KR101612157B1 - Light harvesting pigment from cyanobacterium and use thereof - Google Patents

Abstract

The present invention relates to a novel optical mathematical colorant derived from Sambrook, and more particularly to a gene and protein of each of optical mathematical pigments GAF2, GAF3 and GAF4 derived from microcoleus sp. And a method for increasing the efficiency of using photosynthesis. The present invention is a novel light harvesting dye capable of absorbing ultraviolet light and green light. It introduces green of visible light into photosynthetic plants such as mosses, green algae, and higher plants which can not be used for photosynthesis energy conversion, To increase the productivity of crops.

Description

[0001] This invention relates to an optical mathematical coloring matter derived from a < RTI ID = 0.0 >

The present invention relates to a novel optical mathematical colorant derived from Sambrook , and more particularly to optical mathematical pigments GAF2, GAF3, and GAF4 derived from Microcoleus sp. To a method for increasing efficiency.

Light not only provides energy to plants through photosynthesis, but also directly affects the environmental adaptation and survival of plants as an external stimulus source. Among the oxygen-producing photosynthetic organisms, green algae and terrestrial higher plants, which do not have phycoerythrin as a light harvesting pigment but have auxotrophy of chlorophyll b, use blue light and red light as main light sources for photosynthesis. However, ultraviolet light and green light are used as chlorophyll It can not be used for photosynthesis. Therefore, it is expected that the photosynthetic light utilization efficiency will be significantly increased if the nematode, green algae and terrestrial higher plants that do not have pico erythrin absorb ultraviolet and green light.

Phytochromes and cryptochromes are typical photoreceptors in plants. Phytochrome, which is present in the cytoplasm of plants and algae, recognizes the qualitative aspect of light and causes a molecular reaction. Phytochrome absorbs red light and red light and regulates the mechanism associated with photoperiods such as flowering and germination. This phytochrome is a protein having a size of about 120 kDa, and has a site at which a chromophore binds as a light-recognizing part at the N-terminus, and a histidine kinase domain at the C-terminal site . The chromophores of phytochrome have an open-chain tetrapyrrole structure. Synechocystis sp. Known photoreceptors for PCC6803 include cph I (cyanobacteria phytochrome I; position slr0473), cph II (cyanobacteria phytochrome II; position sll0821) and plpA (cyanobacteria phytochrome III; position sll1124).

Among these, Cph I has a structure similar to the phytochrome of common plants, while Cph II has slightly different structural differences. Cph II has a size of 145 kDa and an amino acid sequence similar to the N-terminal region of the plant phytochrome exists at the N-terminal region, but no kinase domain exists at the C-terminal region.

In recent years, attempts have been made to identify photoreceptors in the case of S-bacillus, and as a result, it has been found that Synechocystis sp. PCC6803 and Nostoc A number of photoreceptors cyanobacteriochrome (CBCR), which absorb ultraviolet, blue, green, orange, and red, have been reported in punctiforme ATCC 29133. Song JY et al. 2011. Near-UV cyanobacteriochrome signaling system elicits negative phototaxis in the cyanobacterium Synechocystis sp. PCC 6803. Proc Natl Acad Sci 108: 10780-10785; Rockwell NC et al. 2012. Phycoviolobilin formation and spectral tuning in the DXCF cyanobacteriochrome subfamily. Biochemistry 51: 1449-1463).

Although a variety of photoreceptors have been found in S. aureus, a multi-photoreceptor protein (multi-GAF domain holoprotein) containing a plurality of photoreceptors has not yet been elucidated. In particular, it has not been disclosed that photoreceptor proteins having three GAF domains corresponding to the phytochrome subfamily are transformed by ultraviolet and green light, and that the absorbed light energy transfers energy to the adjacent GAF domain.

Korean Patent Publication No. 10-2012-0041408

Accordingly, the present invention has been made in view of the above-described needs, and the present inventors have found that, among the four photoreceptor candidates (GAF domains) constituting the optical mathematical color protein derived from Microcoleus sp., 2 and 3 are orange 605 nm). In the fourth step, it was confirmed that the morphological conversion reversibly occurs due to the photoreactive activity of ultraviolet / green light. In particular, seven variants of amino acid residue substitutions in GAF # 4 confirmed that cysteine 768 is essential for ultraviolet / green light absorption, while cysteine 713 and tyrosine 769 are involved in the absorption domain determination. Tyrosine 741 should be present for light conversion, and tyrosine 674, which has not been studied to date, is likely to be a novel motif involved in light absorption and conversion of the GAF domain. In addition, GAF2 / 3/4 recombinant protein with 2, 3 and 4 GAF mutations were found to absorb UV and orange color. The fluorescence emission from GAF2 / 3/4 protein was confirmed to be the same as that of GAF2 and GAF3 by chlorophyll fluorescence analysis at room temperature. Thus, it was confirmed that the green light absorbed in GAF4 was domain transferred to GAF2 and 3, .

In order to achieve the above object, the present invention provides a light harvesting dye GAF2 polynucleotide having the nucleotide sequence of SEQ ID NO: 1.

In one embodiment of the present invention, the gene may be derived from S. cerevisiae, Microcoleus B353.

In one embodiment of the present invention, the light harvesting dye is capable of absorbing orange light having a wavelength range of 600 to 605 nm.

In addition, the present invention provides a light harvesting dye GAF2 domain having the amino acid sequence of SEQ ID NO: 2.

In addition, the present invention provides a light harvesting dye GAF3 polynucleotide having the nucleotide sequence of SEQ ID NO: 3.

In one embodiment of the present invention, the gene may be derived from S. cerevisiae, Microcoleus B353.

In one embodiment of the present invention, the light harvesting dye is capable of absorbing orange light having a wavelength range of 600 to 605 nm.

In addition, the present invention provides a light harvesting dye GAF3 domain having the amino acid sequence of SEQ ID NO: 4.

Further, the present invention provides a light harvesting dye GAF4 polynucleotide having the nucleotide sequence of SEQ ID NO: 5.

In one embodiment of the present invention, the gene may be derived from S. cerevisiae, Microcoleus B353.

In one embodiment of the present invention, the light harvesting dye may have optical reversible activity of ultraviolet / green light.

In one embodiment of the present invention, the photo reversible activity of the ultraviolet / green light may be a state of being able to absorb green light upon absorption of ultraviolet light and returning to a state capable of absorbing ultraviolet light upon absorption of green light .

In one embodiment of the present invention, the light harvesting dye may be an auxiliary dye capable of transferring the absorbed energy to other dye molecules.

In addition, the present invention provides a light harvesting dye GAF4 domain having the amino acid sequence of SEQ ID NO: 6.

Further, the present invention provides a polynucleotide comprising a GAF2 polynucleotide having the nucleotide sequence of SEQ ID NO: 1; A GAF3 polynucleotide having the nucleotide sequence of SEQ ID NO: 3; And a GAF4 polynucleotide having the nucleotide sequence of SEQ ID NO: 5.

Further, the present invention provides a recombinant vector comprising a GAF2 domain having the amino acid sequence of SEQ ID NO: 2; A GAF3 domain having the amino acid sequence of SEQ ID NO: 4; And a GAF4 domain having the amino acid sequence of SEQ ID NO: 6. The present invention also provides an ultraviolet / green light harvesting pigment protein having an amino acid sequence of SEQ ID NO: 10.

In addition, the present invention provides a recombinant vector comprising the ultraviolet / green light harvest pigment gene.

In addition, the present invention provides a method for introducing the vector into a plant to increase the utilization efficiency of photosynthesis.

In one embodiment of the present invention, the plant is selected from the group consisting of Ms. Green algae having chlorophyll b as an auxiliary pigment; And terrestrial higher plants.

In one embodiment of the present invention, the efficiency of utilizing the photosynthesis can be increased by introducing ultraviolet / green light harvest pigment genes into plants and using ultraviolet light and green light as light sources for photosynthesis.

The present invention is a novel light harvesting dye capable of absorbing ultraviolet light and green light. The pigment is introduced into photosynthetic plants such as mosses, green algae, and higher plants that can not be used for photosynthesis energy conversion of visible light, By dramatically increasing the efficiency, it can contribute to productivity improvement of crops.

Figure 1 shows the domain structure of the GHP protein from Microcoleus and a well-preserved CH motif. 1A is a schematic diagram of a GHP protein composed of four GAF domains and one Methyl accepting chemotaxis protein (MCP). Fig. 1B shows the amino acid sequences of the four GAF domains of GHP, the S. cerevisiae phytochrome Cph1, and the GAF domain of the ultraviolet / green photoreceptor UirS. In the CH motif, GAF1 has no cysteine residue, GAF2 and 3 have CH, and GAF4 has CY Residue. It also has Cys713 instead of the DXCF motif in UirS.
FIG. 2 is a photograph showing recombinant His6GHP-GAF2, His6GHP-GAF3, His6GHP-GAF4 and His6GHP-GAF2 / 3/4 solutions isolated from Escherichia coli producing PCBs.
FIG. 3 is a graph showing the results of the expression of Coomassie brilliant blue (CBB) of seven mutants (FIG. 3B) of GAF2, GAF3, GAF4 and GAF2 / 3/4 domain proteins of GHP isolated from E. coli (FIG. This is a photograph showing the staining and zinc (Zn2 +) -dependent fluorescence emission.
4 is a graph showing the spectroscopic characteristics of recombinant micro-core GHP. Absorption (A) of recombinant His6GHP-GAF2, His6GHP-GAF3, His6GHP-GAF4 and His6GHP-GAF2 / 3/4 isolated from PCB-producing E. coli, Absorption after protein structural change (B) , And a room temperature emission fluorescent spectrum (D).
FIG. 5 shows the absorption spectra of seven protein variants produced through substitution of the conserved motif amino acids of the GAF4 domain of the GHP protein.
FIG. 6 is a schematic diagram showing the absorption and transmission of light energy between the GAF domains of the GHP proteins. FIG. 6 is a schematic diagram showing that light energy is transferred to GAF2 or GAF3 after green light is absorbed by GAF4.
Fig. 7 shows the vector map of recombinant plasmids in which the base sequence (GAF2, GAF3, GAF4 or GAF2 / 3/4) of each GAF domain is linked to pBAD / mycHisC.

The present invention is characterized by providing an ultraviolet / green light harvesting pigment protein including all of the novel light harvesting pigments GAF2, GAF3, GAF4, and GAF2 to GAF4.

More specifically, the present invention relates to a polynucleotide of each of light harvesting pigments GAF2, GAF3 and GAF4 having a specific base sequence and an ultraviolet / green light harvesting pigment gene comprising both of them. And domains of light harvesting pigments GAF2, GAF3, and GAF4 having specific amino acid sequences, and ultraviolet / green light harvesting pigment proteins including both of them.

In the present invention, the 'light harvesting pigment' is a concept collectively referred to as a light harvesting pigment, which is a concept that collects light in photosynthesis and transmits the energy to the reaction center.

In the present invention, 'ultraviolet / green light harvesting pigment' means a pigment capable of absorbing ultraviolet light and green light. In the following, this UV / Green Light Harvesting Pigment is briefly referred to as 'GHP'.

The inventors of the present invention have found that fungal strains of Microcoleus sp. Based on the results of the genome analysis of B353, it was first identified that the GHP protein of the microorganism has four GAF domains and one methylation-accepting chemotaxis protein (MCP). Especially, among the four GAF domains, 3 Were found to have photoreceptor functions, and in the present invention they were named as GAF 2, GAF 3 and GAF 4, respectively.

First, the present invention provides a light-harvesting dye GAF2 polynucleotide having the nucleotide sequence of SEQ ID NO: 1 or a variant of the above-mentioned nucleotide sequence. Specifically, the polynucleotide has a nucleotide sequence having a sequence homology of 70% or more, more preferably 80% or more, still more preferably 90% or more, and most preferably 95% or more, with the nucleotide sequence of SEQ ID NO: 1 . ≪ / RTI > "% Of sequence homology to polynucleotides" is ascertained by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences (I. E., A gap) relative to the < / RTI >

Those skilled in the art will appreciate that as long as the nucleotide sequence that retains at least 70% homology by such an artificial modification retains the light harvesting activity for gene expression desired in the present invention, the GAF2 polynucleotide of the present invention And the scope of the invention is not limited thereto.

In one embodiment of the present invention, the GAF2 polynucleotide may be derived from a microorganism , and more particularly, may be derived from a microcoleus sp., And more particularly, Microcoleus sp. B353, Lt; / RTI >

Such GAF2 polynucleotides may have a light harvesting pigment function to absorb the orange color of the 600 to 605 nm wavelength band.

The present invention also provides a light harvesting dye GAF2 domain having the amino acid sequence of SEQ ID NO: 2.

In the present invention, the GAF2 domain may include not only a wild type but also a functional homolog having a light harvesting activity with an amino acid homology of 90% or more.

As used herein, the term "homology" refers to a similar degree to the amino acid sequence of a wild type protein, wherein the light harvesting dye GAF2 domain (polypeptide) of the present invention comprises a wild-type amino acid as defined by the sequence of SEQ ID NO: 2 And more preferably 70% or more, preferably 90% or more, more preferably 95% or more. This comparison of homology can be performed using a visual program or a comparison program that is easy to purchase. Commercially available computer programs can calculate homology between two or more sequences as a percentage, and homology (%) can be calculated for adjacent sequences.

One of ordinary skill in the art will readily recognize that this artificial modification retains at least 90% homology and is an equivalent domain (polypeptide) as long as it retains light harvesting activity in the present invention.

The GAF2 domains (polypeptides) of the present invention may comprise amino acid sequence variants thereof as long as they retain light harvesting activity. By mutant herein is meant a protein having a sequence that differs by natural amino acid sequence and one or more amino acid residues by deletion, insertion, non-conservative or conservative substitution, or a combination thereof.

Such variants include functional homologues having substantially equivalent activity to the wild type or proteins having modifications that increase or decrease physicochemical properties. Preferably, the physicochemical properties of the protein are modified variants. For example, physical factors such as temperature, moisture, pH, electrolyte, reducing sugar, pressurization, drying, freezing, interfacial tension, lightning, repetition of freezing and thawing, high concentration conditions, and acid, alkali, neutral salt, organic solvent, , Oxidation-reduction agent, protease, and the like are increased in structural stability to the external environment. It may also be a mutant having increased activity due to a mutation in the amino acid sequence.

The GAF2 domain (polypeptide) of the present invention can be prepared by directly isolating from Microcoleus sp. B353, chemically synthesized, or obtained using a gene recombination technique.

First, the GAF2 domain (polypeptide) of the present invention can be prepared by directly isolating from Microcoleus B353. The separation and purification of the GAF2 domain (polypeptide) of the present invention contained in the cells can be carried out by a number of known methods.

The GAF2 domain (polypeptide) of the present invention can be chemically synthesized. When chemically synthesized, it can be obtained by using a polypeptide synthesis method well known in the art. Polypeptides can be prepared using conventional stepwise liquid or solid phase synthesis, fractional condensation, F-MOC or T-BOC chemistry. In particular, the preferred method of preparing the polypeptide is by using solid phase syntheses. The GAF2 domain (polypeptide) can be synthesized by a condensation reaction between the protected amino acids in a conventional solid-phase method, starting from the C-terminus and progressing sequentially according to the sequence. After the condensation reaction, the protecting group and the carrier to which the C-terminal amino acid is linked can be removed by a known method such as acid decomposition or aminolysis. The above-mentioned polypeptide synthesis methods are described in detail in the relevant publications.

The GAF2 domain (polypeptide) of the present invention may also be obtained using a gene recombination technique. When a gene recombination technique is used, a polynucleotide (nucleic acid) encoding the GAF2 domain (polypeptide) of the present invention is inserted into an appropriate expression vector, and the vector is transformed into a host cell to transform the GAF2 domain protein of the present invention into a host cell , And then recovering the protein from the host cell. The protein may be expressed in a selected host cell and then subjected to conventional biochemical separation techniques such as treatment with a protein precipitant (salting-out method), centrifugation, ultrasonic disruption, ultrafiltration, dialysis, (Gel filtration), adsorption chromatography, ion exchange chromatography, affinity chromatography, and the like. In order to separate proteins having high purity, they are used in combination.

The present invention also provides a light harvesting dye GAF3 polynucleotide having the nucleotide sequence of SEQ ID NO: 3 or a variant of the above base sequence. Specifically, the polynucleotide has a nucleotide sequence having a sequence homology of 70% or more, more preferably 80% or more, still more preferably 90% or more, and most preferably 95% or more, with the nucleotide sequence of SEQ ID NO: 3 . ≪ / RTI >

In one embodiment of the present invention, the GAF3 polynucleotide may be derived from a bacterium belonging to the genus Microcleus , and more specifically, may be derived from a microcoleus sp., And more particularly, Microcoleus sp. B353, Lt; / RTI >

Such GAF3 polynucleotides may have a light harvesting pigment function to absorb orange color of 600 to 605 nm wavelength band.

The present invention also provides a light harvesting dye GAF3 domain (polypeptide) having the amino acid sequence of SEQ ID NO: 4.

In the present invention, the GAF3 domain (polypeptide) may include not only a wild type but also a functional homolog having a light harvesting activity with an amino acid homology of 90% or more.

The present invention also provides a light-harvesting dye GAF4 polynucleotide having the nucleotide sequence of SEQ ID NO: 5 or a variant of the above-mentioned nucleotide sequence. Specifically, the polynucleotide has a nucleotide sequence having a sequence homology of 70% or more, more preferably 80% or more, still more preferably 90% or more, and most preferably 95% or more, with the nucleotide sequence of SEQ ID NO: 5 . ≪ / RTI >

In one embodiment of the present invention, the GAF4 polynucleotide may be derived from a bacterium belonging to the genus Bacillus , and more specifically, may be derived from a microcoleus sp., More specifically, Microcoleus sp. B353, Lt; / RTI >

The GAF4 polynucleotide of the present invention may have a photoreactive function of ultraviolet / green light.

In the present invention, the optical reversible function of the ultraviolet / green light means a function capable of absorbing green light upon absorption of ultraviolet light, accompanied by a type conversion of light returning to a state capable of absorbing ultraviolet light again upon absorption of green light do.

The GAF4 polynucleotide may also have an auxiliary pigment function capable of transferring the absorbed energy to other pigment molecules.

The present invention also provides a light harvesting dye GAF4 domain (polypeptide) having the amino acid sequence of SEQ ID NO: 6.

In the present invention, the GAF4 domain (polypeptide) may include not only a wild type but also a functional homolog having a light harvesting activity with an amino acid homology of 90% or more.

The present invention also provides an ultraviolet / green light harvesting dye (GHP) gene having the nucleotide sequence of SEQ. ID. NO: 9 including all of the GAF2, GAF3 and GAF4 genes, or a mutant of the above nucleotide sequence. Specifically, the gene has a nucleotide sequence having a sequence homology of at least 70%, more preferably at least 80%, further preferably at least 90%, most preferably at least 95% with the nucleotide sequence of SEQ ID NO: 9 .

The ultraviolet / green light harvesting dye (GHP) gene of the present invention may be derived from a microorganism belonging to the genus Soc ., And may be derived from Microcoleus sp., More specifically Microcoleus sp. B353 ). ≪ / RTI >

The present invention also provides an ultraviolet / green light harvesting pigment (GHP) protein having the amino acid sequence of SEQ ID NO: 10, including both the GAF2, GAF3 and GAF4 domains.

In the present invention, the ultraviolet / green light harvesting dye (GHP) protein may include not only a wild type but also a functional homolog having a light harvesting activity with an amino acid homology of 90% or more.

The present invention also provides a recombinant vector comprising the above ultraviolet / green light harvesting dye (GHP) gene.

The ultraviolet / green light harvesting dye (GHP) gene of the present invention can be provided by a vector expressing it and expressed as a protein.

The term "vector" as used herein refers to a DNA construct containing a base sequence of a gene operably linked to a suitable regulatory sequence so as to be capable of expressing the gene of interest in a suitable host, said regulatory sequence being capable of initiating transcription A promoter, any operator sequence for modulating such transcription, a sequence encoding a suitable mRNA ribosome binding site, and a sequence controlling the termination of transcription and translation.

In the present invention, "operably linked" refers to a functional linkage between a nucleic acid expression control sequence and a nucleic acid sequence encoding a desired protein to perform a general function. For example, a nucleic acid sequence encoding a promoter and a protein or RNA may be operably linked to affect the expression of the coding sequence. The operative linkage with the recombinant vector can be produced using genetic recombination techniques well known in the art, and site-specific DNA cleavage and linkage are made using enzymes generally known in the art.

The vector of the present invention may include expression regulatory elements such as a promoter, an initiation codon, a stop codon, a polyadenylation signal and an enhancer, a secretion signal, and the like, and may be variously manufactured according to the purpose. The initiation codon and the termination codon must be operative in the individual when the gene construct is administered and in the coding sequence and in frame.

The vector used in the present invention is not particularly limited as long as it is replicable in a host, and any vector known in the art can be used. For example, LMG-pPL-PCB, which is a plasmid, cosmid, phage particle, viral vector, or simply a potential genome insert, and preferably E. coli for protein expression that forms a picocyananolchromophore can be exemplified, But is not limited thereto. The vector may be transcribed into a suitable host, then replicated or functional, independent of the host genome, and integrated into the genome itself.

In addition, the vector of the present invention may further include a selection marker. The selection marker is used to select a cell transformed with a vector, that is, to confirm whether a target gene has been inserted, and to give a selectable phenotype such as resistance to drug resistance, nutritional requirement, tolerance to cytotoxic agent, or surface protein expression May be used. In the environment treated with the selective agent, only the cells expressing the selection marker survive or express different phenotypes, so that the transformed cells can be selected.

The present invention also provides a transformant transformed with said vector. The transformant of the present invention can be constructed by introducing the vector into the host cell in such a manner that the promoter can function.

The term "transformed" in the present invention means that DNA is introduced into the host and the DNA becomes replicable as an extrachromosomal factor or by chromosome integration completion. Transformation includes any method of introducing a nucleic acid molecule into an organism, cell, tissue or organ, and can be carried out by selecting a suitable standard technique depending on the host cell as is known in the art. Such methods include electroporation, CaPO4 precipitation, CaCl2 precipitation, microinjection, polyethylene glycol (PEG), DEAE-dextran, cationic liposomes, and lithium acetate- DMSO, and the like, but are not limited thereto.

Since the expression amount of the protein and the expression are different depending on the host cell transformed with the expression vector, the host cell most suitable for the purpose may be selected and used. The host cells that can be used in the present invention include yeast, fungi, bacteria, or algae. For example, Escherichia coli), Bacillus subtilis (Bacillus subtili s), Streptomyces (Streptomyces), Pseudomonas (Pseudomonas), Proteus Mira Billy's (Proteus mirabilis), Staphylococcus (Staphylococcus), Aspergillus way Russ (Aspergillus), Paracoccus (Paracoccus), Flavobacterium (Flavobacterium), Agrobacterium (Agrobacterium), alkali jeneseu (Alcaligenes), El Winiah (Erwinia), blood Chiapas pastoris (Pichia pastoris), Saccharomyces access to my Celebi jiae (Saccharomyces cerevisiae), investigating car break in Rome Seth (Schizosaccharomyces), Castello La Neuro Chrysler Corporation (Neurospora crassa ). Preferably, Escherichia coli ), but is not limited thereto.

The present invention also provides a method for introducing the recombinant vector into a plant to increase the utilization efficiency of photosynthesis.

In one embodiment of the present invention, the plant can not use ultraviolet light or green light as a light source for photosynthesis, and in detail, it can be used as a light source such as an antiseptic, Plants and the like.

Examples of the species of mosses that do not have the above-mentioned picoeritrin as a light harvesting dye include Synechocystis sp., Synechococcus sp., And Prochlorococcus sp .; Examples of the green algae having the chlorophyll b as an auxiliary dye include Chlamydomonas, Chlorella, Nannochloropsis, and Ettlia; Examples of the land higher plants include corn, sugar cane, cotton, tobacco, rice, barley and potato, but the present invention is not limited thereto.

In the present invention, the efficiency of utilization of the photosynthetic activity can be enhanced by using ultraviolet light and green light as a light source for photosynthesis through the introduction of ultraviolet / green light harvesting dye (GHP) gene, and specific examples thereof will be described with reference to the following examples.

Advantages and features of the present invention and methods of achieving them will become apparent with reference to the embodiments described in detail below. Hereinafter, the present invention will be described in detail with reference to examples. However, these examples are for illustrating the present invention specifically, and the scope of the present invention is not limited to these examples.

< Example >

&Lt; Materials and methods >

One. Microcore  B353 strain culture

Oscillatoriales Neck Microcoleus sp. B353 is a filamentous fungus collected from the carbonate lake of Khilganta, Russia. Unlike most cyanobacteria distributed in freshwater or seawater, it can grow in high alkali and high salt environments. The inventors of the present invention found that Microcoleus sp. IPPAS-B353 was obtained from the Institute of Plant Physiology, Russian Academy of Science, Moscow (GenBank ac.No. DQ222209), Russian Academy of Plant Physiology.

Micro Collet mouse B353 (B353 Microcoleus) strain is a white light (20 μmolm-2s-1) agar medium with 30 ℃ temperature conditions in the (KCl 1.0g / L; NaHCO3 16.8g / L; K2HPO4 0.5g / L; NaNO3 2.5g / L; K2SO4 1.0 g / L; NaCl 30 g / L; MgSO4 0.1 g / L; CaCl2 0.04 g / L; FeSO4 0.01 g / L; EDTA 0.08 g / L; Trace metal (A5 + Co) % Agar).

2. Genome DNA  Separation and ultraviolet / Green light  Harvest coloring ( GHP ) GAF  domain Cloning

For genomic DNA isolation, the cells were harvested by centrifugation (3,500 rpm, 15 min, 4 ° C) of the microcoleus B353 cells of the exponential growth period, and then 200 μl of TEN buffer (10 mM Tris-Cl, 150 mM NaCl, 100 mM EDTA). 10 μl lysozyme (20 mg / ml) was added thereto, and the cells were disrupted by reacting at 37 ° C for 15 minutes, followed by addition of 5 μl of 10% SDS and shaking. 5 μl of protease K (20 μg / ml) was added to the reaction mixture, and reacted at 60 ° C for 1 hour. 400 μl of phenol was added thereto, and the supernatant was transferred to a new tube by centrifugation (3,500 rpm, 3 minutes at room temperature). The same amount of phenol: chloroform: isoamyl alcohol as the supernatant was added and the mixture was stirred at room temperature for 3-5 minutes. After centrifugation (3,500 rpm, 3 minutes, room temperature), the supernatant was mixed with 20 μl of 3M sodium acetate (pH 5.2) and 400 μl of ethanol, and centrifuged (3,500 rpm, 3 min, .

In order to clone the GAF domain of the ultraviolet / green light harvesting dye (GHP), primers (see Table 1 below) were first prepared so that the protein sequences of each GAF domain could be conserved. At this time, restriction enzyme recognition sites Respectively. The restriction enzymes used were selected not to be present in the nucleotide sequence to be cloned, and XhoI and EcoRI were used in this study. Each primer was used to obtain a fragment of each base sequence that can encode each GAF domain through polymerase chain reaction. Subsequently, subcloning was performed on DH5a E. coli to obtain a recombinant plasmid in which the base sequences of the respective GAF domains obtained by the polymerase chain reaction with the vector pBAD / mycHisC (Invitrogen, USA) were ligated (see FIG. 7). Then, the plasmid was extracted and it was confirmed whether the nucleotide sequence was correctly inserted into the plasmid prepared by the sequencing method. After final confirmation that the desired sequence was conserved, it was transformed into E. coli (LMG-pPL-PCB) for protein expression to form phycocyanobilin (briefly referred to as PCB) chromophores (Gambetta , GA and JC Lagaris, 2001. Genetic engineering of phytochrome biosynthesis in bacteria, Proceedings of the National Academy of Science, USA, 98, 10566-10571).

The primer sequences used in PCR gene Primer sequence SEQ ID NO: GAF2 Forward Gt; 11 Reverse Gt; 12 GAF3 Forward Gt; 13 Reverse Gt; 14 GAF4 Forward 5'-CATCTCGAGTCGGGAATCTCTGAACTTAG-3 ' 15 Reverse 5'-CATCTGCAGCGGGAATCTCTGAACTTAG-3 ' 16 GAF2 / 3/4 Forward Gt; 17 Reverse 5'-CATCTGCAGCGGGAATCTCTGAACTTAG-3 ' 18

3. PCB  produce In E. coli  UV-rays/ Green light  Harvest coloring ( GHP ) GAF  Domain expression and pure isolation

Escherichia coli for the expression of the transfected protein was cultured in 100 ml of RM liquid medium (2% casamino acids; 1X M9 salts; 1 mM MgCl2; 0.2% glucose; Thiamine 0.1 mM) supplemented with antibiotic ampicillin (200 ug / L) and kanamycin And the cells were inoculated at OD600 = 0.1 and cultured at 37 DEG C until OD600 = 0.4-0.6. To the 500 mL of the LB medium, 100 mL of the RM medium was added and the cultured E. coli for expression of the protein was mixed. Then, 25 μM of FAC, 25 μM of ALA, 0.1 mM of IPTG, 200 μg / L of ampicillin and 50 μg / L of kanamycin were added thereto. Time. After incubation at 28 ° C for 5 hours with 0.2% Arabinose, E. coli for protein expression was harvested by centrifugation at 7000 rpm at 4 ° C and cells were frozen at 70 ° C. All the procedures for protein extraction were carried out at 4 ° C under dark conditions. To the 20 mL of cell lysis buffer (50 mM Tris pH 7.85, 100 mM NaCl, 10% Glycerol, 0.1% NP40), 0.5 tablets of Complete, EDTA-free protease inhibitor (Roche, Switzerland) and 0.1 mM DDT were added, Escherichia coli for protein expression was disrupted with a crusher, and the supernatant was transferred to a new tube by centrifugation at 15000 rpm for 10 minutes at 4 ° C. To the supernatant, 500 μl of Ni-NTA agarose (Quiagen, Germany) beads was added and mixed with protein for 1 hour and 30 minutes at 4 ° C. Centrifuge at 3500 rpm, 4 ° C for 2 minutes to submerge the protein-bound bead and remove the supernatant. The bound beads were transferred to Poly-Prep chromatography column (Bio-Rad, USA) and 10 mL of wash buffer (50 mM Tris pH 7.85, 300 mM NaCl, 10% Glycerol, 20 mM imidazole, 0.1% NP40) . Proteins were extracted by passing 1.5 mL of Eultion buffer (50 mM Tris pH 8.0, 300 mM NaCl, 250 mM imidazole) and centrifuged using Amicon Ultra centrifugal filter units 10K membrane (Millipore, USA) to obtain concentrated proteins. Separated pure proteins were used to measure absorption and fluorescence spectra.

4. Electrophoresis and Zn ^{2 +} Blotting

12% SDS-PAGE gels were prepared, and 3 μl of the separated purified protein and 1 μl of 4 × sample buffer were mixed and heated at 100 ° C. for 5 minutes. 4ul loading per lane and 2h electrophoresis at 120mV. Zn2 + blotting was performed in 20 mM Tris pH 7.35, 20 mM Zinc acetate solution for 5 minutes under dark condition, and the band was confirmed under UV. The cells were stained with Coomassie Brilliant Blue Solution (CBB) for 1 hour and then reacted for 12 hours or more with a de-staining solution.

5. Absorption spectrum  Measure

UV-vis spectrophotometer S-3100 (Scinco, Korea) was used to measure the spectra absorbed in the range of 190 to 1100 nm using LabPro software. Initial values were set in Tris pH 8.0 solution. After calibrated at a value of 750 nm, the absorption spectrum within 300 to 800 nm was drawn by Origin 8.0.

6. Room temperature fluorescence measurement

Fluorescence spectrometer Fluorescence spectra were measured using LS-55 (Perkin Elmer, USA). The excitation at 560 nm and the emission spectrum emitted between 500 and 750 nm were measured. The initial value was set to Tris pH 8.0. After calibrating at a value of 750 nm, the fluorescence spectrum was plotted with Origin 8.0.

< Example  1>

Microcore  From B353 New  UV-rays/ Green light  Harvest coloring ( GHP ) Identification of genes

The present inventors analyzed a novel genome of Microcoleus B353. Based on the results of the genome analysis, Synechocystis sp. A gene having homology to Cph1 and ETR1 of PCC6803 was BLAST-screened.

As a result, 21 homologous genes could be detected. Among them, UV / green light harvesting pigment (GHP) protein consisted of 4 GAF domains and 1 methyl accepting chemotaxis protein (MCP) I could confirm. Also, the GHP protein was confirmed by the following experiment that three domains out of four GAF domains had photoreceptor functions, and these were designated as GAF2, GAF3, and GAF4, respectively (see FIG. 1).

Also, for reference, in the present invention, For the GAF2, GAF3 and GAF4 domains, the nucleotide sequence (polynucleotide) for GAF2 is shown in SEQ ID NO: 1 and the GAF2 polypeptide sequence is shown in SEQ ID NO: 2. The nucleotide sequence (polynucleotide) of GAF3 is represented by SEQ ID NO: 3, the GAF3 polypeptide sequence is represented by SEQ ID NO: 4, the nucleotide sequence for GAF4 (polynucleotide) is represented by SEQ ID NO: 5 and the polypeptide sequence of GAF4 is represented by SEQ ID NO: Number 6. The nucleotide sequence (polynucleotide) for GAF / 2/3/4 is shown in SEQ ID NO: 7, and the polypeptide sequence for GAF / 2/3/4 is shown in SEQ ID NO:

In addition, in the present invention, the sequence of an ultraviolet / green light harvest pigment (GHP) protein consisting of four GAF domains including GAF2, GAF3 and GAF4 and one methyl accepting chemotaxis protein (MCP) The polynucleotide sequence of GHP is shown in SEQ ID NO: 9, and the polypeptide of GHP is shown in SEQ ID NO: 10.

< Example  2>

Microcore  B353 strain-derived ultraviolet / Green light  Harvest coloring ( GHP ) Are ultraviolet, green and orange absorption pigments

As a result of the amino acid sequence alignment analysis, it was found that it belongs to the cyanobacteriochromes (CBCRs) subfamily having one conserved cysteine residue in the 2, 3 and 4 GAF domains except the 1 GAF domain. In general, the first cysteine residues are known to have a CH motif with histidine (Rockwell, NC, and Lagarias, JC (2010) ChemPhysChem 11, 1172-1180) As a result, it was confirmed that GAF 2 and GAF 3 had a CH motif as expected, but GAF4 was a CY variant having tyrosine instead of histidine (see FIG. 1).

Furthermore, in order to test the importance of the cysteine residues of the CH and CY motifs to the light-sensing activity of the GAF domain, the present inventors have found that GAF2, GAF3, GAF4 and the three GAF domains RTI ID = 0.0 &gt; GAF2 / 3/4 &lt; / RTI &gt; Most GAF domains were blue in the native 15Z form. The GAF2 / 3/4 protein, which expressed GAF2, GAF3 and GAF4 domains simultaneously and three domains simultaneously, was colorless (15Z form) in cancer condition and light pink (15E form) when exposed to UV It was observed that a changing light circulation occurred (see Fig. 2). The separated proteins were electrophoresed on 12% SDS-PAGE gel, and CBB staining and Zinc blotting were performed. As a result, it was confirmed that PCBs were bonded to the three kinds of GAF proteins (see FIG. 3).

< Example  3>

Microcore  B353 strain-derived Light harvest  Pigment GAF4  Domains are UV / Green Optically reversible Cyanobacte Identify Rio Chrome

Absorption spectra were measured using a UV-spectrophotometer in order to confirm the absorption of light of a certain wavelength as a photoreceptor by concentrating the separated protein at a high concentration. GAF2 and GAF3 (native 15Z) in the ground state absorbed wavelengths of around 600-605 nm and gave wavelengths near 600 nm, but the absorption wavelength regions remained unchanged. On the other hand, GAF4 having a CY residue showed a peak at 330 nm and 386 nm in the ground state (native 15Z), and changed into a form capable of absorbing light at 560 nm (native 15E) when irradiated with UV (~ 380 nm). It was confirmed that when this light is again irradiated with 560 nm light, it returns to a ground state (native 15Z) capable of absorbing 330 nm and 386 nm (see FIG. 4A). Therefore, only GAF4 among the three GAF domains was found to be capable of circulating UV / green light.

In the acidic conditions of the GAF domain, denaturation studies were performed to identify the combined chromophore structure. When transformed into acidic factor (8M urea / hydrochloric acid, pH 2.0) in the dark state, no photoconversion activity was observed in GAF domains 2 and 3, but phototransformation activity was observed in GAF4 and GAF2 / 3/4. The ultraviolet absorbing type was transformed into a type absorbing green light having a peak at 603 nm (see FIG. 4B). Irradiation of orange or green light on the denatured sample revealed that the denatured Pg state was photoactive (see FIG. 4B). The green light-minus dark difference spectrum of denatured form of GAF3 and GAF2 / 3/4 recombinant GAF is identical to that of acid denatured Cph1 (see FIG. 4C), indicating that the PCB is a chromophore of GAF4 it means. Taken together, this means that GHP is a PCB and thioether-coupled photoreceptor through the GAF4 domain and that the chromophore of m GAF2 and 3 is detected by Zn2 + -dependent fluorescence technique, .

< Example  4>

Microcore  B353 strain-derived ultraviolet / Green light  Harvest coloring ( GHP ) GAF4  Absorbed in the domain Green light Is energy GAF2  Or 3

When GAF4 absorbed UV, the fluorescence spectra of each protein were measured by focusing on the overlap of GAF2 and GAF3 with the absorption region. The light energy delivered to each protein is absorbed and emits redshifted fluorescence. Fluorescence spectra were obtained by quantifying the same number of moles of protein (when excited at 560 nm), the amount of fluorescence emitted by GAF2 and GAF3 was about 20 FU, and no light conversion occurred. However, GAF4 showed a low value of less than 10FU when absorbing UV (15Z), but about 40FU when absorbing 580nm (15E). These results indicate that the efficiency of emission of fluorescence by the light energy absorbed by the Z and E type GAF4 proteins is different. GAF2 / 3/4 (protein with GAF2, 3 and 4 all linked) showed an absorption peak at 603 nm in the form of a 15Z state at the bottom, and a form (15E) in which UV light could absorb GAF4 at 560 nm (Fig. 4D). In the fluorescence spectrum, similarly to the shape of GAF4, when the UV was irradiated, it changed into a form capable of absorbing the 560 nm region (15E), and thus the fluorescence Which is about twice as high as that of the conventional method. Notably, fluorescence values were emitted to the same extent as GAF4, although the absorption spectrum of GAF2 / 3/4 could absorb 560 nm. On the other hand, when GAF2 / 3/4 in the GAF2 / 3/4 was able to absorb 560nm in the GAF2 / 3/4, GAF2 / 3/4 was released in a similar amount to GAF2 and 3 in the GAF2 / 3 / Respectively. This is different from the fact that the peak of the fluorescence spectrum of GAF4 is shifted to the left compared to GAF2, 3 at about 640 nm. Therefore, when GAF2 / 3/4 are together, GAF2, 3 is thought to absorb light emitted from GAF4 when GAF4 becomes a form (15E) capable of absorbing 560nm by UV. These results are also shown in the difference spectra of denaturation of each protein structure using 8M Urea. Of the three GAF domains, only GAF4 was associated with PCBs indicating photochromic activity (FIG. 4B). Therefore, the CY residue of GAF4 forms a thioester bond with the C3 Cystein of the PCB chromophore. When UV is irradiated, it transforms the C15 of the chromophore into the cis form, which is a form (15E) capable of absorbing light of 560 nm, On the main surface, C15 is transformed into a trans form and converted into a form (15Z) capable of absorbing UV light. Therefore, MBR3854 was able to activate GAF2, 3 when UV was detected in GAF4, and it was found that MBR3854 could act to transmit signals (Fig. 5).

< Example  5>

Microcore GAF4  Of the domain Of the CY moiety Tyrosine silver On light conversion  Essential amino acid The residue  Identification

In the GAF4 domain, a cysteine residue capable of binding to chromophores is present at positions 713 and 768. [ In the previous study, the motif of the chromophore binding of the GAF domain was a well conserved cysteine residue at 741 and 768th. On the other hand, the GAF4 domain of the GHP protein has a tyrosine residue in place of cysteine at position 741 and a cysteine residue at position 713 in the site where about 30 amino acids are not found in other GAF domains. These results suggest that two cysteine residues are involved in chromophore binding and light circulation by green / ultraviolet light.

To investigate the major amino acid residues involved in the chromophore binding and photoconversion of the GAF4 domain, studies were carried out to replace each amino acid residue in a known motif. There are a total of three cysteine residues in GAF4. When the 672nd cysteine residue was replaced with alanine (Fig. 5B), the zinc-dependent reaction and absorption spectrum showed the same results as GAF4. On the other hand, when the 674th tyrosine was converted to histidine (FIG. 5C), it was confirmed that it binds to chromophor through a zinc-dependent reaction, but the absorption spectrum showed no significant absorption peak. These results indicate that the portion containing 672 th cysteine and 674 th tyrosine is a novel motif that plays an important role in the absorption of the GAF domain. When the 713th cysteine residue in the newly proposed insertion sequence was replaced with alanine (Fig. 5D), the absorption spectrum of the PCB showed a maximum absorption peak at 347 nm and 623 nm. This was very different when compared to the absorption region of GAF4 and no light conversion was observed. Based on these results, it is judged that 713th cysteine determines the absorption region of GAF4 domain and plays an important role in light conversion. GAF4 has a tyrosine residue at position 741, but the cysteine residues at the same position are well conserved in the various GAF domains identified in previous studies. When the 741th tyrosine was converted to cysteine (FIG. 5E), the absorption spectrum of the region similar to GAF4 was shown. However, when the UV was changed to 15E by 2 minutes, the region below 400 nm showed a high absorption spectrum. Irreversible absorption spectra were obtained that did not convert to 15Z form when given. On the other hand, when the tyrosine at the same position was substituted with phenylalanine (Fig. 5F), the result was the same as that of the absorption spectrum of GAF4. Thus, the 741th tyrosine is considered to play an important role in the light conversion of GAF4, and the GAF domain having 713th cysteine is also conserved as tyrosine residue at 741th place in the GAF domain of other S.coli. When cysteine 768 was converted to serine (Fig. 5G), no chromopore was bound and no absorption spectrum was observed. When tyrosine 769 was substituted with histidine (FIG. 5H), chromopore was bound, but the absorption spectrum measurement result was very different from that of GAF4 and no light conversion was observed. Therefore, GAF domain having 713 cysteine residues was thought to be able to perform its function although it had to have tyrosine residues at 741 and 769.

To date, the chromophore binding of the GAF4 domain of the GHP protein has been shown to be mediated by cysteine 768, while cysteine 713 and tyrosine 769 are thought to be involved in the absorption domain. It should be noted that residues other than cysteine must be located at position 741 and it is very conserved in tyrosine in many species of S.coli. In addition, mutant studies of 672 cysteine and 674 tyrosine, which have not been studied so far, confirm that there are new motifs involved in light absorption and conversion of the GAF domain.

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

cph I: cyanobacteria phytochrome I
cph II: cyanobacteria phytochrome II
plpA: cyanobacteria phytochrome Ⅲ
GHP: UV / Green Light Harvesting Pigment
PCB: phycocyanobilin

<110> The Industry & Academic Cooperation in Chungnam National University (IAC) <120> Light harvesting pigment from cyanobacterium and use thereof <130> PN1212-588 <160> 18 <170> Kopatentin 2.0 <210> 1 <211> 558 <212> DNA <213> Light harvesting pigment GAF2 polynucleotide sequence from Microcoleus sp. B353 <400> 1 cgggtgattg agcggattcg tcaatccctg aatatcgata ttatcttccg caccaccaca 60 aaggaagtcc ggttactgct gaaatgcgat cgcgtggccg tctatcaatt taaccccgac 120 tttagcggtc agtttgtggc tgagtccgtc acctccggct gggtgaaact cgttggcgag 180 gaggtcgaac gggtttggaa agatacctac ctcgaagaaa accaaggggg acgctacgcc 240 aaccatgaaa cctacgccgt cagtgatatc tataccatcg gtcatgaccc ctgtcacgtc 300 gaactcctcg aacagttcga ggcccgggcc tataccctag ttcccatttt ccaaggagaa 360 catctctggg ggattctcgc cgcctatcaa aacgacggtc cccgggaatg gcaaaatagc 420 gaagtggatt tgctgcaacg gattgccgac caattggggg tggccctgca acaagccgaa 480 accgtgaccc aactgcgtcg tcagtccgat cgcattcgcc aagccgccga gcgcgatcgc 540 accgtagccc aagtcatc 558 <210> 2 <211> 186 <212> PRT <213> Light harvesting pigment GAF2 polypeptide sequence from Microcoleus sp. B353 <400> 2 Arg Val Ile Glu Arg Ile Arg Gln Ser Leu Asn Ile Asp Ile Ile Phe   1 5 10 15 Arg Thr Thr Lys Glu Val Arg Leu Leu Leu Lys Cys Asp Arg Val              20 25 30 Ala Val Tyr Gln Phe Asn Pro Asp Phe Ser Gly Gln Phe Val Ala Glu          35 40 45 Ser Val Thr Ser Gly Trp Val Lys Leu Val Gly Glu Glu Val Glu Arg      50 55 60 Val Trp Lys Asp Thr Tyr Leu Glu Glu Asn Gln Gly Gly Arg Tyr Ala  65 70 75 80 Asn His Glu Thr Tyr Ala Val Ser Asp Ile Tyr Thr Ile Gly His Asp                  85 90 95 Pro Cys His Val Glu Leu Leu Glu Gln Phe Glu Ala Arg Ala Tyr Thr             100 105 110 Leu Val Pro Ile Phe Gln Gly Glu His Leu Trp Gly Ile Leu Ala Ala         115 120 125 Tyr Gln Asn Asp Gly Pro Arg Glu Trp Gln Asn Ser Glu Val Asp Leu     130 135 140 Leu Gln Arg Ile Asp Gln Leu Gly Val Ala Leu Gln Gln Ala Glu 145 150 155 160 Thr Val Thr Gln Leu Arg Arg Gln Ser Asp Arg Ile Arg Gln Ala Ala                 165 170 175 Glu Arg Asp Arg Thr Val Ala Gln Val Ile             180 185 <210> 3 <211> 570 <212> DNA <213> Light harvesting pigment GAF3 polynucleotide sequence from Microcoleus sp. B353 <400> 3 cgcatccgtc aatccctcga tatcaacagt atcttcaacg tcaccaccaa agaagtacga 60 ctgttgctgc aatgcgatcg catcgccgtc tatcaattca accccgactt tagcggtcag 120 tttgtggccg agtccgtcac tcccggctgg gggaaactcg tcggccccga agtcgaacga 180 gtctggaaag acacctacct cgaagaaaac cagggaggac gttatgccta ccgggaaacc 240 tacgccgtca ccgatatcta cgaagccggc catgacccct gtcacgtcga actcctcgaa 300 cagttcgagg cccgggccta taccctggtt cctattttcc aaggggaaac cctctggggc 360 attctcgccg cctatcaaaa cgacggccct cgggaatggc aagaggacga agtgcaactg 420 cttgtacgca tcgccgacca attaggggtc gccctgcaac aagccgaaac cgtccaggaa 480 ctgcgccgcc agtccgaacg aattgcccgc accgccgaac gggaacgcgc cgtcgctcag 540 gtcatcgaac ggattcggga atctctgaac 570 <210> 4 <211> 190 <212> PRT <213> Light harvesting pigment GAF3 polypeptide sequence from Microcoleus sp. B353 <400> 4 Arg Ile Arg Gln Ser Leu Asp Ile Asn Ser Ile Phe Asn Val Thr Thr   1 5 10 15 Lys Glu Val Arg Leu Leu Leu Gln Cys Asp Arg Ile Ala Val Tyr Gln              20 25 30 Phe Asn Pro Asp Phe Ser Gly Gln Phe Val Ala Glu Ser Val Thr Pro          35 40 45 Gly Trp Gly Lys Leu Val Gly Pro Glu Val Glu Arg Val Trp Lys Asp      50 55 60 Thr Tyr Leu Glu Glu Asn Gln Gly Gly Arg Tyr Ala Tyr Arg Glu Thr  65 70 75 80 Tyr Ala Val Thr Asp Ile Tyr Glu Ala Gly His Asp Pro Cys His Val                  85 90 95 Glu Leu Leu Glu Gln Phe Glu Ala Arg Ala Tyr Thr Leu Val Pro Ile             100 105 110 Phe Gln Gly Glu Thr Leu Trp Gly Ile Leu Ala Ala Tyr Gln Asn Asp         115 120 125 Gly Pro Arg Glu Trp Gln Glu Asp Glu Val Gln Leu Leu Val Arg Ile     130 135 140 Ala Asp Gln Leu Gly Val Ala Leu Gln Gln Ala Glu Thr Val Gln Glu 145 150 155 160 Leu Arg Arg Gln Ser Glu Arg Ile Ala Arg Thr Ala Glu Arg Glu Arg                 165 170 175 Ala Val Ala Gln Val Ile Glu Arg Ile Arg Glu Ser Leu Asn             180 185 190 <210> 5 <211> 720 <212> DNA <213> Light harvesting pigment GAF4 polynucleotide sequence from Microcoleus sp. B353 <400> 5 cgggaatctc tgaacttaga caccatcttc gacaccacca ccaccgaagt ccgacgactc 60 ctcaaaactg atcgcgtctg tgtctatcaa ttcgccgaag actggagtgg gtcctttatt 120 gccgagtccg tcggggcgac ctgggccccc ctggtgcagc gacaacaaca aattcccgct 180 ctacgggaca atgtcagcca atgcgacaac atgacaagcc tgattgagcg gcaacagtcc 240 ggccaacttg ccccccaagc cagcggcccc gccagtcggg atacctatct gcaagagacg 300 gaagggggac gcttccagac tgaacaagcc ttctccgttg aagatatcta tcaagcgggc 360 tttagcacct gttatctcga agtcctcgaa caatatcaat gtcgcgccta tgccattgtt 420 cccatcttca agggtcctca actctgggga ctcctggccg cctatcagaa caacgggcca 480 cgggactggg aaaacgagga agtcacccta ctgcggcaaa tcggaaccca actgggagtc 540 gccattcagc agagtgaata cctgcaacaa ctgcaagagc agtccctgca actcgaagct 600 gctgcccaac gggataaggc cgccaaagaa caactgcaaa aacgggctct ggaactgctg 660 atggccgtcc gcccagccct ggacggggat ttaacagtgc ggattcccgt gacagaagat 720                                                                          720 <210> 6 <211> 240 <212> PRT <213> Light harvesting pigment GAF4 polypeptide sequence from Microcoleus sp. B353 <400> 6 Arg Glu Ser Leu Asn Leu Asp Thr Ile Phe Asp Thr Thr Thr Thr Glu   1 5 10 15 Val Arg Arg Leu Leu Lys Thr Asp Arg Val Cys Val Tyr Gln Phe Ala              20 25 30 Glu Asp Trp Ser Gly Ser Phe Ile Ala Glu Ser Val Gly Ala Thr Trp          35 40 45 Ala Pro Leu Val Gln Arg Gln Gln Gln Ile Pro Ala Leu Arg Asp Asn      50 55 60 Val Ser Gln Cys Asp Asn Met Thr Ser Leu Ile Glu Arg Gln Gln Ser  65 70 75 80 Gly Gln Leu Ala Pro Gln Ala Ser Gly Pro Ala Ser Arg Asp Thr Tyr                  85 90 95 Leu Gln Glu Thr Glu Gly Gly Arg Phe Gln Thr Glu Gln Ala Phe Ser             100 105 110 Val Glu Asp Ile Tyr Gln Ala Gly Phe Ser Thr Cys Tyr Leu Glu Val         115 120 125 Leu Glu Gln Tyr Gln Cys Arg Ala Tyr Ala Ile Val Pro Ile Phe Lys     130 135 140 Gly Pro Gln Leu Trp Gly Leu Leu Ala Ala Tyr Gln Asn Asn Gly Pro 145 150 155 160 Arg Asp Trp Glu Asn Glu Glu Val Thr Leu Leu Arg Gln Ile Gly Thr                 165 170 175 Gln Leu Gly Val Ala Ile Gln Gln Ser Glu Tyr Leu Gln Gln Leu Gln             180 185 190 Glu Gln Ser Leu Gln Leu Glu Ala Ala Ala Gln Arg Asp Lys Ala Ala         195 200 205 Lys Glu Gln Leu Gln Lys Arg Ala Leu Glu Leu Leu Met Ala Val Arg     210 215 220 Pro Ala Leu Asp Gly Asp Leu Thr Val Arg Ile Pro Val Thr Glu Asp 225 230 235 240 <210> 7 <211> 1836 <212> DNA <213> Light harvesting pigment GAF2 / 3/4 polynucleotide sequence from Microcoleus sp. B353 <400> 7 cgggtgattg agcggattcg tcaatccctg aatatcgata ttatcttccg caccaccaca 60 aaggaagtcc ggttactgct gaaatgcgat cgcgtggccg tctatcaatt taaccccgac 120 tttagcggtc agtttgtggc tgagtccgtc acctccggct gggtgaaact cgttggcgag 180 gaggtcgaac gggtttggaa agatacctac ctcgaagaaa accaaggggg acgctacgcc 240 aaccatgaaa cctacgccgt cagtgatatc tataccatcg gtcatgaccc ctgtcacgtc 300 gaactcctcg aacagttcga ggcccgggcc tataccctag ttcccatttt ccaaggagaa 360 catctctggg ggattctcgc cgcctatcaa aacgacggtc cccgggaatg gcaaaatagc 420 gaagtggatt tgctgcaacg gattgccgac caattggggg tggccctgca acaagccgaa 480 accgtgaccc aactgcgtcg tcagtccgat cgcattcgcc aagccgccga gcgcgatcgc 540 accgtagccc aagtcatcga ccgcatccgt caatccctcg atatcaacag tatcttcaac 600 gtcaccacca aagaagtacg actgttgctg caatgcgatc gcatcgccgt ctatcaattc 660 aaccccgact ttagcggtca gtttgtggcc gagtccgtca ctcccggctg ggggaaactc 720 gtcggccccg aagtcgaacg agtctggaaa gacacctacc tcgaagaaaa ccagggagga 780 cgttatgcct accgggaaac ctacgccgtc accgatatct acgaagccgg ccatgacccc 840 tgtcacgtcg aactcctcga acagttcgag gcccgggcct ataccctggt tcctattttc 900 caaggggaaa ccctctgggg cattctcgcc gcctatcaaa acgacggccc tcgggaatgg 960 caagaggacg aagtgcaact gcttgtacgc atcgccgacc aattaggggt cgccctgcaa 1020 caagccgaaa ccgtccagga actgcgccgc cagtccgaac gaattgcccg caccgccgaa 1080 cgggaacgcg ccgtcgctca ggtcatcgaa cggattcggg aatctctgaa cttagacacc 1140 atcttcgaca ccaccaccac cgaagtccga cgactcctca aaactgatcg cgtctgtgtc 1200 tatcaattcg ccgaagactg gagtgggtcc tttattgccg agtccgtcgg ggcgacctgg 1260 gcccccctgg tgcagcgaca acaacaaatt cccgctctac gggacaatgt cagccaatgc 1320 gacaacatga caagcctgat tgagcggcaa cagtccggcc aacttgcccc ccaagccagc 1380 ggccccgcca gtcgggatac ctatctgcaa gagacggaag ggggacgctt ccagactgaa 1440 caagccttct ccgttgaaga tatctatcaa gcgggcttta gcacctgtta tctcgaagtc 1500 ctcgaacaat atcaatgtcg cgcctatgcc attgttccca tcttcaaggg tcctcaactc 1560 tggggactcc tggccgccta tcagaacaac gggccacggg actgggaaaa cgaggaagtc 1620 accctactgc ggcaaatcgg aacccaactg ggagtcgcca ttcagcagag tgaatacctg 1680 caacaactgc aagagcagtc cctgcaactc gaagctgctg cccaacggga taaggccgcc 1740 aaagaacaac tgcaaaaacg ggctctggaa ctgctgatgg ccgtccgccc agccctggac 1800 ggggatttaa cagtgcggat tcccgtgaca gaagat 1836 <210> 8 <211> 612 <212> PRT <213> Light harvesting pigment GAF2 / 3/4 polypeptide sequence from Microcoleus sp. B353 <400> 8 Arg Val Ile Glu Arg Ile Arg Gln Ser Leu Asn Ile Asp Ile Ile Phe   1 5 10 15 Arg Thr Thr Lys Glu Val Arg Leu Leu Leu Lys Cys Asp Arg Val              20 25 30 Ala Val Tyr Gln Phe Asn Pro Asp Phe Ser Gly Gln Phe Val Ala Glu          35 40 45 Ser Val Thr Ser Gly Trp Val Lys Leu Val Gly Glu Glu Val Glu Arg      50 55 60 Val Trp Lys Asp Thr Tyr Leu Glu Glu Asn Gln Gly Gly Arg Tyr Ala  65 70 75 80 Asn His Glu Thr Tyr Ala Val Ser Asp Ile Tyr Thr Ile Gly His Asp                  85 90 95 Pro Cys His Val Glu Leu Leu Glu Gln Phe Glu Ala Arg Ala Tyr Thr             100 105 110 Leu Val Pro Ile Phe Gln Gly Glu His Leu Trp Gly Ile Leu Ala Ala         115 120 125 Tyr Gln Asn Asp Gly Pro Arg Glu Trp Gln Asn Ser Glu Val Asp Leu     130 135 140 Leu Gln Arg Ile Asp Gln Leu Gly Val Ala Leu Gln Gln Ala Glu 145 150 155 160 Thr Val Thr Gln Leu Arg Arg Gln Ser Asp Arg Ile Arg Gln Ala Ala                 165 170 175 Glu Arg Asp Arg Thr Val Ala Gln Val Ile Asp Arg Ile Arg Gln Ser             180 185 190 Leu Asp Ile Asn Ser Ile Phe Asn Val Thr Thr Lys Glu Val Arg Leu         195 200 205 Leu Leu Gln Cys Asp Arg Ile Ala Val Tyr Gln Phe Asn Pro Asp Phe     210 215 220 Ser Gly Gln Phe Val Ala Glu Ser Val Thr Pro Gly Trp Gly Lys Leu 225 230 235 240 Val Gly Pro Glu Val Glu Arg Val Trp Lys Asp Thr Tyr Leu Glu Glu                 245 250 255 Asn Gln Gly Gly Arg Tyr Ala Tyr Arg Glu Thr Tyr Ala Val Thr Asp             260 265 270 Ile Tyr Glu Ala Gly His Asp Pro Cys His Val Glu Leu Leu Glu Gln         275 280 285 Phe Glu Ala Arg Ala Tyr Thr Leu Val Pro Ile Phe Gln Gly Glu Thr     290 295 300 Leu Trp Gly Ile Leu Ala Ala Tyr Gln Asn Asp Gly Pro Arg Glu Trp 305 310 315 320 Gln Glu Asp Glu Val Glu Leu Leu Val Arg Ile Ala Asp Gln Leu Gly                 325 330 335 Val Ala Leu Gln Gln Ala Glu Thr Val Gln Glu Leu Arg Arg Gln Ser             340 345 350 Glu Arg Ile Ala Arg Thr Ala Glu Arg Glu Arg Ala Val Ala Gln Val         355 360 365 Ile Glu Arg Ile Arg Glu Ser Leu Asn Leu Asp Thr Ile Phe Asp Thr     370 375 380 Thr Thr Glu Val Arg Arg Leu Leu Lys Thr Asp Arg Val Cys Val 385 390 395 400 Tyr Gln Phe Ala Glu Asp Trp Ser Gly Ser Phe Ile Ala Glu Ser Val                 405 410 415 Gly Ala Thr Trp Ala Pro Leu Val Gln Arg Gln Gln Gln Ile Pro Ala             420 425 430 Leu Arg Asp Asn Val Ser Gln Cys Asp Asn Met Thr Ser Leu Ile Glu         435 440 445 Arg Gln Gln Ser Gly Gln Leu Ala Pro Gln Ala Ser Gly Pro Ala Ser     450 455 460 Arg Asp Thr Tyr Leu Gln Glu Thr Glu Gly Gly Arg Phe Gln Thr Glu 465 470 475 480 Gln Ala Phe Ser Val Glu Asp Ile Tyr Gln Ala Gly Phe Ser Thr Cys                 485 490 495 Tyr Leu Glu Val Leu Glu Gln Tyr Gln Cys Arg Ala Tyr Ala Ile Val             500 505 510 Pro Ile Phe Lys Gly Pro Gln Leu Trp Gly Leu Leu Ala Ala Tyr Gln         515 520 525 Asn Asn Gly Pro Arg Asp Trp Glu Asn Glu Glu Val Thr Leu Leu Arg     530 535 540 Gln Ile Gly Thr Gln Leu Gly Val Ala Ile Gln Gln Ser Glu Tyr Leu 545 550 555 560 Gln Gln Leu Gln Glu Gln Ser Leu Glu Leu Glu Ala Ala Ala Gln Arg                 565 570 575 Asp Lys Ala Ala Lys Glu Gln Leu Gln Lys Arg Ala Leu Glu Leu Leu             580 585 590 Met Ala Val Arg Pro Ala Leu Asp Gly Asp Leu Thr Val Arg Ile Pro         595 600 605 Val Thr Glu Asp     610 <210> 9 <211> 3555 <212> DNA <213> Light harvesting pigment GHP polynucleotide sequence from Microcoleus sp. B353 <400> 9 atgtctcctc gcgaatccac ctccccagcc acaaccaacc tctcggcccc cattgctaaa 60 agcttagaag aacaactcaa agcgattcgt cagcaggcgg ctaagattac ccaacccctc 120 caacaggcca acagcttaga ggcggcctgt cgggatgtgg tggtacaact tcagcaggtg 180 tttgagtgcg atcgcgccct catttaccgc ctgtgcgatc gcagtattga tgcctttatt 240 gaccaattcc aaaatcccat gtctggattg gccttcaatg gggggggaaa tggcgtcaat 300 cgtcctggtt taaatagcgc aattgaatct ggactgcctc taacaaatgc caatggtgcg 360 aatttaacgg ggactgtggt tgccgaagcc gtcggacgag gttggacacc atctttgggc 420 gatcgcctcc acattgccgc ctttggtttt gaaaccctag acgactatca aaacctgggg 480 tttatctcca tcgccgacca atctcgcacc aaccttaccc cacaccagca gcaaatcctc 540 gatcgctatc aagtccaagc ggccctaacc attcccctca aactgggtca aaaactttgg 600 ggggttttgt ctgtgcagca ttgtcgtaac ccctattcct ggtcagaacg agaaatcgcc 660 attttgcgac aagtggccgc cgaattaatt agtttccagc aaagtctcaa tggtcatggg 720 acttccatgc tagaccccaa cccccagcaa gttgaggatg tcctcgaacg gttgcgcaac 780 tcctttgacc gagttcaaga ccgcgatcgc agcgtagcac gggtgattga gcggattcgt 840 caatccctga atatcgatat tatcttccgc accaccacaa aggaagtccg gttactgctg 900 aaatgcgatc gcgtggccgt ctatcaattt aaccccgact ttagcggtca gtttgtggct 960 ggtccgtca cctccggctg ggtgaaactc gttggcgagg aggtcgaacg ggtttggaaa 1020 gatacctacc tcgaagaaaa ccaaggggga cgctacgcca accatgaaac ctacgccgtc 1080 agtgatatct ataccatcgg tcatgacccc tgtcacgtcg aactcctcga acagttcgag 1140 gcccgggcct ataccctagt tcccattttc caaggagaac atctctgggg gattctcgcc 1200 gcctatcaaa acgacggtcc ccgggaatgg caaaatagcg aagtggattt gctgcaacgg 1260 attgccgacc aattgggggt ggccctgcaa caagccgaaa ccgtgaccca actgcgtcgt 1320 cagtccgatc gcattcgcca agccgccgag cgcgatcgca ccgtagccca agtcatcgac 1380 cgcatccgtc aatccctcga tatcaacagt atcttcaacg tcaccaccaa agaagtacga 1440 ctgttgctgc aatgcgatcg catcgccgtc tatcaattca accccgactt tagcggtcag 1500 tttgtggccg agtccgtcac tcccggctgg gggaaactcg tcggccctga agtcgaacga 1560 gtctggaaag acacctacct cgaagaaaac cagggaggac gttacgccta ccgggaaacc 1620 tacgccgtca ccgatatcta cgaagccggc catgacccct gtcacgtcga actcctcgaa 1680 cagttcgagg cccgggccta taccctggtt cctattttcc aaggggaaac cctctggggc 1740 attctcgccg cctatcaaaa cgacggccct cgggaatggc aagaggacga agtgcaactg 1800 cttgtacgca tcgccgacca attaggggtc gccctgcaac aagccgaaac cgtccaggaa 1860 ctgcgccgcc agtccgaacg aattgcccgc accgccgaac gggaacgcgc cgtcgctcag 1920 gtcatcgaac ggattcggga atctctgaac ttagacacca tcttcgacac caccaccacc 1980 gaagtccgac gactcctcaa aactgatcgc gtctgtgtct atcaattcgc cgaagactgg 2040 agtgggtcct ttattgccga gtccgtcggg gcgacctggg cccccctggt gcagcgacaa 2100 caacaaattc ccgctctacg ggacaatgtc agccaatgcg acaacatgac aagcctgatt 2160 gagcggcaac agtccggcca acttgccccc caagccagcg gccccgccag tcgggatacc 2220 tatctgcaag agacggaagg gggacgcttc cagactgaac aagccttctc cgttgaagat 2280 atctatcaag cgggctttag cacctgttat ctcgaagtcc tcgaacaata tcaatgtcgc 2340 gcctatgcca ttgttcccat cttcaagggt cctcaactct ggggactcct ggccgcctat 2400 cagaacaacg ggccacggga ctgggaaaac gaggaagtca ccctactgcg gcaaatcgga 2460 acccaactgg gagtcgccat tcagcagagt gaatacctgc aacaactgca agagcagtcc 2520 ctgcaactcg aagctgctgc ccaacgggat aaggccgcca aagaacaact gcaaaaacgg 2580 gctctggaac tgctgatggc cgtccgccca gccctggacg gggatttaac agtgcggatt 2640 cccgtgacag aagatgaaat gggaaccctg gccgactcct tcaacaacac cctgcaaagc 2700 ctgcgtaaaa tcgtcatgca ggtgcaaacc gccgccggac aggtggtgca aagctcccaa 2760 agtagtgacg cgtctctgat ggaattgtca gaccaagcgc aaaagcaatt cgaggaaatt 2820 agcaaaaccc tcgatcgcat tcaggatgtg gtgggcttca cccaaaccac tgccaacaac 2880 gcccaacagg tggaagtggc gctacagggg gccaaccgta ccctagaagc tggggatgat 2940 gccatgaacc gtaccgtgga ggggattgcc gacattcgcc aaaccgtggc cgccgccgcc 3000 aaagggattg cccgccttac ccagtcctcg caaaacatcg ctaaggtggt gaacctgatt 3060 aacaacttcg ccactcagac caacttattg gcgttgaatg cggccattga agccacccga 3120 gccggggaat atgggcgcgg ctttgcggtg gttgccgatg aggtgcgctc cctggctcgt 3180 cagtcagcgg atgcgaccac cgaaattgaa aatctggtcc aggaaattca ggcagaaacg 3240 aaggatgtga ccctggcgat ggaaatggcc tcggaacagg tgacgaaggg gacaaatctg 3300 gtgggcgaaa cccggcaaac tctcaatgag attgtgacgg cgacggccca aatcggggaa 3360 ctggtgcagg gaattaccca tgcgacgcag gtacaaacgg aacaagctca atcggtgaca 3420 gagatgatgc gctcaattgc agaaattgcc aaccaaacct cggaaaactc catcgcgatt 3480 tcctcgcgct ttaaggatgc tctgaccaca gctcaggagt tacaatcaag tgcgggacaa 3540 tttaaggtga gttag 3555 <210> 10 <211> 1184 <212> PRT <213> Light harvesting pigment GHP polypeptide sequence from Microcoleus sp. B353 <400> 10 Met Ser Pro Arg Glu Ser Thr Ser Pro Ala Thr Thr Asn Leu Ser Ala   1 5 10 15 Pro Ile Ala Lys Ser Leu Glu Glu Gln Leu Lys Ala Ile Arg Gln Gln              20 25 30 Ala Ala Lys Ile Thr Gln Pro Leu Gln Gln Ala Asn Ser Leu Glu Ala          35 40 45 Ala Cys Arg Asp Val Val Val Gln Leu Gln Gln Val Phe Glu Cys Asp      50 55 60 Arg Ala Leu Ile Tyr Arg Leu Cys Asp Arg Ser Ile Asp Ala Phe Ile  65 70 75 80 Asp Gln Phe Gln Asn Pro Met Ser Gly Leu Ala Phe Asn Gly Gly Gly                  85 90 95 Asn Gly Val Asn Arg Pro Gly Leu Asn Ser Ala Ile Glu Ser Gly Leu             100 105 110 Pro Leu Thr Asn Ala Asn Gly Ala Asn Leu Thr Gly Thr Val Val Ala         115 120 125 Glu Ala Val Gly Arg Gly Trp Thr Pro Ser Leu Gly Asp Arg Leu His     130 135 140 Ile Ala Phe Gly Phe Glu Thr Leu Asp Asp Tyr Gln Asn Leu Gly 145 150 155 160 Phe Ile Ser Ile Ala Asp Gln Ser Arg Thr Asn Leu Thr Pro His Gln                 165 170 175 Gln Gln Ile Leu Asp Arg Tyr Gln Val Gln Ala Leu Thr Ile Pro             180 185 190 Leu Lys Leu Gly Gln Lys Leu Trp Gly Val Leu Ser Val Gln His Cys         195 200 205 Arg Asn Pro Tyr Ser Trp Ser Glu Arg Glu Ile Ala Ile Leu Arg Gln     210 215 220 Val Ala Ala Glu Leu Ile Ser Phe Gln Gln Ser Leu Asn Gly His Gly 225 230 235 240 Thr Ser Met Leu Asp Pro Asn Pro Gln Gln Val Glu Asp Val Leu Glu                 245 250 255 Arg Leu Arg Asn Ser Phe Asp Arg Val Gln Asp Arg Asp Arg Ser Val             260 265 270 Ala Arg Val Ile Glu Arg Ile Arg Gln Ser Leu Asn Ile Asp Ile Ile         275 280 285 Phe Arg Thr Thr Thr Lys Glu Val Arg Leu Leu Leu Lys Cys Asp Arg     290 295 300 Val Ala Val Tyr Gln Phe Asn Pro Asp Phe Ser Gly Gln Phe Val Ala 305 310 315 320 Glu Ser Val Thr Ser Gly Trp Val Lys Leu Val Gly Glu Glu Val Glu                 325 330 335 Arg Val Trp Lys Asp Thr Tyr Leu Glu Glu Asn Gln Gly Gly Arg Tyr             340 345 350 Ala Asn His Glu Thr Tyr Ala Val Ser Asp Ile Tyr Thr Ile Gly His         355 360 365 Asp Pro Cys His Val Glu Leu Leu Glu Gln Phe Glu Ala Arg Ala Tyr     370 375 380 Thr Leu Val Pro Ile Phe Gln Gly Glu His Leu Trp Gly Ile Leu Ala 385 390 395 400 Ala Tyr Gln Asn Asp Gly Pro Arg Glu Trp Gln Asn Ser Glu Val Asp                 405 410 415 Leu Leu Gln Arg Ile Ala Asp Gln Leu Gly Val Ala Leu Gln Gln Ala             420 425 430 Glu Thr Val Thr Gln Leu Arg Arg Gln Ser Asp Arg Ile Arg Gln Ala         435 440 445 Ala Glu Arg Asp Arg Thr Val Ala Gln Val Ile Asp Arg Ile Arg Gln     450 455 460 Ser Leu Asp Ile Asn Ser Ile Phe Asn Val Thr Thr Lys Glu Val Arg 465 470 475 480 Leu Leu Leu Gln Cys Asp Arg Ile Ala Val Tyr Gln Phe Asn Pro Asp                 485 490 495 Phe Ser Gly Gln Phe Val Ala Glu Ser Val Thr Pro Gly Trp Gly Lys             500 505 510 Leu Val Gly Pro Glu Val Glu Arg Val Trp Lys Asp Thr Tyr Leu Glu         515 520 525 Glu Asn Gln Gly Gly Arg Tyr Ala Tyr Arg Glu Thr Tyr Ala Val Thr     530 535 540 Asp Ile Tyr Glu Ala Gly His Asp Pro Cys His Val Glu Leu Leu Glu 545 550 555 560 Gln Phe Glu Ala Arg Ala Tyr Thr Leu Val Pro Ile Phe Gln Gly Glu                 565 570 575 Thr Leu Trp Gly Ile Leu Ala Ala Tyr Gln Asn Asp Gly Pro Arg Glu             580 585 590 Trp Gln Glu Asp Glu Val Gln Leu Leu Val Arg Ile Ala Asp Gln Leu         595 600 605 Gly Val Ala Leu Gln Gln Ala Glu Thr Val Gln Glu Leu Arg Arg Gln     610 615 620 Ser Glu Arg Ile Ala Arg Thr Ala Glu Arg Glu Arg Ala Val Ala Gln 625 630 635 640 Val Ile Glu Arg Ile Arg Glu Ser Leu Asn Leu Asp Thr Ile Phe Asp                 645 650 655 Thr Thr Thr Glu Val Arg Arg Leu Leu Lys Thr Asp Arg Val Cys             660 665 670 Val Tyr Gln Phe Ala Glu Asp Trp Ser Gly Ser Phe Ile Ala Glu Ser         675 680 685 Val Gly Ala Thr Trp Ala Pro Leu Val Gln Arg Gln Gln Gln Ile Pro     690 695 700 Ala Leu Arg Asp Asn Val Ser Gln Cys Asp Asn Met Thr Ser Leu Ile 705 710 715 720 Glu Arg Gln Gln Ser Gly Gln Leu Ala Pro Gln Ala Ser Gly Pro Ala                 725 730 735 Ser Arg Asp Thr Tyr Leu Gln Glu Thr Glu Gly Gly Arg Phe Gln Thr             740 745 750 Glu Gln Ala Phe Ser Val Glu Asp Ile Tyr Gln Ala Gly Phe Ser Thr         755 760 765 Cys Tyr Leu Glu Val Leu Glu Gln Tyr Gln Cys Arg Ala Tyr Ala Ile     770 775 780 Val Pro Ile Phe Lys Gly Pro Gln Leu Trp Gly Leu Leu Ala Ala Tyr 785 790 795 800 Gln Asn Asn Gly Pro Arg Asp Trp Glu Asn Glu Glu Val Thr Leu Leu                 805 810 815 Arg Gln Ile Gly Thr Gln Leu Gly Val Ala Ile Gln Gln Ser Glu Tyr             820 825 830 Leu Gln Gln Leu Gln Glu Gln Ser Leu Gln Leu Glu Ala Ala Ala Gln         835 840 845 Arg Asp Lys Ala Ala Lys Glu Gln Leu Gln Lys Arg Ala Leu Glu Leu     850 855 860 Leu Met Ala Val Arg Pro Ala Leu Asp Gly Asp Leu Thr Val Arg Ile 865 870 875 880 Pro Val Thr Glu Asp Glu Met Gly Thr Leu Ala Asp Ser Phe Asn Asn                 885 890 895 Thr Leu Gln Ser Leu Arg Lys Ile Val Met Gln Val Gln Thr Ala Ala             900 905 910 Gly Gln Val Val Gln Ser Ser Gln Ser Ser Asp Ser Ser Leu Met Glu         915 920 925 Leu Ser Asp Gln Ala Gln Lys Gln Phe Glu Glu Ile Ser Lys Thr Leu     930 935 940 Asp Arg Ile Gln Asp Val Val Gly Phe Thr Gln Thr Thr Ala Asn Asn 945 950 955 960 Ala Gln Gln Val Glu Val Ala Leu Gln Gly Ala Asn Arg Thr Leu Glu                 965 970 975 Ala Gly Asp Asp Ala Met Asn Arg Thr Val Glu Gly Ile Ala Asp Ile             980 985 990 Arg Gln Thr Val Ala Ala Ala Lys Gly Ile Ala Arg Leu Thr Gln         995 1000 1005 Ser Ser Gln Asn Ile Ala Lys Val Val Asn Leu Ile Asn Asn Phe Ala    1010 1015 1020 Thr Gln Thr Asn Leu Leu Ala Leu Asn Ala Ala Ile Glu Ala Thr Arg 1025 1030 1035 1040 Ala Gly Glu Tyr Gly Arg Gly Phe Ala Val Val Ala Asp Glu Val Arg                1045 1050 1055 Ser Leu Ala Arg Gln Ser Ala Asp Ala Thr Thr Glu Ile Glu Asn Leu            1060 1065 1070 Val Gln Glu Ile Gln Ala Glu Thr Lys Asp Val Thr Leu Ala Met Glu        1075 1080 1085 Met Ala Ser Glu Gln Val Thr Lys Gly Thr Asn Leu Val Gly Glu Thr    1090 1095 1100 Arg Gln Thr Leu Asn Glu Ile Val Thr Ala Thr Ala Gln Ile Gly Glu 1105 1110 1115 1120 Leu Val Gln Gly Ile Thr His Ala Thr Gln Val Gln Thr Glu Gln Ala                1125 1130 1135 Gln Ser Val Thr Glu Met Met Arg Ser Ile Ala Glu Ile Ala Asn Gln            1140 1145 1150 Thr Ser Glu Asn Ser Ile Ala Ile Ser Ser Arg Phe Lys Asp Ala Leu        1155 1160 1165 Thr Thr Ala Gln Glu Leu Gln Ser Ser Ala Gly Gln Phe Lys Val Ser    1170 1175 1180 <210> 11 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> GAF2 forward primer <400> 11 gtactcgaga cgggtgattg agcggattc 29 <210> 12 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> GAF2 reverse primer <400> 12 gtgaattcga tgacttgggc tacggtg 27 <210> 13 <211> 28 <212> DNA <213> Artificial Sequence <220> <223> GAF3 forward primer <400> 13 gtactcgagc cgcatccgtc aatccctc 28 <210> 14 <211> 26 <212> DNA <213> Artificial Sequence <220> <223> GAF3 reverse primer <400> 14 gtgaattcgt tcagagattc ccgaat 26 <210> 15 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> GAF4 forward primer <400> 15 catctcgagt cgggaatctc tgaacttag 29 <210> 16 <211> 28 <212> DNA <213> Artificial Sequence <220> <223> GAF4 reverse primer <400> 16 catctgcagc gggaatctct gaacttag 28 <210> 17 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> GAF2 / 3/4 forward primer <400> 17 gtactcgaga cgggtgattg agcggattc 29 <210> 18 <211> 28 <212> DNA <213> Artificial Sequence <220> <223> GAF2 / 3/4 reverse primer <400> 18 catctgcagc gggaatctct gaacttag 28

Claims (20)

A light harvesting dye GAF2 polynucleotide having the nucleotide sequence of SEQ ID NO: 1. The method according to claim 1,
Wherein the polynucleotide is derived from Microclause B353 which is a S. cereus. 3. The method according to claim 1 or 2,
Wherein the light harvesting dye absorbs an orange color at a wavelength range of 600 to 605 nm. A light harvesting dye GAF2 domain having the amino acid sequence of SEQ ID NO: 2. A light harvesting dye GAF3 polynucleotide having the nucleotide sequence of SEQ ID NO: 3. 6. The method of claim 5,
Wherein the polynucleotide is derived from Microclause B353 which is a microorganism of the genus S. cerevisiae. The method according to claim 5 or 6,
Wherein the light harvesting dye absorbs an orange color of 600 to 605 nm wavelength band. A light harvesting dye GAF3 domain having the amino acid sequence of SEQ ID NO: 4. A light harvesting dye GAF4 polynucleotide having the nucleotide sequence of SEQ ID NO: 5. 10. The method of claim 9,
Wherein the polynucleotide is derived from Microclause B353 which is a bacterium of the genus S. cerevisiae. 11. The method according to claim 9 or 10,
Wherein said light harvesting dye has a photoreactive activity of ultraviolet / green light. &Lt; RTI ID = 0.0 &gt; 8. &lt; / RTI &gt; 12. The method of claim 11,
Wherein the light reversible activity of the ultraviolet / green light is in a state capable of absorbing green light upon absorption of ultraviolet light and returned to a state capable of absorbing ultraviolet light again upon absorption of green light. 10. The method of claim 9,
Wherein the light harvesting dye is an auxiliary dye capable of transferring the absorbed energy to other dye molecules. &Lt; Desc / Clms Page number 20 &gt; A light harvesting dye GAF4 domain having the amino acid sequence of SEQ ID NO: 6. A GAF2 polynucleotide having the nucleotide sequence of SEQ ID NO: 1;
A GAF3 polynucleotide having the nucleotide sequence of SEQ ID NO: 3; And
9. An ultraviolet / green light harvesting pigment gene having the nucleotide sequence of SEQ ID NO: 9, which comprises a GAF4 polynucleotide having the nucleotide sequence of SEQ ID NO: 5. A GAF2 domain having the amino acid sequence of SEQ ID NO: 2;
A GAF3 domain having the amino acid sequence of SEQ ID NO: 4; And
An ultraviolet / green light harvesting pigment protein having an amino acid sequence of SEQ ID NO: 10 comprising a GAF4 domain having the amino acid sequence of SEQ ID NO: 6. 18. A recombinant vector comprising the ultraviolet / green light harvest pigment gene of claim 15. A method for introducing the recombinant vector of claim 17 into plants to increase efficiency of photosynthetic utilization. 19. The method of claim 18,
Wherein the plant is one selected from the group consisting of Cucurbitaceae having no light harvesting pigment as a light harvesting pigment, green algae having a chlorophyll-b as an auxiliary dye, and a terrestrial higher plant. 19. The method of claim 18,
Wherein the increase in the efficiency of using photosynthesis is achieved by using ultraviolet light and green light as a light source for photosynthesis through the introduction of ultraviolet / green light harvesting dye gene.

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