattttt tatgttatgg aatatatgag agaagaactt 1080

aagagagaag gatctgaaga atggggactt gctcttgctt ttggaccagg aattactttt 1140

gaaggaattc ttcttagatc tctt 1164

<210> 121

<211> 388

<212>PRT

<213> Aquilaria sinensis

<400> 121

Met Ser Gln Ala Ile Ala Asp Asn Ala Tyr Arg His His Leu Lys Arg

1 5 10 15

Ala Pro Thr Pro Gly Lys Ala Thr Val Leu Ala Leu Gly Lys Ala Phe

20 25 30

Pro Lys Gln Val Ile Pro Gln Glu Asn Leu Val Glu Gly Tyr Ile Arg

35 40 45

Asp Thr Lys Cys Glu Asp Val Ser Ile Lys Glu Lys Leu Glu Arg Leu

50 55 60

Cys Lys Thr Thr Thr Val Lys Thr Arg Tyr Thr Val Met Ser Lys Glu

65 70 75 80

Ile Leu Asp Asn Tyr Pro Glu Leu Val Thr Glu Gly Ser Pro Thr Ile

85 90 95

Arg Gln Arg Leu Glu Ile Ala Asn Pro Ala Val Val Glu Met Ala Lys

100 105 110

Glu Ala Ser Leu Ala Cys Ile Lys Gln Trp Gly Arg Pro Ala Gly Asp

115 120 125

Ile Thr His Ile Val Tyr Val Ser Ser Ser Glu Ile Arg Leu Pro Gly

130 135 140

Gly Asp Leu Tyr Leu Ala Asn Glu Leu Gly Leu Lys Asn Asp Ile Asn

145 150 155 160

Arg Ile Met Leu Tyr Phe Leu Gly Cys Tyr Gly Gly Val Thr Gly Leu

165 170 175

Arg Val Ala Lys Asp Ile Ala Glu Asn Asn Pro Gly Ser Arg Ile Leu

180 185 190

Leu Thr Thr Ser Glu Thr Thr Ile Leu Gly Phe Arg Pro Pro Asn Lys

195 200 205

Ser Arg Pro Tyr Asp Leu Val Gly Ala Ala Leu Phe Gly Asp Gly Ala

210 215 220

Ala Ala Val Ile Ile Gly Ala Asn Pro Glu Ile Gly Arg Glu Ser Pro

225 230 235 240

Phe Met Glu Leu Asn Phe Ala Leu Gln Gln Phe Leu Pro Gly Thr His

245 250 255

Gly Val Ile Asp Gly Arg Leu Ser Glu Glu Gly Ile Asn Phe Lys Leu

260 265 270

Gly Arg Asp Leu Pro Gln Lys Ile Glu Asp Asn Ile Glu Asp Phe Cys

275 280 285

Arg Lys Leu Met Ile Lys Ala Asp Gly Asp Leu Lys Glu Phe Asn Glu

290 295 300

Leu Phe Trp Ala Val His Pro Gly Gly Pro Ala Ile Leu Asn Arg Leu

305 310 315 320

Glu Ser Ile Leu Asp Leu Lys Asn Gly Lys Leu Glu Cys Ser Arg Arg

325 330 335

Ala Leu Met Asp Tyr Gly Asn Val Ser Ser Asn Thr Ile Phe Tyr Val

340 345 350

Met Glu Tyr Met Arg Glu Glu Leu Lys Arg Glu Gly Ser Glu Glu Trp

355 360 365

Gly Leu Ala Leu Ala Phe Gly Pro Gly Ile Thr Phe Glu Gly Ile Leu

370 375 380

Leu Arg Ser Leu

385

<210> 122

<211> 1185

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Arabidopsis thaliana,

optimized for expression in Nicotiana benthamiana

<400> 122

atgtctaatt ctagaatgaa tggagttgaa aagctttctt ctaagtctac tagaagagtt 60

gctaatgctg gaaaggctac tcttcttgct cttggaaagg cttttccatc tcaagttgtt 120

ccacaagaaa atcttgttga aggatttctt agagatacta agtgtgatga tgcttttatt 180

aaggaaaagc ttgaacatct ttgtaagact actactgtta agactagata tactgttctt 240

actagagaaa ttcttgctaa gtatccagaa cttactactg aaggatctcc aactattaag 300

caaagacttg aaattgctaa tgaagctgtt gttgaaatgg ctcttgaagc ttctcttgga 360

tgtattaagg aatggggaag accagttgaa gatattactc atattgttta tgtttcttct 420

tctgaaatta gacttccagg aggagatctt tatctttctg ctaagcttgg acttagaaat 480

gatgttaata gagttatgct ttattttctt ggatgttatg gaggagttac tggacttaga 540

gttgctaagg atattgctga aaataatcca ggatctagag ttcttcttac tacttctgaa 600

actactattc ttggatttag accaccaaat aaggctagac catatgatct tgttggagct 660

gctctttttg gagatggagc tgctgctgtt attattggag ctgatccaag agaatgtgaa 720

gctccatta tggaacttca ttatgctgtt caacaatttc ttccaggaac tcaaaatgtt 780

attgaaggaa gacttactga agaaggaatt aattttaagc ttggaagaga tcttccacaa 840

aagattgaag aaaatattga agaattttgt aagaagctta tgggaaaggc tggagatgaa 900

tctatggaat ttaatgatat gttttgggct gttcatccag gaggaccagc tattcttaat 960

agacttgaaa ctaagcttaa gcttgaaaag gaaaagcttg aatcttctag aagagctctt 1020

gttgattatg gaaatgtttc ttctaatact attctttatg ttatggaata tatgagagat 1080

gaacttaaga agaagggaga tgctgctcaa gaatggggac ttggacttgc ttttggacca 1140

ggaattactt ttgaaggact tcttattaga tctcttactt cttct 1185

<210> 123

<211> 395

<212>PRT

<213> Arabidopsis thaliana

<400> 123

Met Ser Asn Ser Arg Met Asn Gly Val Glu Lys Leu Ser Ser Lys Ser

1 5 10 15

Thr Arg Arg Val Ala Asn Ala Gly Lys Ala Thr Leu Leu Ala Leu Gly

20 25 30

Lys Ala Phe Pro Ser Gln Val Val Pro Gln Glu Asn Leu Val Glu Gly

35 40 45

Phe Leu Arg Asp Thr Lys Cys Asp Asp Ala Phe Ile Lys Glu Lys Leu

50 55 60

Glu His Leu Cys Lys Thr Thr Thr Val Lys Thr Arg Tyr Thr Val Leu

65 70 75 80

Thr Arg Glu Ile Leu Ala Lys Tyr Pro Glu Leu Thr Thr Glu Gly Ser

85 90 95

Pro Thr Ile Lys Gln Arg Leu Glu Ile Ala Asn Glu Ala Val Val Glu

100 105 110

Met Ala Leu Glu Ala Ser Leu Gly Cys Ile Lys Glu Trp Gly Arg Pro

115 120 125

Val Glu Asp Ile Thr His Ile Val Tyr Val Ser Ser Ser Glu Ile Arg

130 135 140

Leu Pro Gly Gly Asp Leu Tyr Leu Ser Ala Lys Leu Gly Leu Arg Asn

145 150 155 160

Asp Val Asn Arg Val Met Leu Tyr Phe Leu Gly Cys Tyr Gly Gly Val

165 170 175

Thr Gly Leu Arg Val Ala Lys Asp Ile Ala Glu Asn Asn Pro Gly Ser

180 185 190

Arg Val Leu Leu Thr Thr Ser Glu Thr Thr Ile Leu Gly Phe Arg Pro

195 200 205

Pro Asn Lys Ala Arg Pro Tyr Asp Leu Val Gly Ala Ala Leu Phe Gly

210 215 220

Asp Gly Ala Ala Ala Val Ile Ile Gly Ala Asp Pro Arg Glu Cys Glu

225 230 235 240

Ala Pro Phe Met Glu Leu His Tyr Ala Val Gln Gln Phe Leu Pro Gly

245 250 255

Thr Gln Asn Val Ile Glu Gly Arg Leu Thr Glu Glu Gly Ile Asn Phe

260 265 270

Lys Leu Gly Arg Asp Leu Pro Gln Lys Ile Glu Glu Asn Ile Glu Glu

275 280 285

Phe Cys Lys Lys Leu Met Gly Lys Ala Gly Asp Glu Ser Met Glu Phe

290 295 300

Asn Asp Met Phe Trp Ala Val His Pro Gly Gly Pro Ala Ile Leu Asn

305 310 315 320

Arg Leu Glu Thr Lys Leu Lys Leu Glu Lys Glu Lys Leu Glu Ser Ser

325 330 335

Arg Arg Ala Leu Val Asp Tyr Gly Asn Val Ser Ser Asn Thr Ile Leu

340 345 350

Tyr Val Met Glu Tyr Met Arg Asp Glu Leu Lys Lys Lys Gly Asp Ala

355 360 365

Ala Gln Glu Trp Gly Leu Gly Leu Ala Phe Gly Pro Gly Ile Thr Phe

370 375 380

Glu Gly Leu Leu Ile Arg Ser Leu Thr Ser Ser

385 390 395

<210> 124

<211> 1197

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Hydrangea macrophylla,

optimized for expression in Nicotiana benthamiana

<400> 124

atggctacta agtctgttgc tgttgaagaa atgtgtaagg ctcaaaaggc tggaggacca 60

gctactattc ttgctattgg aactgctgtt ccatctaatt gttattatca atctgaatat 120

ccagattttt attttagagt tactaagtct gatcatctta ctgatcttaa gtctaagttt 180

aagagaatgt gtgaaagatc ttctattaag aagagatata tgcatcttac tgaagaaatt 240

cttgaagaaa atccaaatat gtgtactttt gctgctccat ctattgatgg aagacaagat 300

attgttgtta aggaaattcc aaagcttgct aaggaagctg cttctaaggc tattaaggaa 360

tggggacaac caaagtctaa tattactcat cttgtttttt gtactacttc tggagttgat 420

atgccaggat gtgattatca acttactaga cttcttggac ttagaccatc tattaagaga 480

cttatgatgt atcaacaagg atgtcatgct ggaggaactg gacttagact tgctaaggat 540

cttgctgaaa ataataaggg agctagagtt cttgttgttt gttctgaaat gactgttatt 600

aattttagag gaccatctga agctcatatg gattctcttg ttggacaatc tctttttgga 660

gatggagctt ctgctgttat tgttggatct gatccagatc tttctactga acatccactt 720

tatcaaatta tgtctgcttc tcaaattatt gttgctgatt ctgaaggagc tattgatgga 780

catcttagac aagaaggact tacttttcat cttagaaagg atgttccatc tcttgtttct 840

gataatattg aaaatactct tgttgaagct tttactccaa ttcttatgga ttctattgat 900

tctattattg attggaattc tattttttgg attgctcatc caggaggacc agctattctt 960

aatcaagttc aagctaaggt tggacttaag gaagaaaagc ttagagtttc tagacatatt 1020

ctttctgaat atggaaatat gtcttctgct tgtgtttttt ttattatgga tgaaatgaga 1080

aagagatcta tggaagaagg aaagggaact actggagaag gacttgaatg gggagttctt 1140

tttggatttg gaccaggatt tactgttgaa actattgttc ttcattctgt tccaatt 1197

<210> 125

<211> 399

<212>PRT

<213> Hydrangea macrophylla

<400> 125

Met Ala Thr Lys Ser Val Ala Val Glu Glu Met Cys Lys Ala Gln Lys

1 5 10 15

Ala Gly Gly Pro Ala Thr Ile Leu Ala Ile Gly Thr Ala Val Pro Ser

20 25 30

Asn Cys Tyr Tyr Gln Ser Glu Tyr Pro Asp Phe Tyr Phe Arg Val Thr

35 40 45

Lys Ser Asp His Leu Thr Asp Leu Lys Ser Lys Phe Lys Arg Met Cys

50 55 60

Glu Arg Ser Ser Ile Lys Lys Arg Tyr Met His Leu Thr Glu Glu Ile

65 70 75 80

Leu Glu Glu Asn Pro Asn Met Cys Thr Phe Ala Ala Pro Ser Ile Asp

85 90 95

Gly Arg Gln Asp Ile Val Val Lys Glu Ile Pro Lys Leu Ala Lys Glu

100 105 110

Ala Ala Ser Lys Ala Ile Lys Glu Trp Gly Gln Pro Lys Ser Asn Ile

115 120 125

Thr His Leu Val Phe Cys Thr Thr Ser Gly Val Asp Met Pro Gly Cys

130 135 140

Asp Tyr Gln Leu Thr Arg Leu Leu Gly Leu Arg Pro Ser Ile Lys Arg

145 150 155 160

Leu Met Met Tyr Gln Gln Gly Cys His Ala Gly Gly Thr Gly Leu Arg

165 170 175

Leu Ala Lys Asp Leu Ala Glu Asn Asn Lys Gly Ala Arg Val Leu Val

180 185 190

Val Cys Ser Glu Met Thr Val Ile Asn Phe Arg Gly Pro Ser Glu Ala

195 200 205

His Met Asp Ser Leu Val Gly Gln Ser Leu Phe Gly Asp Gly Ala Ser

210 215 220

Ala Val Ile Val Gly Ser Asp Pro Asp Leu Ser Thr Glu His Pro Leu

225 230 235 240

Tyr Gln Ile Met Ser Ala Ser Gln Ile Ile Val Ala Asp Ser Glu Gly

245 250 255

Ala Ile Asp Gly His Leu Arg Gln Glu Gly Leu Thr Phe His Leu Arg

260 265 270

Lys Asp Val Pro Ser Leu Val Ser Asp Asn Ile Glu Asn Thr Leu Val

275 280 285

Glu Ala Phe Thr Pro Ile Leu Met Asp Ser Ile Asp Ser Ile Ile Asp

290 295 300

Trp Asn Ser Ile Phe Trp Ile Ala His Pro Gly Gly Pro Ala Ile Leu

305 310 315 320

Asn Gln Val Gln Ala Lys Val Gly Leu Lys Glu Glu Lys Leu Arg Val

325 330 335

Ser Arg His Ile Leu Ser Glu Tyr Gly Asn Met Ser Ser Ala Cys Val

340 345 350

Phe Phe Ile Met Asp Glu Met Arg Lys Arg Ser Met Glu Glu Gly Lys

355 360 365

Gly Thr Thr Gly Glu Gly Leu Glu Trp Gly Val Leu Phe Gly Phe Gly

370 375 380

Pro Gly Phe Thr Val Glu Thr Ile Val Leu His Ser Val Pro Ile

385 390 395

<210> 126

<211> 1254

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Physcomitrella patens,

optimized for expression in Nicotiana benthamiana

<400> 126

atggcttcta gaagagttga agctgctttt gatggacaag ctgttgaact tggagctact 60

attccagctg ctaatggaaa tggaactcat caatctatta aggttccagg acatagacaa 120

gttactccag gaaagactac tattatggct attggaagag ctgttccagc taatactact 180

tttaatgatg gacttgctga tcattatatt caagaattta atcttcaaga tccagttctt 240

caagctaagc ttagaagact ttgtgaaact actactgtta agactagata tcttgttgtt 300

aataaggaaa ttcttgatga acatccagaa tttcttgttg atggagctgc tactgtttct 360

caaagacttg ctattactgg agaagctgtt actcaacttg gacatgaagc tgctactgct 420

gctattaagg aatggggaag accagcttct gaaattactc atcttgttta tgtttcttct 480

tctgaaatta gacttccagg aggagatctt tatcttgctc aacttcttgg acttagatct 540

gatgttaata gagttatgct ttatatgctt ggatgttatg gaggagcttc tggaattaga 600

gttgctaagg atcttgctga aaataatcca ggatctagag ttcttcttat tacttctgaa 660

tgtactctta ttggatataa gtctctttct ccagatagac catatgatct tgttggagct 720

gctctttttg gagatggagc tgctgctatg attatgggaa aggatccaat tccagttctt 780

gaaagagctt tttttgaact tgattgggct ggacaatctt ttattccagg aactaataag 840

actattgatg gaagactttc tgaagaagga atttctttta agcttggaag agaacttcca 900

aagcttattg aatctaatat tcaaggattt tgtgatccaa ttcttaagag agctggagga 960

cttaagtata atgatatttt ttgggctgtt catccaggag gaccagctat tcttaatgct 1020

gttcaaaagc aacttgatct tgctccagaa aagcttcaaa ctgctagaca agttcttaga 1080

gattatggaa atatttcttc ttctacttgt atttatgttc ttgattatat gagacatcaa 1140

tctcttaagc ttaaggaagc taatgataat gttaatactg aaccagaatg gggacttctt 1200

cttgcttttg gaccaggagt tactattgaa ggagctcttc ttagaaatct ttgt 1254

<210> 127

<211> 397

<212>PRT

<213> Physcomitrella patens

<400> 127

Met Ser Lys Thr Val Glu Asp Arg Ala Ala Gln Arg Ala Lys Gly Pro

1 5 10 15

Ala Thr Val Leu Ala Ile Gly Thr Ala Thr Pro Ala Asn Val Val Tyr

20 25 30

Gln Thr Asp Tyr Pro Asp Tyr Tyr Phe Arg Val Thr Lys Ser Glu His

35 40 45

Met Thr Lys Leu Lys Asn Lys Phe Gln Arg Met Cys Asp Arg Ser Thr

50 55 60

Ile Lys Lys Arg Tyr Met Val Leu Thr Glu Glu Leu Leu Glu Lys Asn

65 70 75 80

Leu Ser Leu Cys Thr Tyr Met Glu Pro Ser Leu Asp Ala Arg Gln Asp

85 90 95

Ile Leu Val Pro Glu Val Pro Lys Leu Gly Lys Glu Ala Ala Asp Glu

100 105 110

Ala Ile Ala Glu Trp Gly Arg Pro Lys Ser Glu Ile Thr His Leu Ile

115 120 125

Phe Cys Thr Thr Cys Gly Val Asp Met Pro Gly Ala Asp Tyr Gln Leu

130 135 140

Thr Lys Leu Leu Gly Leu Arg Ser Ser Val Arg Arg Thr Met Leu Tyr

145 150 155 160

Gln Gln Gly Cys Phe Gly Gly Gly Thr Val Leu Arg Leu Ala Lys Asp

165 170 175

Leu Ala Glu Asn Asn Ala Gly Ala Arg Val Leu Val Val Cys Ser Glu

180 185 190

Ile Thr Thr Ala Val Asn Phe Arg Gly Pro Ser Asp Thr His Leu Asp

195 200 205

Leu Leu Val Gly Leu Ala Leu Phe Gly Asp Gly Ala Ala Ala Val Ile

210 215 220

Val Gly Ala Asp Pro Asp Pro Thr Leu Glu Arg Pro Leu Phe Gln Ile

225 230 235 240

Val Ser Gly Ala Gln Thr Ile Leu Pro Asp Ser Glu Gly Ala Ile Asn

245 250 255

Gly His Leu Arg Glu Val Gly Leu Thr Ile Arg Leu Leu Lys Asp Val

260 265 270

Pro Gly Leu Val Ser Met Asn Ile Glu Lys Cys Leu Met Glu Ala Phe

275 280 285

Ala Pro Met Gly Ile His Asp Trp Asn Ser Ile Phe Trp Ile Ala His

290 295 300

Pro Gly Gly Pro Thr Ile Leu Asp Gln Val Glu Ala Lys Leu Gly Leu

305 310 315 320

Lys Glu Glu Lys Leu Lys Ser Thr Arg Ala Val Leu Arg Glu Tyr Gly

325 330 335

Asn Met Ser Ser Ala Cys Val Leu Phe Ile Leu Asp Glu Val Arg Lys

340 345 350

Arg Ser Met Glu Glu Gly Lys Thr Thr Thr Gly Glu Gly Phe Asp Trp

355 360 365

Gly Val Leu Phe Gly Phe Gly Pro Gly Phe Thr Val Glu Thr Val Val

370 375 380

Leu His Ser Met Pro Ile Pro Lys Ala Asp Glu Gly Arg

385 390 395

<210> 128

<211> 1179

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Polygonum cuspidatum,

optimized for expression in Nicotiana benthamiana

<400> 128

atggctccag ctgttgctga tattagaaag gctcaaagag ctgaaggacc agctactgtt 60

cttgctattg gaactgctac tccaccaaat tgtgtttatc aaaaggatta tccagattat 120

tattttagag ttactaattc tgatcatatg actgatctta aggaaaagtt tagaagaatg 180

tgtgaaaagt ctaatattga aaagagatat atgtatctta ctgaagaaat tcttaaggaa 240

aatccaaata tgtgttctta tatgcaaact tcttctcttg atactagaca agatatggtt 300

gtttctgaag ttccaagact tggaaaggaa gctgctcaaa aggctattaa ggaatgggga 360

caaccaaagt ctaagattac tcatgttatt atgtgtacta cttctggagt tgatatgcca 420

ggagctgatt atcaacttac taagcttctt ggacttcatc catctgttaa gagatttatg 480

atgtatcaac aaggatgttt tgctggagga actgttctta gacttgctaa ggatcttgct 540

gaaaataata gaggagctag agttcttgtt gtttgttctg aaattactgc tatttgtttt 600

agaggaccaa ctgatactca tccagattct atggttggac aagctctttt tggagatgga 660

tctggagctg ttattattgg agctgatcca gatctttcta ttgaaaagcc aatttttgaa 720

cttgtttgga ctgctcaaac tattcttcca gattctgaag gagctattga tggacatctt 780

agagaagttg gacttacttt tcatcttctt aaggatgttc caggacttat ttctaagaat 840

attgaaaaga atcttactga agctttttct ccacttaatg tttctgattg gaattctctt 900

ttttggattg ctcatccagg aggaccagct attcttgatc aagttgaaac taagcttgga 960

cttaaggaag aaaagcttaa ggctactaga caagttctta atgattatgg aaatatgtct 1020

tctgcttgtg ttctttttat tatggatgaa atgagaaaga agtctgttga aaatggacat 1080

gctactactg gagaaggact tgaatgggga gttctttttg gatttggacc aggacttact 1140

gttgaaactg ttgttcttca ttctgttcca gttgctaat 1179

<210> 129

<211> 393

<212>PRT

<213> Polygonum cuspidatum

<400> 129

Met Ala Pro Ala Val Ala Asp Ile Arg Lys Ala Gln Arg Ala Glu Gly

1 5 10 15

Pro Ala Thr Val Leu Ala Ile Gly Thr Ala Thr Pro Pro Asn Cys Val

20 25 30

Tyr Gln Lys Asp Tyr Pro Asp Tyr Tyr Phe Arg Val Thr Asn Ser Asp

35 40 45

His Met Thr Asp Leu Lys Glu Lys Phe Arg Arg Met Cys Glu Lys Ser

50 55 60

Asn Ile Glu Lys Arg Tyr Met Tyr Leu Thr Glu Glu Ile Leu Lys Glu

65 70 75 80

Asn Pro Asn Met Cys Ser Tyr Met Gln Thr Ser Ser Leu Asp Thr Arg

85 90 95

Gln Asp Met Val Val Ser Glu Val Pro Arg Leu Gly Lys Glu Ala Ala

100 105 110

Gln Lys Ala Ile Lys Glu Trp Gly Gln Pro Lys Ser Lys Ile Thr His

115 120 125

Val Ile Met Cys Thr Thr Ser Gly Val Asp Met Pro Gly Ala Asp Tyr

130 135 140

Gln Leu Thr Lys Leu Leu Gly Leu His Pro Ser Val Lys Arg Phe Met

145 150 155 160

Met Tyr Gln Gln Gly Cys Phe Ala Gly Gly Thr Val Leu Arg Leu Ala

165 170 175

Lys Asp Leu Ala Glu Asn Asn Arg Gly Ala Arg Val Leu Val Val Cys

180 185 190

Ser Glu Ile Thr Ala Ile Cys Phe Arg Gly Pro Thr Asp Thr His Pro

195 200 205

Asp Ser Met Val Gly Gln Ala Leu Phe Gly Asp Gly Ser Gly Ala Val

210 215 220

Ile Ile Gly Ala Asp Pro Asp Leu Ser Ile Glu Lys Pro Ile Phe Glu

225 230 235 240

Leu Val Trp Thr Ala Gln Thr Ile Leu Pro Asp Ser Glu Gly Ala Ile

245 250 255

Asp Gly His Leu Arg Glu Val Gly Leu Thr Phe His Leu Leu Lys Asp

260 265 270

Val Pro Gly Leu Ile Ser Lys Asn Ile Glu Lys Asn Leu Thr Glu Ala

275 280 285

Phe Ser Pro Leu Asn Val Ser Asp Trp Asn Ser Leu Phe Trp Ile Ala

290 295 300

His Pro Gly Gly Pro Ala Ile Leu Asp Gln Val Glu Thr Lys Leu Gly

305 310 315 320

Leu Lys Glu Glu Lys Leu Lys Ala Thr Arg Gln Val Leu Asn Asp Tyr

325 330 335

Gly Asn Met Ser Ser Ala Cys Val Leu Phe Ile Met Asp Glu Met Arg

340 345 350

Lys Lys Ser Val Glu Asn Gly His Ala Thr Thr Gly Glu Gly Leu Glu

355 360 365

Trp Gly Val Leu Phe Gly Phe Gly Pro Gly Leu Thr Val Glu Thr Val

370 375 380

Val Leu His Ser Val Pro Val Ala Asn

385 390

<210> 130

<211> 1152

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of the L132S PKS mutant from Rheum

palmatum, optimized for expression in Nicotiana benthamiana

<400> 130

atggctactg aagaaatgaa gaagcttgct actgttatgg ctattggaac tgctaatcca 60

ccaaattgtt attatcaagc tgattttcca gatttttatt ttagagttac taattctgat 120

catcttatta atcttaagca aaagtttaag agactttgtg aaaattctag aattgaaaag 180

agatatcttc atgttactga agaaattctt aaggaaaatc caaatattgc tgcttatgaa 240

gctacttctc ttaatgttag acataagatg caagttaagg gagttgctga acttggaaag 300

gaagctgctc ttaaggctat taaggaatgg ggacaaccaa agtctaagat tactcatctt 360

attgtttgtt gttctgctgg agttgatatg ccaggagctg attatcaact tactaagctt 420

cttgatcttg atccatctgt taagagattt atgttttatc atcttggatg ttatgctgga 480

ggaactgttc ttagacttgc taaggatatt gctgaaaata ataagggagc tagagttctt 540

attgtttgtt ctgaaatgac tactacttgt tttagaggac catctgaaac tcatcttgat 600

tctatgattg gacaagctat tcttggagat ggagctgctg ctgttattgt tggagctgat 660

ccagatctta ctgttgaaag accaattttt gaacttgttt ctactgctca aactattgtt 720

ccagaatctc atggagctat tgaaggacat cttcttgaat ctggactttc ttttcatctt 780

tataagactg ttccaactct tatttctaat aatattaaga cttgtctttc tgatgctttt 840

actccactta atatttctga ttggaattct cttttttgga ttgctcatcc aggaggacca 900

gctattcttg atcaagttac tgctaaggtt ggacttgaaa aggaaaagct taaggttact 960

agacaagttc ttaaggatta tggaaatatg tcttctgcta ctgttttttt tattatggat 1020

gaaatgagaa agaagtctct tgaaaatgga caagctacta ctggagaagg acttgaatgg 1080

ggagttcttt ttggatttgg accaggaatt actgttgaaa ctgttgttct tagatctgtt 1140

ccagttattt ct 1152

<210> 131

<211> 384

<212>PRT

<213> Artificial sequence

<220>

<223> Amino acid sequence of the L132S PKS mutant from Rheum

palmatum, optimized for expression in Nicotiana benthamiana

<400> 131

Met Ala Thr Glu Glu Met Lys Lys Leu Ala Thr Val Met Ala Ile Gly

1 5 10 15

Thr Ala Asn Pro Pro Asn Cys Tyr Tyr Gln Ala Asp Phe Pro Asp Phe

20 25 30

Tyr Phe Arg Val Thr Asn Ser Asp His Leu Ile Asn Leu Lys Gln Lys

35 40 45

Phe Lys Arg Leu Cys Glu Asn Ser Arg Ile Glu Lys Arg Tyr Leu His

50 55 60

Val Thr Glu Glu Ile Leu Lys Glu Asn Pro Asn Ile Ala Ala Tyr Glu

65 70 75 80

Ala Thr Ser Leu Asn Val Arg His Lys Met Gln Val Lys Gly Val Ala

85 90 95

Glu Leu Gly Lys Glu Ala Ala Leu Lys Ala Ile Lys Glu Trp Gly Gln

100 105 110

Pro Lys Ser Lys Ile Thr His Leu Ile Val Cys Cys Ser Ala Gly Val

115 120 125

Asp Met Pro Gly Ala Asp Tyr Gln Leu Thr Lys Leu Leu Asp Leu Asp

130 135 140

Pro Ser Val Lys Arg Phe Met Phe Tyr His Leu Gly Cys Tyr Ala Gly

145 150 155 160

Gly Thr Val Leu Arg Leu Ala Lys Asp Ile Ala Glu Asn Asn Lys Gly

165 170 175

Ala Arg Val Leu Ile Val Cys Ser Glu Met Thr Thr Thr Cys Phe Arg

180 185 190

Gly Pro Ser Glu Thr His Leu Asp Ser Met Ile Gly Gln Ala Ile Leu

195 200 205

Gly Asp Gly Ala Ala Ala Val Ile Val Gly Ala Asp Pro Asp Leu Thr

210 215 220

Val Glu Arg Pro Ile Phe Glu Leu Val Ser Thr Ala Gln Thr Ile Val

225 230 235 240

Pro Glu Ser His Gly Ala Ile Glu Gly His Leu Leu Glu Ser Gly Leu

245 250 255

Ser Phe His Leu Tyr Lys Thr Val Pro Thr Leu Ile Ser Asn Asn Ile

260 265 270

Lys Thr Cys Leu Ser Asp Ala Phe Thr Pro Leu Asn Ile Ser Asp Trp

275 280 285

Asn Ser Leu Phe Trp Ile Ala His Pro Gly Gly Pro Ala Ile Leu Asp

290 295 300

Gln Val Thr Ala Lys Val Gly Leu Glu Lys Glu Lys Leu Lys Val Thr

305 310 315 320

Arg Gln Val Leu Lys Asp Tyr Gly Asn Met Ser Ser Ala Thr Val Phe

325 330 335

Phe Ile Met Asp Glu Met Arg Lys Lys Ser Leu Glu Asn Gly Gln Ala

340 345 350

Thr Thr Gly Glu Gly Leu Glu Trp Gly Val Leu Phe Gly Phe Gly Pro

355 360 365

Gly Ile Thr Val Glu Thr Val Val Leu Arg Ser Val Pro Val Ile Ser

370 375 380

<210> 132

<211> 1173

<212>PRT

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Rheum tataricum,

optimized for expression in Nicotiana benthamiana

<400> 132

Ala Thr Gly Gly Cys Thr Cys Cys Ala Gly Ala Ala Gly Ala Ala Thr

1 5 10 15

Cys Thr Ala Gly Ala Cys Ala Thr Gly Cys Thr Gly Ala Ala Ala Cys

20 25 30

Thr Gly Cys Thr Gly Thr Thr Ala Ala Thr Ala Gly Ala Gly Cys Thr

35 40 45

Gly Cys Thr Ala Cys Thr Gly Thr Thr Cys Thr Thr Gly Cys Thr Ala

50 55 60

Thr Thr Gly Gly Ala Ala Cys Thr Gly Cys Thr Ala Ala Thr Cys Cys

65 70 75 80

Ala Cys Cys Ala Ala Ala Thr Thr Gly Thr Thr Ala Thr Thr Ala Thr

85 90 95

Cys Ala Ala Gly Cys Thr Gly Ala Thr Thr Thr Thr Cys Cys Ala Gly

100 105 110

Ala Thr Thr Thr Thr Thr Ala Thr Thr Thr Thr Ala Gly Ala Gly Cys

115 120 125

Thr Ala Cys Thr Ala Ala Thr Thr Cys Thr Gly Ala Thr Cys Ala Thr

130 135 140

Cys Thr Thr Ala Cys Thr Cys Ala Thr Cys Thr Thr Ala Ala Gly Cys

145 150 155 160

Ala Ala Ala Ala Gly Thr Thr Thr Ala Ala Gly Ala Gly Ala Ala Thr

165 170 175

Thr Thr Gly Thr Gly Ala Ala Ala Ala Gly Thr Cys Thr Ala Thr Gly

180 185 190

Ala Thr Thr Gly Ala Ala Ala Ala Gly Ala Gly Ala Thr Ala Thr Cys

195 200 205

Thr Thr Cys Ala Thr Cys Thr Thr Ala Cys Thr Gly Ala Ala Gly Ala

210 215 220

Ala Ala Thr Thr Cys Thr Thr Ala Ala Gly Gly Ala Ala Ala Ala Thr

225 230 235 240

Cys Cys Ala Ala Ala Thr Ala Thr Thr Gly Cys Thr Thr Cys Thr Thr

245 250 255

Thr Thr Gly Ala Ala Gly Cys Thr Cys Cys Ala Thr Cys Thr Cys Thr

260 265 270

Thr Gly Ala Thr Gly Thr Thr Ala Gly Ala Cys Ala Thr Ala Ala Thr

275 280 285

Ala Thr Thr Cys Ala Ala Gly Thr Thr Ala Ala Gly Gly Ala Ala Gly

290 295 300

Thr Thr Gly Thr Thr Cys Thr Thr Cys Thr Thr Gly Gly Ala Ala Ala

305 310 315 320

Gly Gly Ala Ala Gly Cys Thr Gly Cys Thr Cys Thr Thr Ala Ala Gly

325 330 335

Gly Cys Thr Ala Thr Thr Ala Ala Thr Gly Ala Ala Thr Gly Gly Gly

340 345 350

Gly Ala Cys Ala Ala Cys Cys Ala Ala Ala Gly Thr Cys Thr Ala Ala

355 360 365

Gly Ala Thr Thr Ala Cys Thr Ala Gly Ala Cys Thr Thr Ala Thr Thr

370 375 380

Gly Thr Thr Thr Gly Thr Thr Gly Thr Ala Thr Thr Gly Cys Thr Gly

385 390 395 400

Gly Ala Gly Thr Thr Gly Ala Thr Ala Thr Gly Cys Cys Ala Gly Gly

405 410 415

Ala Gly Cys Thr Gly Ala Thr Thr Ala Thr Cys Ala Ala Cys Thr Thr

420 425 430

Ala Cys Thr Ala Ala Gly Cys Thr Thr Cys Thr Thr Gly Gly Ala Cys

435 440 445

Thr Thr Cys Ala Ala Cys Thr Thr Thr Cys Thr Gly Thr Thr Ala Ala

450 455 460

Gly Ala Gly Ala Thr Thr Thr Ala Thr Gly Thr Thr Thr Thr Ala Thr

465 470 475 480

Cys Ala Thr Cys Thr Thr Gly Gly Ala Thr Gly Thr Thr Ala Thr Gly

485 490 495

Cys Thr Gly Gly Ala Gly Gly Ala Ala Cys Thr Gly Thr Thr Cys Thr

500 505 510

Thr Ala Gly Ala Cys Thr Thr Gly Cys Thr Ala Ala Gly Gly Ala Thr

515 520 525

Ala Thr Thr Gly Cys Thr Gly Ala Ala Ala Ala Thr Ala Ala Thr Ala

530 535 540

Ala Gly Gly Ala Ala Gly Cys Thr Ala Gly Ala Gly Thr Thr Cys Thr

545 550 555 560

Thr Ala Thr Thr Gly Thr Thr Ala Gly Ala Thr Cys Thr Gly Ala Ala

565 570 575

Ala Thr Gly Ala Cys Thr Cys Cys Ala Ala Thr Thr Thr Gly Thr Thr

580 585 590

Thr Thr Ala Gly Ala Gly Gly Ala Cys Cys Ala Thr Cys Thr Gly Ala

595 600 605

Ala Ala Cys Thr Cys Ala Thr Ala Thr Thr Gly Ala Thr Thr Cys Thr

610 615 620

Ala Thr Gly Gly Thr Thr Gly Gly Ala Cys Ala Ala Gly Cys Thr Ala

625 630 635 640

Thr Thr Thr Thr Thr Gly Gly Ala Gly Ala Thr Gly Gly Ala Gly Cys

645 650 655

Thr Gly Cys Thr Gly Cys Thr Gly Thr Thr Ala Thr Thr Gly Thr Thr

660 665 670

Gly Gly Ala Gly Cys Thr Ala Ala Thr Cys Cys Ala Gly Ala Thr Cys

675 680 685

Thr Thr Thr Cys Thr Ala Thr Thr Gly Ala Ala Ala Gly Ala Cys Cys

690 695 700

Ala Ala Thr Thr Thr Thr Thr Gly Ala Ala Cys Thr Thr Ala Thr Thr

705 710 715 720

Thr Cys Thr Ala Cys Thr Thr Cys Thr Cys Ala Ala Ala Cys Thr Ala

725 730 735

Thr Thr Ala Thr Thr Cys Cys Ala Gly Ala Ala Thr Cys Thr Gly Ala

740 745 750

Thr Gly Gly Ala Gly Cys Thr Ala Thr Thr Gly Ala Ala Gly Gly Ala

755 760 765

Cys Ala Thr Cys Thr Thr Cys Thr Thr Gly Ala Ala Gly Thr Thr Gly

770 775 780

Gly Ala Cys Thr Thr Thr Cys Thr Thr Thr Thr Cys Ala Ala Cys Thr

785 790 795 800

Thr Thr Ala Thr Cys Ala Ala Ala Cys Thr Gly Thr Thr Cys Cys Ala

805 810 815

Thr Cys Thr Cys Thr Thr Ala Thr Thr Thr Cys Thr Ala Ala Thr Thr

820 825 830

Gly Thr Ala Thr Thr Gly Ala Ala Ala Cys Thr Thr Gly Thr Cys Thr

835 840 845

Thr Thr Cys Thr Ala Ala Gly Gly Cys Thr Thr Thr Thr Ala Cys Thr

850 855 860

Cys Cys Ala Cys Thr Thr Ala Ala Thr Ala Thr Thr Thr Cys Thr Gly

865 870 875 880

Ala Thr Thr Gly Gly Ala Ala Thr Thr Cys Thr Cys Thr Thr Thr Thr

885 890 895

Thr Thr Gly Gly Ala Thr Thr Gly Cys Thr Cys Ala Thr Cys Cys Ala

900 905 910

Gly Gly Ala Gly Gly Ala Ala Gly Ala Gly Cys Thr Ala Thr Thr Cys

915 920 925

Thr Thr Gly Ala Thr Gly Ala Thr Ala Thr Thr Gly Ala Ala Gly Cys

930 935 940

Thr Ala Cys Thr Gly Thr Thr Gly Gly Ala Cys Thr Thr Ala Ala Gly

945 950 955 960

Ala Ala Gly Gly Ala Ala Ala Ala Gly Cys Thr Thr Ala Ala Gly Gly

965 970 975

Cys Thr Ala Cys Thr Ala Gly Ala Cys Ala Ala Gly Thr Thr Cys Thr

980 985 990

Thr Ala Ala Thr Gly Ala Thr Thr Ala Thr Gly Gly Ala Ala Ala Thr

995 1000 1005

Ala Thr Gly Thr Cys Thr Thr Cys Thr Gly Cys Thr Thr Gly Thr

1010 1015 1020

Gly Thr Thr Thr Thr Thr Thr Thr Thr Ala Thr Thr Ala Thr Gly

1025 1030 1035

Gly Ala Thr Gly Ala Ala Ala Thr Gly Ala Gly Ala Ala Ala Gly

1040 1045 1050

Ala Ala Gly Thr Cys Thr Cys Thr Thr Gly Cys Thr Ala Ala Thr

1055 1060 1065

Gly Gly Ala Cys Ala Ala Gly Thr Thr Ala Cys Thr Ala Cys Thr

1070 1075 1080

Gly Gly Ala Gly Ala Ala Gly Gly Ala Cys Thr Thr Ala Ala Gly

1085 1090 1095

Thr Gly Gly Gly Gly Ala Gly Thr Thr Cys Thr Thr Thr Thr Thr

1100 1105 1110

Gly Gly Ala Thr Thr Thr Gly Gly Ala Cys Cys Ala Gly Gly Ala

1115 1120 1125

Gly Thr Thr Ala Cys Thr Gly Thr Thr Gly Ala Ala Ala Cys Thr

1130 1135 1140

Gly Thr Thr Gly Thr Thr Cys Thr Thr Thr Cys Thr Thr Cys Thr

1145 1150 1155

Gly Thr Thr Cys Cys Ala Cys Thr Thr Ala Thr Thr Ala Cys Thr

1160 1165 1170

<210> 133

<211> 391

<212>PRT

<213> Rheum tataricum

<400> 133

Met Ala Pro Glu Glu Ser Arg His Ala Glu Thr Ala Val Asn Arg Ala

1 5 10 15

Ala Thr Val Leu Ala Ile Gly Thr Ala Asn Pro Pro Asn Cys Tyr Tyr

20 25 30

Gln Ala Asp Phe Pro Asp Phe Tyr Phe Arg Ala Thr Asn Ser Asp His

35 40 45

Leu Thr His Leu Lys Gln Lys Phe Lys Arg Ile Cys Glu Lys Ser Met

50 55 60

Ile Glu Lys Arg Tyr Leu His Leu Thr Glu Glu Ile Leu Lys Glu Asn

65 70 75 80

Pro Asn Ile Ala Ser Phe Glu Ala Pro Ser Leu Asp Val Arg His Asn

85 90 95

Ile Gln Val Lys Glu Val Val Leu Leu Gly Lys Glu Ala Ala Leu Lys

100 105 110

Ala Ile Asn Glu Trp Gly Gln Pro Lys Ser Lys Ile Thr Arg Leu Ile

115 120 125

Val Cys Cys Ile Ala Gly Val Asp Met Pro Gly Ala Asp Tyr Gln Leu

130 135 140

Thr Lys Leu Leu Gly Leu Gln Leu Ser Val Lys Arg Phe Met Phe Tyr

145 150 155 160

His Leu Gly Cys Tyr Ala Gly Gly Thr Val Leu Arg Leu Ala Lys Asp

165 170 175

Ile Ala Glu Asn Asn Lys Glu Ala Arg Val Leu Ile Val Arg Ser Glu

180 185 190

Met Thr Pro Ile Cys Phe Arg Gly Pro Ser Glu Thr His Ile Asp Ser

195 200 205

Met Val Gly Gln Ala Ile Phe Gly Asp Gly Ala Ala Ala Val Ile Val

210 215 220

Gly Ala Asn Pro Asp Leu Ser Ile Glu Arg Pro Ile Phe Glu Leu Ile

225 230 235 240

Ser Thr Ser Gln Thr Ile Ile Pro Glu Ser Asp Gly Ala Ile Glu Gly

245 250 255

His Leu Leu Glu Val Gly Leu Ser Phe Gln Leu Tyr Gln Thr Val Pro

260 265 270

Ser Leu Ile Ser Asn Cys Ile Glu Thr Cys Leu Ser Lys Ala Phe Thr

275 280 285

Pro Leu Asn Ile Ser Asp Trp Asn Ser Leu Phe Trp Ile Ala His Pro

290 295 300

Gly Gly Arg Ala Ile Leu Asp Asp Ile Glu Ala Thr Val Gly Leu Lys

305 310 315 320

Lys Glu Lys Leu Lys Ala Thr Arg Gln Val Leu Asn Asp Tyr Gly Asn

325 330 335

Met Ser Ser Ala Cys Val Phe Phe Ile Met Asp Glu Met Arg Lys Lys

340 345 350

Ser Leu Ala Asn Gly Gln Val Thr Thr Gly Glu Gly Leu Lys Trp Gly

355 360 365

Val Leu Phe Gly Phe Gly Pro Gly Val Thr Val Glu Thr Val Val Leu

370 375 380

Ser Ser Val Pro Leu Ile Thr

385 390

<210> 134

<211> 1182

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS from Wachendorfia thyrsiflora,

optimized for expression in Nicotiana benthamiana

<400> 134

atggcttcta ctgaaggaat tcaagcttat agaaataata tggctgaagg accagctact 60

attatggcta ttggaactgc taatccacca aatgttgttg atgcttctac ttttccagat 120

tattattgga gagttactaa ttctgaacat ctttctccag aatatagagt taagcttaag 180

agaatttgtg aaagatcttc tattagaaag agacatcttg ttcttactga acaacttctt 240

aaggaaaatc caactcttac tacttatgtt gatgcttctt atgatgaaag acaatctatt 300

gttcttgatg ctgttccaaa gcttgcttgt gaagctgctg ctaaggctat taaggaatgg 360

ggaagaccaa agactgatat tactcatatg gttgtttgta ctggagctgg agttgatgtt 420

ccaggagttg attataagat gatgaatctt cttggacttc caccaactgt taatagagtt 480

atgctttata atgttggatg tcatgcttct ggaactgttc ttagaattgc taaggatctt 540

gctgaaaata ataagggagc tagagttctt gttgtttctt ctgaagtttc tgttatgttt 600

tttagaggac cagctgaagg agatgttgaa attcttcttg gacaagctct ttttggagat 660

ggatctgctg ctattattgt tggagctgat ccaattgaag gagttgaaaa gccaattttt 720

caaatttttt ctgcttctca aatgactctt ccagaaggag aacatcttgt tgctggacat 780

cttagagaac ttggacttac ttttcatctt aagccacaac ttccaaatac tgtttcttct 840

aatattcata agccacttaa gaaggctttt gaaccactta atattactga ttggaattct 900

attttttgga ttgttcatcc aggaggaaga gctattcttg atcaagttca agaaaagatt 960

ggacttgaag aaaataagct tgatgtttct agatatgttc ttgctgaaaa tggaaatatg 1020

atgtctgctt ctgttttttt tattatggat gaaatgagaa agagatctgc tgctcaagga 1080

tgttctacta ctggagaagg acatgaatgg ggagttcttt ttggatttgg accaggactt 1140

tctattgaaa ctgttgttct tcattctgtt ccactttcta tt 1182

<210> 135

<211> 394

<212>PRT

<213> Wachendorfia thyrsiflora

<400> 135

Met Ala Ser Thr Glu Gly Ile Gln Ala Tyr Arg Asn Asn Met Ala Glu

1 5 10 15

Gly Pro Ala Thr Ile Met Ala Ile Gly Thr Ala Asn Pro Pro Asn Val

20 25 30

Val Asp Ala Ser Thr Phe Pro Asp Tyr Tyr Trp Arg Val Thr Asn Ser

35 40 45

Glu His Leu Ser Pro Glu Tyr Arg Val Lys Leu Lys Arg Ile Cys Glu

50 55 60

Arg Ser Ser Ile Arg Lys Arg His Leu Val Leu Thr Glu Gln Leu Leu

65 70 75 80

Lys Glu Asn Pro Thr Leu Thr Thr Tyr Val Asp Ala Ser Tyr Asp Glu

85 90 95

Arg Gln Ser Ile Val Leu Asp Ala Val Pro Lys Leu Ala Cys Glu Ala

100 105 110

Ala Ala Lys Ala Ile Lys Glu Trp Gly Arg Pro Lys Thr Asp Ile Thr

115 120 125

His Met Val Val Cys Thr Gly Ala Gly Val Asp Val Pro Gly Val Asp

130 135 140

Tyr Lys Met Met Asn Leu Leu Gly Leu Pro Pro Thr Val Asn Arg Val

145 150 155 160

Met Leu Tyr Asn Val Gly Cys His Ala Ser Gly Thr Val Leu Arg Ile

165 170 175

Ala Lys Asp Leu Ala Glu Asn Asn Lys Gly Ala Arg Val Leu Val Val

180 185 190

Ser Ser Glu Val Ser Val Met Phe Phe Arg Gly Pro Ala Glu Gly Asp

195 200 205

Val Glu Ile Leu Leu Gly Gln Ala Leu Phe Gly Asp Gly Ser Ala Ala

210 215 220

Ile Ile Val Gly Ala Asp Pro Ile Glu Gly Val Glu Lys Pro Ile Phe

225 230 235 240

Gln Ile Phe Ser Ala Ser Gln Met Thr Leu Pro Glu Gly Glu His Leu

245 250 255

Val Ala Gly His Leu Arg Glu Leu Gly Leu Thr Phe His Leu Lys Pro

260 265 270

Gln Leu Pro Asn Thr Val Ser Ser Asn Ile His Lys Pro Leu Lys Lys

275 280 285

Ala Phe Glu Pro Leu Asn Ile Thr Asp Trp Asn Ser Ile Phe Trp Ile

290 295 300

Val His Pro Gly Gly Arg Ala Ile Leu Asp Gln Val Gln Glu Lys Ile

305 310 315 320

Gly Leu Glu Glu Asn Lys Leu Asp Val Ser Arg Tyr Val Leu Ala Glu

325 330 335

Asn Gly Asn Met Met Ser Ala Ser Val Phe Phe Ile Met Asp Glu Met

340 345 350

Arg Lys Arg Ser Ala Ala Gln Gly Cys Ser Thr Thr Gly Glu Gly His

355 360 365

Glu Trp Gly Val Leu Phe Gly Phe Gly Pro Gly Leu Ser Ile Glu Thr

370 375 380

Val Val Leu His Ser Val Pro Leu Ser Ile

385 390

<210> 136

<211> 1191

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS1 from Piper methysticum,

optimized for expression in Nicotiana benthamiana

<400> 136

atgtctaaga ctgttgaaga tagagctgct caaagagcta agggaccagc tactgttctt 60

gctattggaa ctgctactcc agctaatgtt gtttatcaaa ctgattatcc agattattat 120

tttagagtta ctaagtctga acatatgact aagcttaaga ataagtttca aagaatgtgt 180

gatagatcta ctattaagaa gagatatatg gttcttactg aagaacttct tgaaaagaat 240

ctttctcttt gtacttatat ggaaccatct cttgatgcta gacaagatat tcttgttcca 300

gaagttccaa agcttggaaa ggaagctgct gatgaagcta ttgctgaatg gggaagacca 360

aagtctgaaa ttactcatct tattttttgt actacttgtg gagttgatat gccaggagct 420

gattatcaac ttactaagct tcttggactt agatcttctg ttagaagaac tatgctttat 480

caacaaggat gttttggagg aggaactgtt cttagacttg ctaaggatct tgctgaaaat 540

aatgctggag ctagagttct tgttgtttgt tctgaaatta ctactgctgt taattttaga 600

ggaccatctg atactcatct tgatcttctt gttggacttg ctctttttgg agatggagct 660

gctgctgtta ttgttggagc tgatccagat ccaactcttg aaagaccact ttttcaaatt 720

gtttctggag ctcaaactat tcttccagat tctgaaggag ctattaatgg acatcttaga 780

gaagttggac ttactattag acttcttaag gatgttccag gacttgtttc tatgaatatt 840

gaaaagtgtc ttatggaagc ttttgctcca atgggaattc atgattggaa ttctattttt 900

tggattgctc atccaggagg accaactatt cttgatcaag ttgaagctaa gcttggactt 960

aaggaagaaa agcttaagtc tactagagct gttcttagag aatatggaaa tatgtcttct 1020

gcttgtgttc tttttattct tgatgaagtt agaaagagat ctatggaaga aggaaagact 1080

actactggag aaggatttga ttggggagtt ctttttggat ttggaccagg atttactgtt 1140

gaaactgttg ttcttcattc tatgccaatt ccaaaggctg atgaaggaag a 1191

<210> 137

<211> 397

<212>PRT

<213> Piper methysticum

<400> 137

Met Ser Lys Thr Val Glu Asp Arg Ala Ala Gln Arg Ala Lys Gly Pro

1 5 10 15

Ala Thr Val Leu Ala Ile Gly Thr Ala Thr Pro Ala Asn Val Val Tyr

20 25 30

Gln Thr Asp Tyr Pro Asp Tyr Tyr Phe Arg Val Thr Lys Ser Glu His

35 40 45

Met Thr Lys Leu Lys Asn Lys Phe Gln Arg Met Cys Asp Arg Ser Thr

50 55 60

Ile Lys Lys Arg Tyr Met Val Leu Thr Glu Glu Leu Leu Glu Lys Asn

65 70 75 80

Leu Ser Leu Cys Thr Tyr Met Glu Pro Ser Leu Asp Ala Arg Gln Asp

85 90 95

Ile Leu Val Pro Glu Val Pro Lys Leu Gly Lys Glu Ala Ala Asp Glu

100 105 110

Ala Ile Ala Glu Trp Gly Arg Pro Lys Ser Glu Ile Thr His Leu Ile

115 120 125

Phe Cys Thr Thr Cys Gly Val Asp Met Pro Gly Ala Asp Tyr Gln Leu

130 135 140

Thr Lys Leu Leu Gly Leu Arg Ser Ser Val Arg Arg Thr Met Leu Tyr

145 150 155 160

Gln Gln Gly Cys Phe Gly Gly Gly Thr Val Leu Arg Leu Ala Lys Asp

165 170 175

Leu Ala Glu Asn Asn Ala Gly Ala Arg Val Leu Val Val Cys Ser Glu

180 185 190

Ile Thr Thr Ala Val Asn Phe Arg Gly Pro Ser Asp Thr His Leu Asp

195 200 205

Leu Leu Val Gly Leu Ala Leu Phe Gly Asp Gly Ala Ala Ala Val Ile

210 215 220

Val Gly Ala Asp Pro Asp Pro Thr Leu Glu Arg Pro Leu Phe Gln Ile

225 230 235 240

Val Ser Gly Ala Gln Thr Ile Leu Pro Asp Ser Glu Gly Ala Ile Asn

245 250 255

Gly His Leu Arg Glu Val Gly Leu Thr Ile Arg Leu Leu Lys Asp Val

260 265 270

Pro Gly Leu Val Ser Met Asn Ile Glu Lys Cys Leu Met Glu Ala Phe

275 280 285

Ala Pro Met Gly Ile His Asp Trp Asn Ser Ile Phe Trp Ile Ala His

290 295 300

Pro Gly Gly Pro Thr Ile Leu Asp Gln Val Glu Ala Lys Leu Gly Leu

305 310 315 320

Lys Glu Glu Lys Leu Lys Ser Thr Arg Ala Val Leu Arg Glu Tyr Gly

325 330 335

Asn Met Ser Ser Ala Cys Val Leu Phe Ile Leu Asp Glu Val Arg Lys

340 345 350

Arg Ser Met Glu Glu Gly Lys Thr Thr Thr Gly Glu Gly Phe Asp Trp

355 360 365

Gly Val Leu Phe Gly Phe Gly Pro Gly Phe Thr Val Glu Thr Val Val

370 375 380

Leu His Ser Met Pro Ile Pro Lys Ala Asp Glu Gly Arg

385 390 395

<210> 138

<211> 1182

<212> DNA

<213> Artificial sequence

<220>

<223> Nucleotide sequence of PKS2 from Piper methysticum,

optimized for expression in Nicotiana benthamiana

<400> 138

atgtctaaga tggttgaaga acattgggct gctcaaagag ctagaggacc agctactgtt 60

cttgctattg gaactgctaa tccaccaaat gttctttatc aagctgatta tccagatttt 120

tattttagag ttactaagtc tgaacatatg actcaactta aggaaaagtt taagagaatt 180

tgtgataagt ctgctattag aaagagacat cttcatctta ctgaagaact tcttgaaaag 240

aatccaaata tttgtgctca tatggctcca tctcttgatg ctagacaaga tattgctgtt 300

gttgaagttc caaagcttgc taaggaagct gctactaagg ctattaagga atggggaaga 360

ccaaagtctg atattactca tcttattttt tgtactactt gtggagttga tatgccagga 420

gctgattatc aacttactac tcttcttgga cttagaccaa ctgttagaag aactatgctt 480

tatcaacaag gatgttttgc tggaggaact gttcttagac atgctaagga ttttgctgaa 540

aataatagag gagctagagt tcttgctgtt tgttctgaat ttactgttat gaatttttct 600

ggaccatctg aagctcatct tgattctatg gttggaatgg ctctttttgg agatggagct 660

tctgctgtta ttgttggagc tgatccagat tttgctattg aaagaccact ttttcaactt 720

gtttctacta ctcaaactat tgttccagat tctgatggag ctattaagtg tcatcttaag 780

gaagttggac ttactcttca tcttgttaag aatgttccag atcttatttc taataatatg 840

gataagattc ttgaagaagc ttttgctcca cttggaatta gagatggaa ttctattttt 900

tggactgctc atccaggagg agctgctatt cttgatcaac ttgaagctaa gcttggactt 960

aataaggaaa agcttaagac tactagaact gttcttagag aatatggaaa tatgtcttct 1020

gcttgtgttt gttttgttct tgatgaaatg agaagatctt ctcttgaaga aggaaagact 1080

acttctggag aaggacttga atggggaatt cttcttggat ttggaccagg acttactgtt 1140

gaaactgttg ttcttagatc tgttccaatt tctactgcta at 1182

<210> 139

<211> 394

<212>PRT

<213> Piper methysticum

<400> 139

Met Ser Lys Met Val Glu Glu His Trp Ala Ala Gln Arg Ala Arg Gly

1 5 10 15

Pro Ala Thr Val Leu Ala Ile Gly Thr Ala Asn Pro Pro Asn Val Leu

20 25 30

Tyr Gln Ala Asp Tyr Pro Asp Phe Tyr Phe Arg Val Thr Lys Ser Glu

35 40 45

His Met Thr Gln Leu Lys Glu Lys Phe Lys Arg Ile Cys Asp Lys Ser

50 55 60

Ala Ile Arg Lys Arg His Leu His Leu Thr Glu Glu Leu Leu Glu Lys

65 70 75 80

Asn Pro Asn Ile Cys Ala His Met Ala Pro Ser Leu Asp Ala Arg Gln

85 90 95

Asp Ile Ala Val Val Glu Val Pro Lys Leu Ala Lys Glu Ala Ala Thr

100 105 110

Lys Ala Ile Lys Glu Trp Gly Arg Pro Lys Ser Asp Ile Thr His Leu

115 120 125

Ile Phe Cys Thr Thr Cys Gly Val Asp Met Pro Gly Ala Asp Tyr Gln

130 135 140

Leu Thr Thr Leu Leu Gly Leu Arg Pro Thr Val Arg Arg Thr Met Leu

145 150 155 160

Tyr Gln Gln Gly Cys Phe Ala Gly Gly Thr Val Leu Arg His Ala Lys

165 170 175

Asp Phe Ala Glu Asn Asn Arg Gly Ala Arg Val Leu Ala Val Cys Ser

180 185 190

Glu Phe Thr Val Met Asn Phe Ser Gly Pro Ser Glu Ala His Leu Asp

195 200 205

Ser Met Val Gly Met Ala Leu Phe Gly Asp Gly Ala Ser Ala Val Ile

210 215 220

Val Gly Ala Asp Pro Asp Phe Ala Ile Glu Arg Pro Leu Phe Gln Leu

225 230 235 240

Val Ser Thr Thr Gln Thr Ile Val Pro Asp Ser Asp Gly Ala Ile Lys

245 250 255

Cys His Leu Lys Glu Val Gly Leu Thr Leu His Leu Val Lys Asn Val

260 265 270

Pro Asp Leu Ile Ser Asn Asn Met Asp Lys Ile Leu Glu Glu Ala Phe

275 280 285

Ala Pro Leu Gly Ile Arg Asp Trp Asn Ser Ile Phe Trp Thr Ala His

290 295 300

Pro Gly Gly Ala Ala Ile Leu Asp Gln Leu Glu Ala Lys Leu Gly Leu

305 310 315 320

Asn Lys Glu Lys Leu Lys Thr Thr Arg Thr Val Leu Arg Glu Tyr Gly

325 330 335

Asn Met Ser Ser Ala Cys Val Cys Phe Val Leu Asp Glu Met Arg Arg

340 345 350

Ser Ser Leu Glu Glu Gly Lys Thr Thr Ser Gly Glu Gly Leu Glu Trp

355 360 365

Gly Ile Leu Leu Gly Phe Gly Pro Gly Leu Thr Val Glu Thr Val Val

370 375 380

Leu Arg Ser Val Pro Ile Ser Thr Ala Asn

385 390

<210> 140

<211> 1734

<212> DNA

<213> Arabidopsis thaliana

<400> 140

atggaagaag attataagat ggctccacaa gaacaagctg tttctcaagt tatggaaaag 60

caatctaata ataataattc tgatgttatt tttagatcta agcttccaga tatttatatt 120

ccaaatcatc tttctcttca tgattatatt tttcaaaata tttctgaatt tgctactaag 180

ccatgtctta ttaatggacc aactggacat gtttatactt attctgatgt tcatgttatt 240

tctagacaaa ttgctgctaa ttttcataag cttggagtta atcaaaatga tgttgttatg 300

cttcttcttc caaattgtcc agaatttgtt ctttcttttc ttgctgcttc ttttagagga 360

gctactgcta ctgctgctaa tccatttttt actccagctg aaattgctaa gcaagctaag 420

gcttctaata ctaagcttat tattactgaa gctagatatg ttgataagat taagccactt 480

caaaatgatg atggagttgt tattgtttgt attgatgata atgaatctgt tccaattcca 540

gaaggatgtc ttagatttac tgaacttact caatctacta ctgaagcttc tgaagttatt 600

gattctgttg aaatttctcc agatgatgtt gttgctcttc catattcttc tggaactact 660

ggacttccaa agggagttat gcttactcat aagggacttg ttacttctgt tgctcaacaa 720

gttgatggag aaaatccaaa tctttatttt cattctgatg atgttattct ttgtgttctt 780

ccaatgtttc atatttatgc tcttaattct attatgcttt gtggacttag agttggagct 840

gctattctta ttatgccaaa gtttgaaatt aatcttcttc ttgaacttat tcaaagatgt 900

aaggttactg ttgctccaat ggttccacca attgttcttg ctattgctaa gtcttctgaa 960

actgaaaagt atgatctttc ttctattaga gttgttaagt ctggagctgc tccacttgga 1020

aaggaacttg aagatgctgt taatgctaag tttccaaatg ctaagcttgg acaaggatat 1080

ggaatgactg aagctggacc agttcttgct atgtctcttg gatttgctaa ggaaccattt 1140

ccagttaagt ctggagcttg tggaactgtt gttagaaatg ctgaaatgaa gattgttgat 1200

ccagatactg gagattctct ttctagaaat caaccaggag aaatttgtat tagaggacat 1260

caaattatga agggatatct taataatcca gctgctactg ctgaaactat tgataaggat 1320

ggatggcttc atactggaga tattggactt attgatgatg atgatgaact ttttattgtt 1380

gatagactta aggaacttat taagtataag ggatttcaag ttgctccagc tgaacttgaa 1440

gctcttctta ttggacatcc agatattact gatgttgctg ttgttgctat gaaggaagaa 1500

gctgctggag aagttccagt tgcttttgtt gttaagtcta aggattctga actttctgaa 1560

gatgatgtta agcaatttgt ttctaagcaa gttgtttttt ataagagaat taataaggtt 1620

ttttttactg aatctattcc aaaggctcca tctggaaaga ttcttagaaa ggatcttaga 1680

gctaagcttg ctaatggact ttctggatat ccatatgatg ttccagatta tgct 1734

<210> 141

<211> 578

<212>PRT

<213> Arabidopsis thaliana

<400> 141

Met Glu Glu Asp Tyr Lys Met Ala Pro Gln Glu Gln Ala Val Ser Gln

1 5 10 15

Val Met Glu Lys Gln Ser Asn Asn Asn Asn Ser Asp Val Ile Phe Arg

20 25 30

Ser Lys Leu Pro Asp Ile Tyr Ile Pro Asn His Leu Ser Leu His Asp

35 40 45

Tyr Ile Phe Gln Asn Ile Ser Glu Phe Ala Thr Lys Pro Cys Leu Ile

50 55 60

Asn Gly Pro Thr Gly His Val Tyr Thr Tyr Ser Asp Val His Val Ile

65 70 75 80

Ser Arg Gln Ile Ala Ala Asn Phe His Lys Leu Gly Val Asn Gln Asn

85 90 95

Asp Val Val Met Leu Leu Leu Pro Asn Cys Pro Glu Phe Val Leu Ser

100 105 110

Phe Leu Ala Ala Ser Phe Arg Gly Ala Thr Ala Thr Ala Ala Asn Pro

115 120 125

Phe Phe Thr Pro Ala Glu Ile Ala Lys Gln Ala Lys Ala Ser Asn Thr

130 135 140

Lys Leu Ile Ile Thr Glu Ala Arg Tyr Val Asp Lys Ile Lys Pro Leu

145 150 155 160

Gln Asn Asp Asp Gly Val Val Ile Val Cys Ile Asp Asp Asn Glu Ser

165 170 175

Val Pro Ile Pro Glu Gly Cys Leu Arg Phe Thr Glu Leu Thr Gln Ser

180 185 190

Thr Thr Glu Ala Ser Glu Val Ile Asp Ser Val Glu Ile Ser Pro Asp

195 200 205

Asp Val Val Ala Leu Pro Tyr Ser Ser Gly Thr Thr Gly Leu Pro Lys

210 215 220

Gly Val Met Leu Thr His Lys Gly Leu Val Thr Ser Val Ala Gln Gln

225 230 235 240

Val Asp Gly Glu Asn Pro Asn Leu Tyr Phe His Ser Asp Asp Val Ile

245 250 255

Leu Cys Val Leu Pro Met Phe His Ile Tyr Ala Leu Asn Ser Ile Met

260 265 270

Leu Cys Gly Leu Arg Val Gly Ala Ala Ile Leu Ile Met Pro Lys Phe

275 280 285

Glu Ile Asn Leu Leu Leu Glu Leu Ile Gln Arg Cys Lys Val Thr Val

290 295 300

Ala Pro Met Val Pro Pro Ile Val Leu Ala Ile Ala Lys Ser Ser Glu

305 310 315 320

Thr Glu Lys Tyr Asp Leu Ser Ser Ile Arg Val Val Lys Ser Gly Ala

325 330 335

Ala Pro Leu Gly Lys Glu Leu Glu Asp Ala Val Asn Ala Lys Phe Pro

340 345 350

Asn Ala Lys Leu Gly Gln Gly Tyr Gly Met Thr Glu Ala Gly Pro Val

355 360 365

Leu Ala Met Ser Leu Gly Phe Ala Lys Glu Pro Phe Pro Val Lys Ser

370 375 380

Gly Ala Cys Gly Thr Val Val Arg Asn Ala Glu Met Lys Ile Val Asp

385 390 395 400

Pro Asp Thr Gly Asp Ser Leu Ser Arg Asn Gln Pro Gly Glu Ile Cys

405 410 415

Ile Arg Gly His Gln Ile Met Lys Gly Tyr Leu Asn Asn Pro Ala Ala

420 425 430

Thr Ala Glu Thr Ile Asp Lys Asp Gly Trp Leu His Thr Gly Asp Ile

435 440 445

Gly Leu Ile Asp Asp Asp Asp Glu Leu Phe Ile Val Asp Arg Leu Lys

450 455 460

Glu Leu Ile Lys Tyr Lys Gly Phe Gln Val Ala Pro Ala Glu Leu Glu

465 470 475 480

Ala Leu Leu Ile Gly His Pro Asp Ile Thr Asp Val Ala Val Val Ala

485 490 495

Met Lys Glu Glu Ala Ala Gly Glu Val Pro Val Ala Phe Val Val Lys

500 505 510

Ser Lys Asp Ser Glu Leu Ser Glu Asp Asp Val Lys Gln Phe Val Ser

515 520 525

Lys Gln Val Val Phe Tyr Lys Arg Ile Asn Lys Val Phe Phe Thr Glu

530 535 540

Ser Ile Pro Lys Ala Pro Ser Gly Lys Ile Leu Arg Lys Asp Leu Arg

545 550 555 560

Ala Lys Leu Ala Asn Gly Leu Ser Gly Tyr Pro Tyr Asp Val Pro Asp

565 570 575

Tyr Ala

<---

Claims (42)

1. Fungal luciferin biosynthesis protein, which is caffeoylpyruvate hydrolase, consisting of: a) an amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75; or b) an amino acid sequence that has at least 60% identity with an amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75, where the specified caffeoylpyruvate hydrolase catalyzes the conversion of 6-aryl-2-hydroxy- 4-oxohexa-2,5-dienoic acids, having the structural formula , where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula . 2. The protein according to claim 1, where the amino acid sequence of caffeoylpyruvate hydrolase has at least 65% identity, or at least 70% identity, or at least 75% identity, or at least 80% identity, or at least 85% identity, or at least 90% identity, or at least 95% identity with an amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75. 3. Protein 2, where the amino acid sequence of caffeoylpyruvate hydrolase has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98% or 99% identity with the amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75. 4. Protein according to claim 1, where 3-arylacrylic acid is selected from the group: caffeic acid, cinnamic acid, paracoumaric acid, coumaric acid, umbellic acid, sinapic acid, ferulic acid. 5. Application of fungal luciferin biosynthesis protein, consisting of: a) an amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75; or b) an amino acid sequence that has at least 60% identity with an amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75, in in vitro or in vivo systems as caffeoylpyruvate hydrolase, catalyzing the reaction of conversion of 6-aryl-2-hydroxy-4-oxohexa-2,5-dienoic acids, having the structural formula , where R is aryl or heteroaryl, to 3-arylacrylic acid with the structural formula . 6. Use according to claim 5, where the amino acid sequence of caffeoylpyruvate hydrolase has at least 65% identity, or at least 70% identity, or at least 75% identity, or at least 80% identity, or at least 85% identity, or at least 90% identity, or at least 95% identity with an amino acid sequence selected from the group SEQ ID NO: 65, 67, 69, 71, 73, 75. 7. Use according to claim 5, where 3-arylacrylic acid is selected from the group: caffeic acid, cinnamic acid, paracoumaric acid, coumaric acid, umbellic acid, sinapic acid, ferulic acid. 8. Nucleic acid encoding the protein of fungal luciferin biosynthesis according to claim 1. 9. An expression cassette containing (a) a transcription initiation region functional in the host cell; (b) a nucleic acid according to claim 8; and (c) a transcription termination region functional in a host cell, wherein the host cell is not a human embryonic cell. 10. A host cell containing at least one expression cassette according to claim 9 as part of an extrachromosomal element or integrated into the genome of the cell as a result of the introduction of the specified cassette into the specified cell, where the specified cell expresses at least one of the functional proteins of fungal luciferin biosynthesis, which is caffeoylpyruvate hydrolase, and the cell is not a human embryonic cell. 11. The use of a nucleic acid encoding a protein for the biosynthesis of fungal luciferin, which is caffeoylpyruvate hydrolase, according to claim 8, for obtaining caffeoylpyruvate hydrolase according to claim 1 in in vitro or in vivo systems. 12. Use according to claim 11, where the nucleic acid is used for the expression of a fungal luciferin biosynthesis protein as part of an expression cassette, which also includes a transcription initiation region functional in the host cell and a transcription termination region functional in the host cell, wherein the host cell is not is a human embryonic cell. 13. Use according to claim 12, wherein the expression cassette is used in a host cell, wherein the host cell is not a human embryonic cell. 14. A transgenic organism capable of autonomous bioluminescence, where said organism contains at least one nucleic acid according to claim 8 as part of an extrachromosomal element or integrated into the genome of a cell as a result of the introduction of an expression cassette according to claim 9 into the specified cell, as well as a nucleic acid , encoding: (a) hispidin hydroxylase, capable of catalyzing the conversion of 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula , to 6-(2-arylvinyl)-3,4-dihydroxy-2H-pyran-2-one, having the structural formula , where R is aryl or heteroaryl; and a nucleic acid encoding: (b) luciferase, capable of oxidizing fungal luciferin to release light, and the transgenic organism is not a human being or a human embryo. 15. A transgenic organism according to claim 14, also containing a nucleic acid encoding hispidin synthase capable of catalyzing the conversion of 3-arylacrylic acid with the structural formula , where R is aryl or heteroaryl in 6-(2-arylvinyl)-4-hydroxy-2H-pyran-2-one, having the structural formula , where R is aryl or heteroaryl. 16. The transgenic organism according to claim 15, also containing a nucleic acid encoding type III polyketide synthase capable of catalyzing the synthesis of hispidin from caffeyl-CoA. 17. The transgenic organism according to claim 15, also containing a nucleic acid encoding 4'-phosphopantotheinyl transferase and/or nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid. 18. Transgenic organism according to claim 17, where the enzymes for the biosynthesis of 3-arylacrylic acid are selected from the group: (a) tyrosine ammonium lyase, the amino acid sequence of which is SEQ ID No: 107 or at least 40% identical to the sequence shown in SEQ ID No: 107; HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase components, the amino acid sequences of which are SEQ ID NOs: 109 and 111 or are at least 40% sequence identical to the HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase components of E. coli, shown in SEQ ID NO: 109 and 111; (b) phenylalanine ammonium lyase, the amino acid sequence of which is SEQ ID NO:117 or is at least 40% identical to the amino acid sequence shown in SEQ ID NO:117. 19. The transgenic organism according to claim 16, also containing a nucleic acid encoding coumarate-CoA ligase and/or nucleic acids encoding enzymes for the biosynthesis of 3-arylacrylic acid. 20. Transgenic organism according to claim 19, where the enzymes for the biosynthesis of 3-arylacrylic acid are selected from the group: (a) tyrosine ammonium lyase, the amino acid sequence of which is SEQ ID NO: 107 or at least 40% identical to the sequence shown in SEQ ID NO: 107; the HpaB and HpaC 4-hydroxyphenylacetate 3-monooxygenase reductase components, the amino acid sequences of which are at least 40% identical to the sequences of the E. coli 4-hydroxyphenylacetate 3-monooxygenase reductase components HpaB and HpaC shown in SEQ ID NOs: 109 and 111; (b) phenylalanine ammonium lyase, the amino acid sequence of which is SEQ ID NO: 117 or is at least 40% identical to the amino acid sequence shown in SEQ ID NO: 117.