TW201127961A - Preparation of a compound comprising an amine group from an alpha-keto acid - Google Patents

Abstract

The invention relates to a method for preparing a compound comprising an amine group from an alpha-keto acid, wherein the preparation comprises using at least one reaction step catalysed by a biocatalyst. The invention further relates to a method, wherein an amine is prepared from an aldehyde or an alpha-amino acid. The invention further relates to a method for cyclising or polymerizing the amine.

Description

201127961 VI. Description of the invention: [^' Ming's labeling 3 The present invention relates to a method for preparing an amine group-containing compound (hereinafter referred to as 'amine>) from α-keto acid. The invention is more related to a process for the cyclization of the amines prepared from alpha-keto acids. c Previous winter 3 diamines and amino acids are currently most commonly prepared by chemical synthesis and are prepared from starting materials derived from mineral oil. Such compounds can be used as intermediates for the polymer and/or cyclic compound. In view of the current desire to use materials with higher persistence to prepare materials, it is desirable to provide a process in which an amine such as an amino acid or an amine is prepared using a biochemical process. In addition, it is also desirable to provide a method of lower energy demand than general chemical methods. A captanic amine, a cyclized amino acid, is an internal amine, which can be used to make polyamines such as Nylon-6 or Nylon _6,12 (a caprolactam and twelfth Further copolymers) ° Various methods for preparing caprolactam from a large chemical substance are: It is known in the art to include caprolactam from cyclohexanone, methyl phenol, phenol, cyclohexanol or the like. These intermediate compounds are generally available from minerals. In the current state of the art (4) for the preparation of materials with higher persistence, it is desirable to provide an intermediate compound for the preparation of caprolactam: Taifa, which can be obtained from biological properties. Renewable sources, or intermediate compounds can be converted to caprolactam by biochemical methods. In addition, = can provide a method of less energy demand, and the general use of chemicals from stone source Method phase ^ 匕来201127961 « ^ . A ΟΛ. ^ j acid (6-80) preparation of caprolactam, known from the amino acid 6-aminol l L. A / L 〆-+ [ As described in WO 2005/068643, as described in US-A 6,194,572, 6-ACA can be a 6-amino group by the presence of an enzyme having 〇,β-sterol reductase activity. 2-enoic acid (6-fluorene) is prepared by biochemical conversion. The 6-fluorene can be prepared from an amide acid, such as a biochemical method or a purification synthesis method, although 6-ACA is prepared by reducing 6-oxime at W0. It is feasible in 2005/068643, but the inventors found that 'in the reduction reaction strip The lower-6-AHEA undergoes an irreversible cyclization reaction spontaneously and in large quantities to form undesirable by-products, especially β-homoproline. This cyclization reaction is the manufacture of 6-ACA. Technical bottlenecks can cause considerable yield loss. SUMMARY OF THE INVENTION One object of the present invention is to provide a novel process for the preparation of amines such as an amino acid such as 6-ACA' or a cyclized amine such as Amine amines - especially, which can be used in the preparation of polyamines, or intermediate compounds for the preparation of amines or cyclized amines, can be an alternative to known methods. Another object of the invention is to provide a novel Method, which overcomes one or more of the above disadvantages. One or more other objects that may be solved in accordance with the present invention will be described in the following detailed description. I: Embodiments It is currently known to prepare amines from alpha-keto acids, especially Is an amino acid, that is, it has been found that 6-aminocaproic acid (6-ACA) can be prepared by a method comprising at least one biochemical step, wherein the 6-aminohexanoic acid is prepared from 2-sided oxygen, heptane Acid is also known as 201127961 as cx-keto pimelic acid (akp). The special method of preparing 6 ACAi has been considered by the Ming people to open up a more general method of converting 1 keto acid to amines. In particular, the preparation of the amine can be carried out in two or more steps. A method is provided in which the enthalpy is first converted to _, and then the genus is converted to an amine. The other method is... wherein the keto acid is first converted to an amine hydrazine, and then converted to an amine. For example, In the preparation of 6_8 (: 八, Ακρ can be converted to 5-methylmercapto _ (5_f valeric acid, 5 FVA), then 5-FVA is converted to 6_ACAe. In addition, AKp is also converted to a first. Aminopimelic acid (AAP) is then converted to 6-ACA.

J, the inventors have found that, in principle, amines can be prepared from a-keto acid by chemical means (i.e., no biocatalyst is used). Every-reverse step should be appropriate. The 4* method is provided as follows. However, the inventors have also appreciated that amines can be prepared from a-keto acids using biochemical methods. Accordingly, the present invention is particularly related to a method for preparing an amine-containing compound of the following formula: 2 Ν CH2''--ar is prepared from the following formula: ct-decanoic acid (1) HO - 〇II c-C-a-R 0 (2); A represents a thiol group. The hydrocarbon group may contain one or more substituents, and/or contain one or more impurities. 'In particular, a hydrocarbon group having 2 to 10 carbon atoms; 201127961 R represents a monofunctional group, especially selected from the group consisting of CN, COOH and NH2; and wherein the preparation of the ruthenium comprises the use of at least one biocatalyst catalyzed Reaction step. And may be particularly selected from the group of functional groups which can be converted to another functional group selected from the group consisting of CN, COOH and NH2 by known methods (e.g., using a biocatalyst). The present invention is more related to a method in which a compound containing an amine of the formula (1) is prepared by using a biocatalyst, and a hydrazine--~~-Α--r 〇(3) is prepared from the aldehyde of the following formula; wherein Α and R are as Defined by equations (1) and (2). As described above, the aldehyde of the formula (3) can be obtained from the α-keto acid of the formula (2). The present invention is more related to a process in which the amine of the formula (1) is prepared from an α-amino acid of the following formula using a biocatalyst.

II

H0—C~CH—A—R NH2 (4); wherein A and R are as defined by formula and (2). As described above, the oc-amino acid of the formula (4) can be obtained from the hydrazine-acid of the formula (2). In one embodiment, the amines of formula (1) prepared by the process of the invention are useful in the preparation of cyclic products. For example, the cyclic product may be an internal amine if the amine is an amino acid. This method involves the cyclization of the amine, such as an amino acid, selectively in the presence of a biocatalyst. For example, 6-ACA prepared by the method of the present invention 201127961 can be used to prepare caprolactam. The term amine as used herein refers to a compound containing an amine group. In particular, the term "amine" refers to a compound containing an amine group of formula (1). If the ruler is a COOH group, then the amine of the formula (1) can be called an "amino acid." If the base is used, the amine of the formula (1) can be called a "diamine". If the group A of the formula (1) also contains one or more amine groups, the "diamine" may contain more than one amine group, and may also be referred to as a diamine (when A contains an amine group) , tetraamine (when a contains a diamine group) and the like. The term "OC-amino acid, as used herein, refers to an amino acid in which an amine group is bonded to a homo-acid group to a homo-carbon atom. In particular, the term refers to a compound of formula (4). Formulated according to the process of the invention (1) The amine may be 6-aminocaproic acid (6-ACA). In this case, the (C)-formed CX-keto acid of the amine is α, pimelic acid. The formula prepared according to the method of the present invention may be other compounds. In this case, the A group in the formulae (1) to (4) represents ((: 1^, where \ is selected from one of the groups consisting of 2, 3 5 6 , 7, 8 and 9). One soil, A% One makeup hexane, diamine, ω-amine H, Thread (5 birds also known as 5-amino fiber light), 〇> amine group, heptanoic acid amine amide material = its The amine may be selected from the group consisting of "······················································· The group, the a-keto acid for the amine 1 is preferably α-glutaric acid (AKG), α-dicarboxylic acid (aka) or (d) suberic acid (AKS). For acid esters, such as α-keto acid, amino acid, 6-ACA, 2-aminopimelic acid (α-aminoglycol Acid, hereinafter referred to as 'ΑΑΡ,', 5-FVA or ΑΚΡ, these terms include protonated carboxylic acid groups (ie, neutral groups), their corresponding carboxylic acid esters (conjugated bases thereof), and Salts. When referring to amino acids, such as oc-amino acids or 6-ACA, the term includes zwitterionic forms of amino acids (wherein the amine group is protonated and the carboxyl group is in a deprotonated form) An amino acid in which an amine group is protonated and a carboxyl group is in a neutral form, and an amino acid in which the amine group is in a neutral form and the thiol group is in a deprotonated form, and a salt thereof. According to the present invention, When 6-ACA is formed, and when caprolactam is selectively formed, there is no undesired intermediate product cyclization problem which leads to loss of yield. It is conceivable that the process of the present invention can obtain comparable or better yields, In contrast to the method described in WO 2005 Secret 43. It is envisioned that the method of the present invention preferably employs living organisms, particularly in the growth and maintenance of organisms. In accordance with the method of the present invention, unless otherwise indicated. Unless otherwise stated. Additives, etc.) Including, it is also envisioned that the yield (g/lh) of the amine of formula (1) can be improved in the examples of the present invention. The term "or" is used herein to define "and/or, the term" "Or" a "here" means "at least - when referring to a single noun (such as a compound, - plural. When referring to a compound that has a steric isomer, the compound may be a part of this space isomer) Or a combination thereof. Therefore, when it is mentioned that an acid such as an amino group 8 201127961 has a mirror image isomer, the amino acid may be a mirror image isomer, a mirror image isomer, or a combination thereof. If there is a natural spatial difference The construct, which is preferably a natural steric isomer. When the enzyme is labeled with an enzyme group (EC) in parentheses, the enzyme is classified or classified into a certain group based on the enzyme nomenclature of the Biochemistry and Molecular Biology International Organization Nomenclature Committee (NC_IUBMB). httP://WWW_chem.qmul.ac.uk/iubmb/enzyme/. Other suitable enzymes that have not yet been classified as a specific group but can be classified are also included. The term "homologue" as used herein, in particular, refers to a polynucleotide or a multi-peptide, having a sequence identity of at least 3%, preferably at least 4%, more preferably at least a sail of at least (four), more Preferably at least township, preferably at least, preferably at least 8〇%, more preferably at least 85%, more preferably at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96 %, at least 97%, at least 98%, or at least 99%. Furthermore, homologs generally have significant sequence similarity, typically greater than 3%, especially at least 35%, preferably at least 4%, more preferably at least 60%, and even more preferably at least 65%. More preferably at least %, more preferably at least 80%, more preferably at least 85%, more preferably at least 9%, at least 9%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, At least 97%, at least 98% or at least 99%. 5 The original I has a common specific function with its homologous polynucleotide or polypeptide, such as those encoding the same peptide, which can catalyze the phase reaction (generally converting the same & mass to the same compound) or similar reaction. "Similar reactions" are generally a reaction of the same form, such as decarboxylation, amino transfer, C1-201127961 chain extension reaction, and the like. Therefore, according to the rule of thumb, the homologous enzyme can be classified into the same ec group, and the first three codes (x.y.z) of the shared ec group, such as the carboxylase, are all EC 4.1·1. In general, in similar reactions, 'the same group of acceptors (such as amines, carboxylic acids, amino acids), similar receptors of similar reactions 'generally convertible to homologous compounds' due to products of similar reactions Mostly analogs. Similar reactions include, inter alia, those defined by the same chemical conversion reaction, as defined by the same KEGG RDM path, where the R- and D-atoms describe the chemical transformation (KEGG RDM pathway: Oh, M. et al. (2007) Systematic Analysis of enzyme-catalyzed reaction patterns and prediction of microbial biodegradation pathways. J.

Chem. Inf. Model" 47, 1702-1712). The term homologue includes a nucleic acid sequence (polynucleotide sequence) which differs from another nucleic acid sequence by a degeneration of the genetic code or experimental adaptation. And encoding the same multi-peptide sequence. The term "functional analog" as used herein means that the nucleic acid sequence differs from the specified sequence, wherein the analog encodes a peptide (protein, enzyme) having the same amino acid sequence. Or a homolog of the peptide. In particular, a preferred analog is that the nucleic acid sequence has similar or better expression in a host cell of interest, since the nucleotide sequence is a host cell It is preferred to use 'as a Wei analog. In this view, as seen in the technical field of 'observable _ good performance, if you want to express peptides (proteins, enzymes), usually refers to higher performance However, in the case of a specific implementation, the amount of money may be lower than the amount of the metabolic path of the host cell due to the pre-suppression. Functional analogue 10 201127961 can be natural hair The sequence, ie wild type functional analogue, or gene sequence was to adorn, namely non-wild-type functional analogues, which are designed to be able to achieve the desired amount of performance.

Sequence equivalence or similarity is defined herein as the relationship between two or more multiple sequences, or between two or more nucleotide sequences, to compare the sequences. Generally, sequence equivalence or similarity can compare the full length of the sequence, but can also be compared by simply aligning one of the sequences. Technically, "equivalent" or, "similarity" also refers to the degree of correlation between polypeptide sequences or between nucleic acid sequences, in which case the match between the sequences is preferred. The method of similarity is to provide a maximum match between the sequences to be tested. For the purposes of the present invention, a computer program for determining the degree of equivalence or similarity between two sequences preferably includes BLASTP and BLASTN (Altschul, SF et Al·, J. Mol. Biol· 1990, 275, 403-410, publicly available from NCBI and other sources (BLAST Manual, Altschul, S·, et al., NCBI NLM NIH

Bethesda, MD 20894). The preferred parameters for comparing the multi-peptide sequences used in BLASTP are 1 〇·〇, space extension 0.5, and Blosum62 matrix. The preferred parameters for comparing polynucleotide sequences using BLASTN are open space 10.0, space extension 0.5, DNA complete matrix (DNA equivalence matrix). In accordance with the present invention, at least one step in the method utilizes a biocatalyst which is catalyzed by a biological material or fragment derived from a biological source, such as an organism or a biomolecule derived therefrom. The biocatalyst specifically comprises one or more enzymes. The biocatalyst used may be in any form. In one embodiment, one or more of the enzymes used are isolated from the natural environment (isolated from the organism from which they are made), such as a solution 11 201127961 solution, emulsion, dispersion, freeze-dried cells (suspension) ), its lysate, or fixed to the support. In one embodiment, one or more enzymes form part of a living organism (e.g., intact living cells). Enzymes perform catalytic functions in cells. The enzyme may also be secreted into the culture medium in which the cells are present. The living cells can be cells in growth, resting or dormant cells (such as spores), and cells in a stable state. It is also possible to use a permeabilized cell (that is, the enzyme that is responsible for the enzyme, or the precursor of the enzyme, or the enzyme is permeabilized). The biocatalyst used in the method of the invention can in principle be any - organism' or derived or derived from any organism. The organism can be a eukaryote or a prokaryote. In particular, the biological system is selected from the group consisting of animals (including humans), plants, bacteria, archaea, yeasts and fungi. In one embodiment, the biocatalyst is derived from an animal, especially a portion thereof - such as the liver, pancreas, brain, kidney, heart or other organs. The animal may especially be selected from mammals, and is particularly preferably selected from the group consisting of rabbits, murines, pigs and bovidae. Particularly suitable plants include those selected from the group consisting of CWwrWiaceizej, especially CwcMrfeiia, such as pumpkin (CwrcMr...α mc^c/ιαία), or cucumber (Cwcwm(4); cross Flower family (firim/caceae) 'especially Arabidopsis, such as Arabidopsis thaliana (九ί/ζα/ίαηα); 山散(A/acim'aZi··?), such as perennial mountain models; big wind and longhorn beans (Ceraiom'a) The group consisting of 12 201127961 Suitable bacteria may be selected in particular from the bacterium (νΑπ.ο), Pseudomonas (Ρμμ如如(10)), Bacillus (7) (1), rod-like Bacillus (C〇〇; nMac^WMm), Brevibacterium (J?reW^cieriMm), Enterococcus, Enterococcus, Actinomycetes fAci/nomyceia/ej'J, Klebsiella (Alzheimer's disease) XfitCiOCOCCM·?), Lactobacillus, Clostridium, Escherichia, Anabaena, Microcystis, Synechococcus, Rhizobium (Τί/^i^MmJ, Slow-growing rhizobium (7)ra^r/nz£^ww), thermophilic bacteria (TViermii·?), nitrogen-fixing bacteria (A(3)i〇&aCfer), ballooning (AenjcoccMsj, thermophilic) Bacteria (TTierm (10)), Mycobacterium, Zymomima W, Proteus, Agrakceni/m, GeokciZ/Mi, AciVieiohacier, Rolstonella (/^ /·5ίο«/β), red bacillus (/?/ίί?ύ?οadmin acier), paraffin (Tarflcoccwi), sphingomonas (TVovc^p/nTigc^iMm), nitrite ( Miro (10) monads), Legionella (Legicmd/a), Diplococcus, Rhodopseudomonas, Staphylococcus (•Siap/^/ococcwi), Deinococcus (X^iTiococcMi1), and Salmonella (Salmonella). The archaea may be particularly selected from the group consisting of Archaea (Arc/meog/oi>w5·), aerobacteria, halophile (Tffl/obacieWMm), and Mei/iimoiflrci'na.曱 球 (Afei/iimococcMjj, Thermoplasma (T/iermop/aimaJ, Fire bacillus (T; yri) kcM/Mmj, 曱 暖 暖 (i^ei/iimocaWociJCCM·?), 甲 菌

I 13 201127961 {Methanobacterium), Methanosphaera, genus 炙, hot-milled YMei/icmopyn^) 7 and acier. Suitable fungi may be particularly selected from the group consisting of leather bacteria (Emericella), Ustilago, Cephalosporium, pseudo-green emblem (cadecary aeci/omycej), Trichophyton (7Wc/) Ic7?/i;yiMm), roots of the genus (can be like a corpse), the key emblem (iVewr &lt;9577£?riZ), penicillium and sputum CAiperg out of M5 ·) group. Suitable yeasts may in particular be selected from the group consisting of Schizosaccharomyces (Sc/iizoiacc/ia/Omycei), Sterol yeast (Tic/H'a), Candid, Hansinensis (Tfimsenw/). α), Kluyveromyces, Yarrowia (Fam&gt;vWa) and yeast (Sacc/iaromycej). It will be apparent to those skilled in the art that naturally occurring biocatalysts (wild type) or naturally occurring biocatalyst mutants having the appropriate activities required for the methods of the invention can be used. The characteristics of naturally occurring biocatalysts can be enhanced by biotechnology known to those skilled in the art, such as molecular evolution techniques or rational design techniques. For example, a mutant of a wild-type biocatalyst can be achieved by modifying the coding DNA of the organism which can be used as a biocatalyst or a biocatalyst fragment (such as an enzyme), using mutation techniques known to those skilled in the art (random mutation technique) , localization mutation technology, directed evolution, genetic recombination, etc.). In particular, the DNA may be modified to encode an enzyme different from the wild-type enzyme having at least one amino acid, such that it encodes an enzyme comprising one or more amino acid substitutions, deletions and/or insertions, and The wild type 14 201127961 is compared to 'or the mutation is combined with the sequence of two or more original enzymes, or • thereby affecting the performance of the modified DNA in appropriate (host) cells. The latter can be achieved by methods known to those skilled in the art, such as codon optimization or codon pair optimization, as based on the method described in WO 2008/000632. The mutated biocatalyst may have enhanced properties such as one or more of the following properties: selectivity to the substrate, activity, stability, solvent tolerance, pH profile, temperature profile, quality curve, inhibition sensitivity, coenzyme utilization Sex, as well as affinities. Mutants with enhanced properties can be identified by appropriate methods such as high performance screening or methods known to those skilled in the art. &quot; When referring to biocatalysts of specific origin, especially enzymes, recombinant biocatalysts derived from the first organism, but actually made by the second organism (genetically modified), especially enzymes, especially those from A biocatalyst of the first organism, especially an enzyme. In a preferred embodiment of the invention, the method of preparation comprises a biocatalytic (usually enzymatic) reaction which catalyzes the alpha-keto acid or amine oxime (the imprint contains at least one carboxylic acid group and at least one amine) The compound of the base) is in the presence of a biocatalyst for the decarboxylation reaction. Therefore, an enzyme having such catalytic activity may be referred to as an α-keto acid decarboxylase or an amino acid decarboxylase. The hexanoic acid is preferably a diacid, wherein the biocatalyst selectively acts on a ketone- or amine-based acid group. If R in the formula (2) ct-keto acid is COOH, the appropriate decarboxylase has ketodicarboxylic acid decarboxylase activity, which can catalyze the conversion of the keto acid to the aldehyde of formula (3), or - acid retardation Ester dehydrogenase activity, can catalyze the conversion of oc-keto acid to formula (3), 15 201127961 or amine-salt acid can deactivate enzyme activity, can catalyze the conversion of &amp; keto acid to amine of formula (1) class. An enzyme capable of decarboxylating oxonic acid or amino acid, preferably selected from the group consisting of de-enzyme (EC 4.U), more preferably selected from glutamic acid decarboxylase (Ec 4.1·1.15) ), grass 醯 acetic acid decarboxylase (EC 4.1.1.3), diamine pimelic acid de-suppressing (EC4.U.2〇), aspartic acid 1-decarboxyl (EC 4.1.1.11), branch 阙Acid destabilizing enzyme (EC4.1.l_72), ot-ketoisovalerate decarboxylase (EC 1.2.4.4), α_glutaric acid decarboxylase (EC 4.1.1.71), and pyruvate decarboxylase (ec 4.1.1.1) The group consisting of. One or more suitable decarboxylases may be selected from the group consisting of oxalate decarboxylase (Ec 4.1·1·2), acetamidine decarboxylase (EC 4.1.1.4), proline decarboxylase/leucine Retarase (EC 4.1.1.14), 3-hydroxyglutamate decarboxylase (EC 4丄1.10), ornithine decarboxylase (EC 41117), lysine decarboxylase (EC 4·1·1· 18), arginine decarboxylase (EC 4.1.1.19), 2-oxoglutaric acid decarboxylase (EC 4.1.1.71), and diaminobutyrate decarboxylase (ec4.L1.86) The group of people. The decarboxylase is particularly a decarboxylase of an organism selected from the group consisting of pumpkin, cucumber, yeast, fungus, such as cucurbit from e), yeast (such as Saccharomyces cerevisiae ceav/Wfle),硪 硪 如 C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C Phytophthora (such as 爪 圭 圭 Λ Λ ι ι ι ι ι ι ι 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 2011 Brain; and bacteria such as Escherichia (such as the large intestine rod halogen (five k, lactobacillus (Xaciococciuj (such as Lactobacillus leucocephala, dynasty _ fMycc ^ acien 'Mm) (such as Mycobacterium tuberculosis (both; m &amp; r (3) Zcw · Chuan, Clostridium (c / still in the mountains), Lactobacillus {Lactobacillus, _ (SirepiococcM ·?), Pseudomonas (Pjewiiomcmiij) (such as the genus Pseudomonas (仏 (10) Claw (10) like)), as well as the group consisting of sports singular bacteria mo along _s). a hexpyruvate de- _ can be obtained from Saccharomyces Cerevisiae cereviW Aej or exercise early morning pain (Zymomowiw. In particular, can be used from Z. mobilis (Z; ymom 〇 则 办! (8) of the C-acid detoxase mutant I472A. Can be used from E. coli ( £:. c〇/〇 glutamic acid decarboxylase, diaminopimelate detoxase or aspartyl detoxification enzyme. Can be used from the rough round spore emblem (iVewrc^pora Bacillus (A^ycMacieWMm /epr-like), Clostridium perfringens per/Wnge/w), Lactobacillus brevis ([Shen Ruo/?reW·?), Mycobacterium tuberculosis Μ厶Μ厶cc/im.i ), Streptococcus CSirepMcocci^), or Lactobacillus (L&lt;3cmCOCCMiS) glutamic acid decarboxylation S#. Examples of lactic acid bacteria species derived from sulphate deactivating enzymes' specifically include Lactococcus lactis/&lt;3&lt;: along), such as Lactococcus lactis (LflciococcM·? /ac) B1157, L. lactis (I/flciococcM·? Zaci/i) IFPL730, especially LaciococcwWacii·? var· e 17 201127961 maltigenes (JL is called Streptococcus lactis var. maltigenes.) In particular, oxalic acid acetate depleted enzyme derived from Pseudomonas can be used. A branched α-keto acid decarboxylase derived from Lactococcus lactis (Z^ciOCOCCM·?^3Ciz\s) can be used. More specifically, a keto-p-isovalerate decarboxylase derived from Lactococcus lactis (/^(^)(9)" can be used. Particularly useful from Mycobacterium tuberculosis. Both; cc^ac( eri.Mm iw ercM/oWW α-ketoglutarate decarboxylase. In a preferred method of the invention, the preparation of the amine of formula (1) comprises carrying out an enzyme reaction in the presence of a biocatalyst, the biocatalyst The catalytic activity of the catalytic amine transfer reaction and/or the reductive amination reaction, preferably the enzyme has at least one selected from the group consisting of an aminotransferase (EC 2.0) and an amino acid dehydrogenase (EC 1.4.1). The catalytic activity of the constituent group. Further, the enzyme reaction can be carried out in the presence of an amine-based donor. If the amine is 6-ACA, the appropriate aminotransferase has a 6-amino group of 6-aminohexanoic acid. The activity of the transferase can be converted to 6_ACA by t5_FVA, or the 2-aminotransferase activity of α-aminopimelate can catalyze the conversion of akp to AAP. The aminotransferase can be selected, in particular, from β-amino group. Butyric acid: α-ketoglutarate aminotransferase, β-alanine aminotransferase, aspartate aminotransferase, 4-amino-butyrate aminotransferase Enzyme (EC 2.6丄19), L-Amino Acid 6-Aminotransferase (EC 2.6.1.36), 2-Aminoadipate Aminotransferase (EC 2.6.1.39), 5-Aminovaleric Acid Aminotransferase (EC 2 6 1 48), 2-aminocaproic acid aminotransferase (EC 2.6.1.67), lysine: pyruvate 6-aminotransferase (EC 2.6.1.71) ', and aromatic a group of amino acid aminotransferases (E(: 2 6 1 57) 18 201127961. In the embodiment, the aminotransferase is selected from the group consisting of alanine aminotransferase (EC 2_6丄2) ), leucine aminotransferase (EC 2.6.1.6), alanine-side oxy-acid aminotransferase (EC 2.6.1.12), β-alanine-pyruvate aminotransferase (EC 2.6丄18) ), 〇5)-3-amino-2-mercaptopropionic acid aminotransferase (£0 2.6.1.22), L,L-diaminopimelate aminotransferase (EC 2.6.1.83) An enzyme having a catalytic activity for catalyzing an amine transfer reaction and/or a reductive amination reaction, such as an aminotransferase or an amino acid dehydrogenase, may be selected from the following group of organisms selected from the group consisting of Vibrio (cardiac); Pseudomonas (cadaverium ΟΑ/ΟΑηΟ/ϊβ*?); Bacillus licheniformis (5&lt;3C Chuan Mi) ; Shan Yi; Iron Fern (Aw/em'wm); Carob (〇Γ3<3, 〇Γ&lt;3ίσηζ'α);Mammal;Streptomyces;Escherichia; Thermophilic bacterium (TTienwws); iS^cc/mromyces); Brevibacterium (fiz-eWbizcien'wwi); Corynebacterium; Proteus (corpse; Agrobacterium (AgrMacieriwm); Bacillus licheniformis; Acinetobacter; Ralstonella such as ma); Salmonella OSVi/mime/Za), Rhodococcus, and Staphylococcus aureus Sia/?/i;y/oC(?(:cwi) group, especially selected from Bacillus subtilis), Bacillus licheniformis 5acz7/«j weihenstephanensis, sphaeroides, staphylococcus aureus, Legionella pneumophila (Legi(10)e//fl, European nitrite, Nitrosomonas ewropaea), gonorrhea 19 201127961 Gcmorr/ioeae), Pseudomonas syringae (the corpse), swamp red fake cell halogen (/?/z〇i/c&gt;pieMiic&gt;mim&lt;2i pa/wsirb), Vibrio fluvialis (11⁄4• The group consisting of η·σ /ZiiWa/b) 'and Pseudomonas aeruginosa (10) j· (3) (10). The aminotransferase may be particularly selected from mammals; Mercwn 'ah··?', especially perennial mountain races (_/ν/^π: «η·&lt;3Ζί·5· perennb), More preferably a perennial mountain drum (the root of Mercwria/^ perenn (4); Asplenium ', especially the singular squid (Asplenium unilateral) or the stellate fern (Asp/em'Mm sepienin'cmiz/e) , Cemicm/a, especially Ceraioma d such as wa / Bacillus (/e/ioiiokcie/·), especially Rhodobacter sphaeroides (/?/z〇i/c^acier sphaeroides), Staphylococcus , especially Staphylococcus aureus &lt;2ΜΓβΜ·ί); Vibrio (ViftWo), especially Vibrio fluvialis (V7 η·ο //wWiz//·?); Pseudomonas, especially patina PjeM^mcmas izen^Moja; Rhodopseudomonas (/?/ι〇pwMi/omimiW), Bacillus licheniformis (β〇^7ΖΜ·5), especially Bacillus licheniformis wd/ienWep/ Ianewj (4), and Bacillus subtilis (5ad / / (10); Legionella (LegioneZZa); nitrite (Miroiomcmcti); dicocci / 4 yeast, especially Saccharomyces cerevisiae (Sacc / iaromyca j amine group Base transferase In the case of a mammalian enzyme, particularly a brain derived from a mammalian kidney, a mammalian liver, a mammalian heart or a mammal. For example, a suitable enzyme may be selected from a β-amino group derived from a mammalian kidney. Isobutyric acid: α-ketoglutarate aminotransferase, especially β-amine from porcine kidneys 201127961 Isobutyric acid: oc-ketoglutarate aminotransferase, β-alanine from mammalian liver Aminotransferases, especially beta-alanine aminotransferases from rabbit liver; aspartate aminotransferases from mammalian hearts, especially aspartate aminotransferase from porcine heart; 4-amino-butyrate aminotransferase in mammalian liver, especially 4-amino-butyrate aminotransferase from pig liver; 4-amino-butyrate aminotransferase from mammalian brain In particular, 4-aminobutyric acid aminotransferase from human, porcine or rabbit brain. In one embodiment, the aminotransferase is selected from the group consisting of α-ketoadipate from Aspergillus sp. Glutamine aminotransferase, especially from Neurospora crassa (//euroyin c&g) t;ra craua) adipic acid: amygdamine aminotransferase; 4-amino-butyric acid aminotransferase from E. co., or from thermophilic bacteria (77 ι^τηΜ) · ί) of cc-aminoadipate aminotransferase, especially from extreme thermophiles (a-aminoadipate aminotransferase of TTiermMs) and from Clostridium (cl〇stridiurn), In particular, it is derived from the group consisting of C-(10)π-extracted 5-aminopentanoate aminotransferase. Appropriate 2 Aminoadipate Aminotransfer i# can be free from Icelandic thermobacter. In particular, the amine group can be selected from the group consisting of ammonia, ammonium ions, amines and amino acids. Suitable amines are primary and secondary amines. The amino acid can have a D- or L-configuration. Examples of amine-based suppliers are alanine, glutamic acid, isopropylamine '2-aminobutyrol, 2-aminoheptanthene, phenylguanamine, 1-phenyl-1-aminoethane, noodles Indoleamine, tyrosine, phenylalanine, aspartate, beta-aminoisobutyrate, beta-alanine, 4-aminobutyrate, and a-aminoadipate.

S 21 201127961 In a preferred embodiment, the amine of formula (1) is subjected to a biocatalytic reaction, in the presence of a catalyzed reductive amination reaction, and in the presence of an ammonia source, Provider's ch-nh2 base &amp;

θ emulsified reductase (EC 1.4), especially 疋 selected from the group consisting of amino acid dehydrogenase (Ec. i 4 . υ. If 6-ACA is prepared, the appropriate amino acid dehydrogenase has 6-amino group (4) A few enzyme activities, can catalyze the conversion of 5-FVA to 6-ACA, or have α~amine, which can convert ΑΚΡ to ΑΑΡ. Particularly suitable amino acid deaminase Is selected from the group consisting of diaminopimelate dehydrogenase (EC L4.U6), lysine 6-dehydrogenase (EC 1.4.1.18), glutamate dehydrogenase (EC 1.4.1.3; Ec 1 4 1 Μ, and a group consisting of leucine dehydrogenase (EC L4 丄 9). The amino acid dehydrogenase is preferably a lysine 6-dehydrogenase (EC 1.4.1.18). The amino acid dehydrogenase may be selected from the group consisting of glutamate dehydrogenase (EC 14丄3) classified as NAD or NADP as a recipient, and glutamic acid as a recipient when NADP is used as a recipient. Hydrogenase (Ec 14 14), leucine dehydrogenase (EC 1.4.1.9), diaminopimelate dehydrogenase (Ec L4.1.16), and lysine 6-dehydrogenase (EC 1.4. 1.18) Amino acid dehydrogenase. Amino acid dehydrogenase may be derived in particular from Corynebacterium Corynebacterium glutamicum (Proteus) ^ Proteus vulgaris (PraMttj / Agrobacterium, bacteria, especially Bacillus thermophilus (GeokciZZM5 • sieiiroi/iermop/u/Mi·); Acinetobacter, especially Acinetobacter ADPl (Ac/neiokcie sp. ADP1); Ralstonella 22 201127961, especially R. solani yo^mac print / Mm) / Salmonella, especially Salmonella Rod 囷 (iS&quot;iz/mcme//a; yeast (Sacc/ziz/Omyce·?), especially Saccharomyces cerevisiae_cerev/s/ae); Brevibacterium (5/-eWki:ieriMm)' especially yellow Brevibacterium (Brev/office; and Bacillus (5 sinking &quot;/milk), especially B. sphaericus (βαα·&quot;Μ·5 印/zaencMd, Bacillus cereus ceaMiy), or Bacillus subtilis (5〇) ^7/如如如7(4) An organism that constitutes a group. For example, a suitable amino acid dehydrogenase may be selected from the group consisting of diaminopimelate dehydrogenase from Bacillus (5ac///MJ), especially from Bacillus sphaericus (7) sinking edge (10), from the genus Brevibacterium (_greW6acieri'wm sp.) diamine heptanoic acid deamination bee, from Dehydrogenation of diaminoheptanoic acid from Corynekcier/wm, especially from Corynebacterium glutamicum (diamine-pimelate dehydrogenase of C^A-ywAacieriMm; from Proteus (Pr〇eMiS) a diaminopimelate dehydrogenase, especially a diaminopimelate dehydrogenase from the common Proteus (/&gt;r(?ieMs vw/gaWj); from Agrokcie Wi/m Amino acid 6-dehydrogenase, especially from Agrobacterium tumefaciens, lysine 6-dehydrogenase from Bacillus licheniformis, especially from Geobacillus stearothermophilus A glutamate dehydrogenase (EC 1.4.1.3) from Acinetobacter (Ac/neiMacie), acting as a coenzyme with NADH or NADPH, especially glutamic acid from ADP1 (AnVieioMcie sp. ADP1) Hydrogenase; from Rolstonella (7? must be called glutamate dehydrogenase (EC 1.4丄3), especially from the sapling S: 23 201127961

Amylin dehydrogenase from Salmonella typhimurium 5O/tmacec!rMm; a face acid dehydrogenase from Salmonella (EC 1.4.1.4) with NADPH as a coenzyme, especially from Salmonella Salmonella typhimurium (Sa/mcmeZ/a's amylin dehydrogenase; from leaven

The crude amino acid dehydrogenase of the parent strain (Sacc/iaromyca) (EC 1.4.1.4), especially the glutamate dehydrogenase from Saccharomyces cerevisiae (iSacc/iaromyce·cerev/ike); from Brevibacterium (Brevi 'kcieri'wm) sulphate dehydrogenase (EC 1.4.1.4), especially glutamate dehydrogenase from Brevibacterium flavum (J5rev digested with acieWwm //avwm); and from Bacillus (BadZ/Mj) Aleucine dehydrogenase, especially from Bacillus licheniformis CBacH/Ms cerew·?) or Bacillus subtilis (10) leucine dehydrogenase along (4). In a particular embodiment, the alpha-keto acid of formula (2) is biologically converted to an aldehyde of formula (3) in the presence of a decarboxylase or other biocatalyst that catalyzes such a reaction. For example, hydrazine can be biocatalyzed to 5-FVA using this method. The decarboxylase used in the present invention may be particularly selected from the group consisting of Lactococcus lactis var. maltigenes / this subsp. (X-keto acid de-enzyme; obtained from Reiter Lactococcus lactis (Z/flCiococcwi1丨B1157 or Lacioc^ccw Zflcik) Branched α-keto acid degrading enzyme of IFPL730; obtained from Saccharomyces cerevisiae, Candida albicans (Cimc^£/a// Are..., pyruvate dehydrogenase of Z. mobilis (ZymomcmiW, H. sphaeroides ίρ.), Xenopus laevis yavamcwj), R. sphaeroides or Marshdamii;自结24 201127961 α-ketoglutaric acid of the bacterium of the genus Mycobacterium bismuth, from the large intestine rod i| (£· co / i), Lactobacillus brevis (Z ^ cic ^ aciZ / Mj reWs), leprosy Bacillus/India (rae), crude sugar-bonded emblem cmwa), or Clostridium perfringens ((: 沁价心·说/^r/n&gt;^elike) glutamate decarboxylase; From Escherichia coli (five C?? (1) aspartate decarboxylase. Especially 'from E. coli cW), Zymomonas mobilis, Saccharomyces cerevisiae c ereWszae), tuberculosis dysenteriae Ci?kCi state (4) iM~rctj〇5(4), Phytosanitary (PMWi/like) genus or Lactobacillus laccase (Lac(7)C0CCMi /aci(4) decarboxylase, found to be quite suitable for catalytic (1) The alpha keto acid is converted to an aldehyde of the formula (3), such as from hydrazine to 5-FVA. More specifically, a biocatalyst can be used, which has a specificity of SEQ ID NO: 31, SEQ ID NO: 34, and SEQ ID NO: 37. SEQ ID NO: 40, SEQ ID NO: 43, SEQ ID NO: 46, SEQ ID NO: 143, SEQ ID NO: 146, SEQ ID NO: 149, SEQ ID NO: 152, or a decarboxylase of a homologous amino acid sequence of these sequences. Decarboxylase is envisioned. It can be used to prepare an amine of the formula (1) from the α-amino acid of the formula (4). The brewing system is then converted to an amine such as the formula (1), such as 5-fva to 6-ACA. This reaction can be carried out chemically. The amine in the south yield can be obtained by a reductive amination reaction of an aldehyde with ammonia by a hydrogenation catalyst such as Ni immobilized on a Si〇2/Al2〇3 support, as in ep_a 628 535 Or in DE 4 322 065, regarding 9-amino decanoic acid (9-amino geranic acid) and 12-aminododecanoic acid (12-amino lauric acid) If the amine of formula (1) is 6-ACA, 6-ACA can also pass 6-boxic acid

S 25 201127961 Hydrogenation by Pt02, 6-decanoic acid can be prepared by reaction of 5-FVA with hydroxylamine (please refer to FO Ayorinde, EY Nana, PD Nicely, AS Woods, EO Price, CP Nwaonicha J. Am) Oil Chem. Soc. 1997, 74, 531-538 for synthesis of the homologous 12-aminododecanoic acid) In one embodiment, the aldehyde of formula (3) is converted to an amine of formula (1), which is based on (1) an amine group. Providing, and (ii) an aminotransferase, an amino acid dehydrogenase, or another biocatalyst catalyzing the conversion reaction, in the presence of a biocatalytic conversion, in particular, in this embodiment The aminotransferase may be selected from the group consisting of Vibrio fluvialis (1⁄2•? η·σ //wWfl/k), Pseudomonas aeruginosa, Bacillus subtilis (such as D), Bacillus weihenstephanensis, gas: Agrobacterium (Escherichia (7) aminotransferase; β-aminoisobutyric acid from pig kidney: (X-ketoglutarate aminotransferase; β-alanine aminotransferase from rabbit liver; from Perennial scent (MercMria/h perewmX) bud adenyltransferase; 4-aminobutyric acid aminotransferase in the liver, or human, rat or pig brain; β-alanine aminotransferase from rabbit liver; and L-lysine: α-ketoglutaric acid- a group consisting of ε-aminotransferase. If an amino acid dehydrogenase is used, the amino acid dehydrogenase may be particularly selected from the group consisting of Agrokcien'wm iMme/ade/i5 or thermophilic Bacillus licheniformis (Geokci'Z/M·? is a group of lysine 6-dehydrogenase. Another suitable amino acid dehydrogenase may be selected from Bacillus licheniformis jp/iaeWcwjj, Brevibacterium Genus (j^reWhacieWwm sp.), Corynebacterium lysine (Corywe acieriwm g/wiflArn'CMm) or Common Proteus (/Voiewj 26 201127961

Diaminopimelate dehydrogenase; from Acinetobacter ADP1 sp. ADP1) or R. solanacearum (and ahcmia (10)/anacearwmj glutamate dehydrogenase, acting as NADH or NADPH Coenzyme (EC 1.4.1.3; from Salmonella typhimurium (heart ^ glutamate dehydrogenase, with NADPH as a coenzyme (EC)

1.4.1.4); glutamate dehydrogenase (EC 1.4.1.4) from Saccharomyces cerevisiae (Sacc/ia/Omycei cerev!Wae), or Brevibacterium flavum CBrevAacien'Mm //flvwmj from Bacillus licheniformis Cerew5) or leucine dehydrogenase of Bacillus licheniformis (BacH/Ms). In a particular embodiment, the conversion of the aldehyde of formula (3) to the amine of formula (1) is catalyzed by a biocatalyst containing an aminotransferase comprising, for example, SEQ ID NO: 2, SEQ ID NO: 5, No. 8, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 69 ' or any homologous amino acid sequence of these sequences. In a particular embodiment, the alpha-keto acid of formula (2) can be converted to the aldehyde of formula (3) by a chemical reaction. The 2-keto acid is converted to the corresponding aldehyde by an effective chemical decarboxylation reaction, which can be carried out by forming an enamine intermediate, using a secondary amine such as morpholine, and removing c〇 immediately after removing the azeotropic water. 2. Based on the method described in Tetrahedron Lett. 1982, 23(4), 459-462. The olefinamide at the end of the intermediate is then hydrolyzed to the corresponding aldehyde. After that, the aldehyde of the formula (3) can be converted into an amine by biological catalysis, and the reductive amination reaction can be carried out by an aminotransfer reaction in the presence of an aminotransferase or using an amino acid dehydrogenase. 'Or in the presence of another biocatalyst that catalyzes such reactions. The aminotransferase or amino acid dehydrogenase may be particularly selected from the above-mentioned biocatalysts when referring to the conversion of the aldehyde of the formula (3) to the amine of the formula (1) S: 27 201127961. Further, the conversion of the aldehyde of the formula (3) to the amine of the formula (1) can be carried out by a chemical method as mentioned above. In a particular embodiment, formula (2) 〇ί-keto acid can be biocatalyzed to convert to the formula (4) alpha-amino acid, in (1) aminotransferase, amino acid dehydrogenase, or another A biocatalyst that catalyzes such conversion, and (ii) an amine based provider, in the presence. In the present invention, an aminotransferase for converting an α-keto acid into an α-amino acid is particularly selected from the group consisting of an aspartate aminotransferase derived from a porcine heart; and from a rough key spore emblem (iVeMrowora) Or yeast (X-ketoadipate: glutamate aminotransferase; aminotransferase from perennial hawthorn (MercwWaZk perenmij bud; 4-aminobutyric acid amine from E. co. (E·Co(1)) Base transferase; α-aminoadipate aminotransferase from extreme thermophilic bacteria (r/iermw i/iermop/ii/M·?); from the genus IpMrtiriowflZe) or unilateral iron horn Aminotransferase of (Asp/eniMm imi/aieraZe); and aminotransferase from Carob (Oraionk Λ77!々Μβ). In a preferred embodiment, the α-keto acid of formula (2) is converted The aminotransferase of the formula (4) α-amino acid is selected from the group consisting of Vibrio, Pseudomonas (Pmw mcmas), Bacillus licheniformis (5aci7/i«), Legionella (LegiimeZ) /fl), nitrite-like), Neisseria 'Rhodobacter, Escherichia and Rhodopseudomonas (10) Transferase. Especially from 'Bacillus subtilis (10) /), Rhodobacter sphaeroides, Legionella pneumoniae, European nitrite (10) like 28 201127961 ewrapaea), gonorrhea, clove pseudomonas Bacteria (·ΡίβΜί^«ι&lt;?ηαί syringae), Rhodopseudomonas palustris, Ho: Vibrio fluvialis, E. coli (£··ϊ«:/^π·ί:/ π·&lt;2, and the aminotransferase of the organism group consisting of Pseudomonas aeruginosa (the corpse·seMi/omorta·?), was found to be particularly suitable for the catalytic formula (2) conversion of α-keto acid to The reaction of the formula (4) α-amino acid, in particular, conversion of hydrazine to hydrazine. In a specific embodiment, in the case of the conversion of the formula (2) α-keto acid to the formula (4) α-amino acid The aminotransferase used includes, for example, SEQ ID NO: 2, SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27. An amino acid sequence of No. 29, or a homolog of these sequences. In yet another embodiment, a formula of the formula (4) α-amino acid is prepared. The method comprises a biocatalytic reaction carried out in the presence of an enzyme capable of catalyzing a reductive amination reaction, and an ammonia source selected from the group consisting of an oxidoreductase (EC 1·4) acting on the CH_NH2 group of the donor, in particular It is a group consisting of amino acid dehydrogenase (EC 1.4.1). If the amine of formula (1) is 6_ACA, the appropriate amino acid dehydrogenase has 0C-aminopimelate 2-dehydrogenase activity which catalyzes the conversion of hydrazine to hydrazine. Particularly suitable amino acid dehydrogenases are selected from the group consisting of diaminopimelate dehydrogenase (EC 1.4.1.16), glutamate dehydrogenase (EC 1.4.1.3; EC 1.4.1.4) and amine A group consisting of acid dehydrogenase (EC 1Α1.9). In one embodiment, the amino acid dehydrogenase is selected from the group consisting of NAD or NADP as a recipient (EC 1.4.1.3), g 29 201127961, face aminate when classified as glutamate dehydrogenase Dehydrogenase acts with NADP as the recipient (EC 1.4.1.4), leucine dehydrogenase (EC 1.4.1.9), and amine group of diaminopimelate dehydrogenase (Ec 1.4.1.16). A group of acid dehydrogenases. Amino acid dehydrogenase may be particularly derived from Corynebacterium, in particular, Corynebacterium glutamicum, Proteus vulgaris (/Voiews; Agrobacterium, Yu Jiang is Agrobacterium tumefaciens (AgrMacierhm iwme/adend) /Gac^czms, especially Bacillus thermophilus (GeokciZ/milk stearothermophilus) · Acinetobacte, especially Acinetobacter ADP1 (AcfV^ic^acie sp. ADP1); Rawls Phytophthora (/?αΖ仙ma), especially R. solani (such as / gamma solanacearum), Salmonella typhimurium, especially 矣I Salmonella typhimurium CSa/moneZZa, · yeast Bacteria (Siicc/iaromycei), especially Saccharomyces cereviiiae); Brevibacterium (BreWZmcierzMm), especially Brevibacterium flavum (fireWZmciaiMm /Z&lt;3VMm) / and Bacillus (βα&lt;:&quot;/Μ·ϊ), especially It is a group of organisms composed of Bacillus sphaericus (fiac/Z/M·? sp/iaericMsJ, Bacillus licheniformis f5ac///(10) cerew), or Bacillus subtilis. For example, a suitable amino acid dehydrogenase may be selected from the diaminopimelate dehydrogenase from Bacillus along Mi), especially from a globular bud rod mill 5·ρ/ζ&lt;36η·£:ΜΛ· Diaminopimelate dehydrogenase from Brevibacterium (5rev/hcieri'Mm sp.); diaminopimelate dehydrogenase from Corynebacterium (C^rynekcierit/m) 30 201127961 , in particular, from Corynebacterium glutamicum (diaminopimelate dehydrogenase of CorpMacienMm; diaminopimelate dehydrogenase from Proteus (PwieM·?), especially from Proteus vulgaris (P) /OieMSVM/gari diaminopimelate dehydrogenase; sulphate dehydrogenase from Acinetobacter (AczV^iMacie) (EC 1.4.1.3), acting with NADH or NADPH as a coenzyme, especially from Amylin dehydrogenase of Acineto ADP1 (AcineioMcie sp. ADP1); glutamate dehydrogenase from Rolstonella (EC 1.4.1.3), especially from Ralstonia solani (/^·ϊί〇λ·π·β dehydrogenation of sulphate

Enzyme; sulphate dehydrogenase from Salmonella (EC)

1.4.1.4), using NADPH as a coenzyme, especially the face acid dehydrogenase from Salmonella typhimurium (Sa/moweZ/β iyp/iimMWMm); from the yeast 〇Sacc/iarom;ycej) Amino acid dehydrogenase (EC 1.4.1.4), especially amylin dehydrogenase from Saccharomyces cerevisiae (iSWc/mromyceA·cerev/hae); glutamate dehydrogenase from Brevibacterium CBreWfcacie Wwm) (EC 1.4) .1.4), especially from Brevibacterium flavum (5reW glutamate dehydrogenase; and leucine dehydrogenase from Bacillus), especially from Bacillus licheniformis CfiacZZ/M·? cereMi or haystoma Alkaloid dehydrogenase of Bacillus (^^7/«·?. Another suitable amino acid dehydrogenase may be selected from Agrobacterium tumefaciens &gt; or Bacillus thermophilus ( Geobacillus stearothermophilus), a lysine 6-deaerator, or a lysine dehydrogenase derived from a butterfly-shaped dental stem (Wan Shen/Xian·cerewj) or Bacillus subtilis (ββ£:ί7/Μ5) .

ffW 31 201127961 The α-amino acid of the formula (4) prepared according to the process of the present invention can be further used for the preparation of the amine of the formula (1). The human body of the present invention recognizes a formula (4) a pulse-amino acid prepared from the formula (2) α-keto acid, which can be converted into an amine of the formula (1) by a decarboxylation reaction. This reaction can be carried out chemically, for example by heating in a high boiling solvent in the presence of a ketone or aldehyde catalyst. For example, the amino acid is decarboxylated in 150-160 Torr 1-2 ν/ν% cyclohexanone to give a high yield of cyclohexanol, as in H. Hashimoto, Υ. Eda, Y. Osanai, Τ Iwai and S. Aoki in C/iem. 1986, 893-896. A similar method is described in Eur. Pat. Appl. 1586553, 2005 by Daiso, and by SD Brandt, D. Mansell, S. Freeman, IA Fleet, JF Alder J. Pharm. Biomed. Anal 2006, 41, 872-882 ° In addition, the decarboxylation of the formula (4) a-amino acid can be carried out to prepare the amine of the formula (1), which can be biocatalyzed in the presence of decarboxylase or other biocatalyst which catalyzes such decarboxylation. . The decarboxylase may be selected from a decarboxylase which catalyzes the decarboxylation of a-amino acid. An enzyme capable of decarboxylation of an α-amino acid, which is particularly selected from the group consisting of decarboxylase (EC 4_1.1), preferably pyruvate decarboxylase (EC 4.1.1.1), monoamine-glycol Diacid decarboxylase (EC411 2(7), diaminopimelate dehydrogenase (EC 4·1.ι·2〇), branch a_嶋__(EC 4]” 72), which includes 〇t-_ Isovaleryl domain enzyme, and a group consisting of a ketoglutarate (EC 4.1.1.71). Or other suitable de-suppressing enzymes may be specifically selected from oxalic acid detoxification enzymes (EC 4·〇·2) ), grass-brewed acetic acid to lang (EC 4.1.1.3), acetic acid de-slowing enzyme (EC 4.1.1.4), aspartic acid 丨 _ _ 4 丨丄 (1), retic acid 32 201127961

Decarboxylase/leucine decarboxylase (EC 4.L1.14), glutamate decarboxylase (EC 4.1.1.15), 3-hydroxyglutamate decarboxylase (EC 4.1.1.16), avian amine Acid decarboxylase (EC 4.1.1.17), lysine decarboxylase (ec 4.1.1J8), arginine decarboxylase (EC 4.1.1.19), 2-oxoglutarate decarboxylase (EC 4) 71), and a group consisting of diaminobutyrate decarboxylase (EC 4.1.1.86). The decarboxylase is especially a decarboxylase of an organism selected from the group consisting of pumpkin 'such as pumpkin ^ ^ ^ ^ as ^ ^ / / (8): cucumber: yeast: fungus, such as Saccharomyces cerevisiae Physic cerev with ae), Candida albicans //arm·), Haemophilus W.), Kluyveromyces cerevisiae, Rhizopus genus (/?/1 such as y'avamcws) and coarse From the brain of mammals, especially mammals; and bacteria such as Escherichia coli (Ac/zer/c/^, L. lactis (z^cioC0CCMiy/Shenyan), M. tuberculosis; (:0^(^7^/72 plus ^ Guang (: "/ still called, pseudo-single-year-old (·Pjewdcwirniw India.), and the group consisting of ZymomoAiai1 mMi·.) Decarbase can be obtained from Saccharomyces cerevisiae or Zymomcrniw (4). In particular, it can be used from Zymomowiw (Iy acid-degrading enzyme I472A). A phytate decarboxylase from Pseudomonas j. A glutamate decarboxylase from E. coli (£. cW!) can be used. Estrogenate decarboxylase from Escherichia coli, or from Acerospora crawa), Escherichia coli / eprae, Clostridium perfringens (C/o vs. nW/wmper/nVige) Like), 33 201127961 Short Lactobacillus rewi) 'MycokcienMm, Streptococcus (^repbcwcw) or Lactobacillus (LaciococcMij's glutamate decarboxylase. Lactobacillus as a source of facial acid detoxification enzymes (LaciococcM ·?) Examples of species include Lactobacillus lactis (LaciococcMs Zacn'i) B1157, Lactococcus lactis (L (3cc (?cc?ccwWflc) IFPL730, especially Lactococcus lactis var. maltigenes (official name) It is Streptococcus lactis var. maltigenes. The diaminopimelate decarboxylase can be derived from an organism which can synthesize lysine from diaminopimelic acid. Such organisms can be found especially in bacteria, archaea and plants. In particular, the diaminopimelate decarboxylase may be derived from a Gram-negative bacterium such as Escherichia coli (five coi.). A branched alpha-keto acid derived from Lactobacillus reesei (jLaciococcwWflciij) may be used. Go to the betrayal. More special That can be used Rett from Lactococcus lactis (Laciococcwi Z &lt; 3ci⑷ of the acid to the branched α-_ carboxylase and cx- ketoisovalerate to carboxylase. The α-ketoglutarate decarboxylase from Mycohacier/Mm can be used. The present inventors have found that ot-g and glutaric acid de-enzyme (Kgd) derived from Mycobacterium tuberculosis can be used for the formula (4) Conversion of an α-amino acid to an amine of the formula (1), especially converting AAP to 6-ACA. In particular, the present inventors have found that this comprises a decarboxylation of a sequence or a functional analogue as shown in SEQ ID NO:46. An enzyme that catalyzes the conversion of an alpha-amino acid of formula (4) to an amine of formula (1), especially converting hydrazine to 6-ACA. The glutamic acid decarboxylase may be particularly selected from the group consisting of pumpkin mosc /iaia), cucumber, yeast or calf brain; and diaminopimelate decarboxylase (EC 4.1.1.20). 34 201127961 The diaminopimelate decarboxylase may be derived from, for example, an organism in which an amide acid can be synthesized from diaminopimelic acid. This organism can be found especially in bacteria, archaea and plants. In particular, the diaminopimelate decarboxylase may be derived from a Gram-negative bacterium such as Escherichia coli (f.co/ί.). In a particular embodiment, the alpha-keto acid of formula (2) is chemically converted to the alpha-amino acid of formula (4). The alpha-amino acid can be prepared from 〇c-keto acid using a Leuckart-Wallach catalyzed reaction as described for analogous compounds. This reaction was carried out with a solution of cerium ammonium in decyl alcohol using [RhCp*Cl2]2 as a homogeneous catalyst (M. Kitamura, D. Lee, S. Hayashi, S. Tan pimelic acid, M. Yoshimura / Og. C/iem. 2002, 67, 8685-8687). In addition, the Leuckart-Wallach reaction can be carried out in an aqueous solution of methacrylate, using [IrmCp*(bpy)H20]S〇4 as a catalyst, such as S. Ogo, K. Uehara and S. Fukuzumi in J. Am. Chem. Soc. Described in 2004, 126, 3020-3021. The conversion of a-keto acid to an amino acid (rich in the image isomer) can also be carried out by reaction with (non-mirrored) benzylamine followed by passage of the imine intermediate through Pd/C or Pd(0H)2/ Hydrogenation is carried out in C. See, for example, R.G. Hiskey, R.C. Northrop./. Am. C/iem. «Soc· 1961, § 3, 4798. The α-amino acid of the formula (4) is then biocatalyzed to be converted to the amine of the formula (1) in the presence of a decarboxylase or another biocatalyst which can carry out the decarboxylation reaction. This decarboxylase may be particularly selected from the biocatalysts mentioned above, when it is described that the biocatalyst can convert rhodium to 6-ACA. Further, the conversion of the α-amino acid of the formula (4) to the amine of the formula (1) is preferably carried out by a chemical method as described above. 35 201127961 In a particular embodiment, the alpha-keto acid of formula (2) is biologically converted to an aldehyde of formula (3), in the presence of a decarboxylase or another biocatalyst capable of undergoing such decarboxylation, after The analogy is converted to an amine of formula (1) in the presence of an amine group with an aminotransferase, an amino acid dehydrogenase, or other biocatalyst that catalyzes such conversion. The defensin enzyme suitable for these reactions may be specifically selected from the group consisting of the above-mentioned de-slowing _ when the biodeformation of the formula (2) α-keto acid is converted into the aldehyde of the formula (3). Suitable amine transferases or amino acid dehydrogenases for converting the aldehydes of formula (3) may be selected in particular from those mentioned above, when describing the conversion of the aldehydes of formula (3) to the amines of formula (1) Time. In a particular embodiment, the alpha-keto acid of formula (2) is biologically converted to an alpha-amino acid of the formula, in an amine group with an aminotransferase, an amino acid dehydrogenase, or other catalyzed In the presence of such a converted biocatalyst, the amino acid can then be converted to an amine of formula (1) in the presence of a decarboxylase or other biocatalyst that catalyzes such conversion. The enzyme suitable for these reactions may be particularly selected from the group consisting of the above-described aminotransferase, amino acid dehydrogenase and decarboxylase, when the keto acid is biologically converted to each ct-amino acid. The α-keto acid used for the preparation of the amine can in principle be obtained in any manner. For example, the acid can be prepared by a method such as that described by H. Hger from Chem. Ber. 1959, bis 2492-2499. For example, Ακρ can be alkylated by diethyl oxalate, sodium ethoxide is used as a base, and the obtained product is refluxed in a strong acid (2 M HCl), and is self-suppressed as a crystallization method. Prepared by recovering the product. The formula (2) ex-keto acid can also be obtained from natural sources, such as from methanogenesis (201111961 bacteria, S. cerevisiae), or the big wind. For example, the (X-keto acid can be extracted from such organisms, or parts thereof, such as seeds extracted from the wind (also 9C (3r/?W·?. Suitable extraction methods, such as AI) Virtanen and AM Berg in Acta Chemica Scandinavica 1954, 6, 1085-1086, based on the method described in which the amino acid and AKP are extracted from Aspergillus oryzae (Asp/em'Mm) using 70% ethanol, as described above In a particular embodiment, formula (2). OC-keto acid can be prepared via a process comprising a step of reacting a smaller carbon (X-keto acid for carbon chain extension reaction. This carbon chain extension reaction can be extended by hydrazine. 'where x is an integer 丨 or greater. The elongation is preferably extended. The extended reaction can be catalyzed by a biocatalyst. For example, α-ketoadipate (ΑΚΑ) can contain α__glutaric acid (AKG) via inclusion It is prepared by a method of extending the step of converting to α-ketoadipate (ΑΚΑ). The oxime can be prepared by converting α-ketoadipate (AKA) to α-ketopimelic acid, etc. aKg can be obtained via Prepared from a carbon source such as an enzyme's biocatalytic reaction in a manner known in the art. Suitable for the preparation of alpha-keto acids from AKG The catalyst may be specifically selected from the group consisting of catalyzing a-ketoglutaric acid for prolonging the reaction' and/or α-adipate acid for C:-prolongation reaction, and converting to α__pimelic acid, and/or making ~ketone The pimelic acid undergoes a Cr extension reaction to convert to a biocatalyst of a-ketosuberic acid. In a particular embodiment, the alpha-keto acid is prepared to contain a. AksA enzyme or it has a high (4) citrate synthase Active analog;

B_ at least one enzyme is selected from the group consisting of AksD enzyme, AksE enzyme, AksD 37 201127961 enzyme homolog, or it has high (8) aconitase activity &lt;AksE enzyme homolog; and c_ AksF or has 咼(4)- A homologue of isocitrate dehydrogenase activity; wherein η is preferably an integer of 1-5, more preferably an integer u, catalyzed by a biocatalyst. Preferably, the catalyst comprises an enzyme selected from the group consisting of AksD enzyme and its homologue, and an enzyme selected from the group consisting of *AksE enzyme and its homologue. The AksD enzyme or a homolog thereof, and the AhE enzyme generally form a heterodimer.

One or more of AksA, AksD, AksE, AksF enzymes or homologues thereof, which may be found in selected from methanogenic archaea, especially tropococcus (Mei/icmococcw·?), sputum (Mei/umoca/i/ococcM·?), sputum bacillus, thermophilic autotrophic carbs, sputum sclerotium, sputum thermophilic bacteria, and acier in. In a particular embodiment, the biocatalyst converts alpha-ketoglutaric acid (AKG) to 〇c-keto acid, comprising an enzyme system that catalyzes the conversion of oc-ketoglutarate to &amp; ketoadipate , wherein the enzyme system forms part of the alpha-amino adipic acid pathway synthesized from the amino acid. The term "enzyme system" is used herein to refer specifically to a single enzyme or group of enzymes that catalyze a specific conversion reaction. The preparation of alpha-keto acid from AKG may comprise one or more biocatalytic reactions with known or unknown intermediates, such as conversion of AKG to hydrazine, or conversion of ΑΚΑ 38 201127961 to AKP. This system can be present in cells or isolated from cells. The enzyme system may be particularly derived from an organism selected from the group consisting of yeasts, fungi, archaea and fine brines, especially from the genus Campylobacter, Cephalosporium, Paecilomyces (Trichophytum) Aspergillus, leather bacterium (limeoc/iaeie), naked cell shell, black stable bacteria (fAsti/agi?), fission yeast (Sc/i&amp; osacc/iflramyceiJ, yeast (Saccharomyces), Candida, Yersinia, methanol yeast, Kluyveromyces, thermophiles (77^rmMi), abnormal cocci (De/MOCOCCM·?), sputum sputum (/^yraCOCCMi) Sulfolobus, Thermomyces cerevisiae (T/iermococcMd, Mei/iimococcMi, Mei/umocaWococcM·?), Saccharomyces cerevisiae, A. sputum, A. brevis (Afei /mwMreWftacierJ, A. serrata, and a group of thermophilic toxins. In a specific embodiment, a biocatalyst for the preparation of the reaction of the formula (2). α-keto acid from οι-ketoglutaric acid can be catalyzed. , comprising an enzyme system that catalyzes the conversion of alpha-ketoglutarate to CC-ketoadipate, wherein At least one enzyme of the enzyme system is derived from a nitrogen-fixing bacterium selected from the group consisting of cyanobacteria, rhizobiales, gamma-proteobacteria and actinobacteria, especially (Ancth like "α", Micracywii, Synechocystis (C-i&lt;9c_y5. along J, Rhizobium, £39 201127961 Slow-growing Rhizobium, Pseudomonas, AzoioMcier ), a group of Klebsiella and Frankia. These Aks enzyme homologs, as well as examples of genes encoding these enzymes, are provided in Tables 1A and 1B on the next page. 40 201127961 Expression Steps Enzymes Gene gene protein 1 AksA Methanocaldococcus jannashii MJ0503 NP_247479 Methanothermobacter thermoautotropicum ΔΗ ΜΊΉ1630 NP_276742 Methanococcus maripaludis S2 MMP0153 NP_987273 C5 (Methanococcus maripaludis C5) MmarC5_.1522 YP—001098033 Methanococcus maripaludis C7 MmarC7 1153 YP—001330370 M. sphaeroides £&gt;SM3Q9\ (Methanospaera stadtmanae DSM 3091) Msp_0199 YP_447259 M. thermophilic strain PNV) (Methanopyrus kandleri AV19) MK1209 NP_614492 文戌 f虎超#^ATCC35061 (Methanobrevibacter smithii ATCC35061) Msm_0722 YP_001273295 Methane SB (Methanococcus vannielii SB) Mevan_1158 YP_001323668 Klebsiella pneumoniae nifV P05345 Azotobacter vinelandii nifV P05342 Pseudomonas stutzerii nifV ABP79047 严虎破^Nankai 3 (Methanococcus Aeolicus Nankai 3) Maeo")994 YP_001325184 2,3 AksD Methanocaldococcus jannashii MJ1003 NP-247997 f Methanothermobacter thermoautotropicum ΔΗ1386 NP_276502 Methanococcus maripaludis S2 Mmpl480 NP_988600 Methanococcus maripaludis C5 MmarC5_0098 YP—001096630 Methanococcus maripaludis Cl MmarC7_0724 YP_0013299 42 M. sphaeroides DSM3Q9\ (Methanospaera stadtmanae DSM 3091) Msp—1486 YP_448499 Methanopyrus kandleri AV19 MK1440 NP_614723 M. cerevisiae ATCC35061 (Methanobrevibacter smithii ATCC35061) Msm_0723 YP—001273296 Methanococcus SB (Methanococcus Vannielii SB) Mevan-0789 YP_001323307 fO^^t'^Nankai 3 (Methmococcus aeolicus) Nankai 3 Maeo—0311 YP_001324511 Methanosarcina acetivorans MA3085* NP—617978* Methanospirillum hungatei JF-1 Mhun A 1800* YP-503240* high temperature f-aerobacteria (Methanosaeta thermophila PT) Mthe_0788* YP_843217* ======= fM^00DSM 3091 (Methanosphaera stadtmanae DSM 3091) Msp_ll〇〇* YP_448126* base, with protein reference The literature can be found at www.ncbi.nlm.nih.gov/ (Genes/proteins marked * can be obtained on March 2, 2010, others: available on April 15, 2008). 41 201127961

Table IB Steps Enzymes Museum 2 3 AksE Methanocaldococcus ' jannashii Methanothermobacter thermoautotropicum ΔΗ Methanococcus maripaludis S2 Methanococcus maripaludis C5) Methanococcus maripaludis Cl Methanospaera stadtmanae DSM 3091 Methanopyremia PN\9 (Methanopyrus kandleri AV19)

Brevibacterium M. sylvestris KiCCi5Q6\ (Methanobrevibacter smithii ATCC35061) Methanococcus vannielii SB f Me Zhengzhen ankai 3 (Methanococcus aeolicus Nankai 3j Methanosarcina acetivorans f. snails JF1 (Methanospirillum hungatei JF -1) Shaped SM 3091

(Methanosphaera stadtmanae DSM 3091) Methanosaeta thermophila PT Gene MJ1271 ΜΊΉ1387 MMP0381 MmarC5_1257 MmarC7_1379 Msp_1485 MK0781 Msm_0847 Mevan-1368 Maeo_0652 Protein NP_248267 NP-276503 NP_987501 YP_001097769 YP—001330593 YP_448498 NP_614065 YP—001273420 YP_001323877 YP_001324848 MA3751* Mhun_1799* Msp_0374* Mthe 0853* NP_618624* YP-503239* γρ_447420* YP 843282*

AksF Methanocaldococcus jannashii M{Methanothermobacter thermoautotropicum ΔΗ Methanococcus maripaludis S2 Methanococcus maripaludis C5 Methanococcus maripaludis C7) M. cerevisiae DSM 3091 (Methanospaera stadtmanae, DSM 3091) M. thermophilus AV19 (Methanopyrus kandleri AV19) Brevibacterium M. oxysporum ATCC35061 (Methanobrevibacter smithii ATCC35061) Methanococcus vannielii SB JCK ^^ ^^Nankai 3 (Methanococcus aeolicus Nankai 3) Methanosarcina acetivorans M. snails JF, 1 (Methanospirillwn hungatei JF-1) F lin, shape town SM 3091 (Methanosphaera stadtmanae DSM 3091) High temperature f-algal bacteria PT (Methanosaeta thermophila PT) Brevibacterium Acetobacter ATCC 35061 (Methanobrevibacter smithii ATCC 35061) MJ1596 MTH184 MMP0880 MmarC5_0688 MmarC7_0128 Msp_0674 MK0782 Msm_0373 Mevan_0040 Maeo_1484 MA3748* Mhun_1797* Msp_0674* Mthe_0855* Msm 1298* NP-248605 NP_275327 NP988000 YP001097214 YP_001329349 YP_447715 NP_614066 YP001272946 YP_001322567 YP_001325672 NP_618621* YP_503237* YP 447715* YP 001273871* References for genes and proteins can be found at www.ncbi.nlm.nih.gov/ (labeled: 曰Get' Other: Available in April 2008 Acquired on the 15th). : Gene/Protein: Available on March 2, 2010 2, 2011, 2011, 961 especially if the method uses one of the serial numbers m, 114, 117, l2〇, 123, 126, 129, 132, 135, 138 or 141 Good results can be obtained by a variety of enzyme methods. Thus, in a preferred embodiment, the invention relates to a method of preparing a compound containing an amine group, wherein one or more enzymes or homologs thereof are used. The present invention also provides a novel gene whose codon pair is optimized, especially for Escherichia coli (the heart is better for Escherichia coli (4) c〇i&quot; therefore the present invention is more related to a polynucleoside An acid comprising any of the following sequences: SEQ ID NO: 112, 115, 118, 121, 124, 127, 130, 133, 136 139 and its analog b analog, similar and identical in Escherichia (five host cells) Or preferred amounts of expression. As far as the inventors are aware, these polynucleotides are not naturally produced. As for those naturally occurring polynucleotides, they are said to be isolated from organisms in which they are naturally produced. The 〇tS acid can be prepared by converting a fatty (di) acid to a keto acid, which can be catalyzed using a biocatalyst, especially a heterologous biocatalyst. The biocatalyst can be a xenogeneic cell. Containing one or more nucleic acid sequences encoding one or more enzymes having catalytic activity for converting a fatty acid to an alpha keto acid. In the preparation of an alpha-acid from a fatty di-acid, one of the carboxylic acid groups of the acid is converted to An aldehyde group to produce (X-keto acid. For example The 2 hydroxysuccinic acid can be converted to an α-keto acid in this manner. For example, AKp can be prepared from 2 hydroxy pimelic acid in this manner. The conversion can include a hydroxylation reaction of the dicarboxylic acid and conversion to a 2-hydroxyl group. a step of alkanoic acid (or alpha-hydroxydicarboxylic acid) which can be catalyzed by a hydroxylase

S 43 201127961. The conversion can be carried out in the step of including the step of oxidizing 2-M cyanoic acid to α-keto acid. The hybridization reaction can be catalyzed by a biocatalyst comprising a oxidoreductase (EC 1,14) selected from the group consisting of a donor (using oxime 2 as an oxidant) and adding or reducing oxygen, acting on cMcH2 The oxidoreductase (EC 1.17) has a hydrolyzing enzyme (ec 3) having pimelic acid hydrolase activity, and one of a group of hydrolyzing enzymes (EC3) having pimelic acid-2 &quot;monooxygenase activity. Further, as the catalysis of the hydroxylation reaction of didecanoic acid, an enzyme selected from the group consisting of the amino acid sequence of SEQ ID NO: 9()_1()9, or any of these sequences can be used. The oxidation can be catalyzed by a biocatalyst comprising an enzyme selected from the group consisting of an oxidoreductase (EC丨丨3) with oxygen as a recipient, such as lactate oxidase or another hydroxy acid oxidase; -L-lactate dehydrogenase (EC 1.1.1.27); - thiopyruvate reductase 'β-pyruvylate reductase; NADH: transpyruvate reductase, and D_glycerate dehydrogenase (EC11) 81); - Malate dehydrogenase [NADP+], NADP+-malic acid enzyme, NADP+-malic acid deoxygenase (in the test of the amine gland), malic acid NADP dehydrogenase NADP+ malate dehydrogenase; NADp+_linked malate dehydrogenase, and malate dehydrogenase (NADp+) (EC 1.1.1.82); 3-isopropyl malate dehydrogenase, β_ Isopropyl malate enzyme; β_isopropyl malate dehydrogenase; threo_Ds_3_isopropyl malate dehydrogenase, 3-carboxy-2-hydroxy-4-methyl-valeric acid: NAD+ oxidation Reductase 44 201127961 1.1.1.85); - tartaric acid dehydrogenase, meso-tartaric acid dehydrogenase (EC ^.: 1.93); - (R)-2-yl-fatty acid dehydrogenase (Ed 1丄98) ; -(S)-2-thiol-fatty acid dehydrogenation Enzyme (Ec丨丨1 99); -1.1.1.172 2-oxo-adipate reductase, 2-ketoadipate reductase, α-adipate reductase, 2-S and adipic acid reductase. -2-light valerate dehydrogenase (EC 1.1.99.2); and -D-2-hydroxy-g complex dehydrogenase (EC 1199.6). This enzyme may be derived in particular from the organism, selected from the group consisting of the human and the balloon (Aer〇c〇cct^, especially the human subfamily, such as humans (good, and the green balloon (Aerococcws). For example Human (Homo sapiens) light acid oxidase (glycolate oxidase) or lactate oxidase _ green balloon bacteria group. 2-Phosphate fatty acid oxidation to α-keto acid reaction, can be catalyzed by an enzyme The enzyme comprises an amino acid sequence such as SEQ ID NO: 85 or 88 (which encodes the nucleotide sequences of SEQ ID NOs: 84 and 86' and SEQ ID NOs: 87 and 89, respectively), or homologs of these sequences. Preferably used in the preparation. The α__acid biocatalyst system-enzyme system can be used to prepare α-keto acid from a suitable carbon source, which can be converted to α_decanoic acid as achieved by a fermentation reaction of the carbon source. In the middle, α-keto acid is prepared by converting whole cells into α-keto acid using whole-cell biotransformation. It is known that dicarboxylic acids can be prepared by oxidative cleavage of long-chain fatty acids. Therefore, such fatty acids can be used as carbon sources. If borrowed 45 £ 201127961 by Providing vegetable oil, fatty acid vinegar (biodiesel) or the like to the biocatalyst of the method of the invention (especially in the case where it is a host cell). For example, the host cell may be selected from those naturally containing such a system - such as Escherichia coli (five co/i·) or B. sphaericus (5. __, or a host cell which can be genetically modified to obtain this system. 2-dibasic fatty acid can also be a naturally occurring compound, and can be used as a non-苇The source of slavery (s|·see references C/iemMoc/zem. 2009 Aug 17;10(12)··2003-10). The biosynthesis of 2-hydroxy and iso-average fatty acids, myxobacteda The sphingolipid (Sphing 〇 Upid) is related (Ring MW' Schwar G, Bode ΗΒ·). When using 2-mercapto fatty acid as a starting material, 'there is no need to bribe (4), the acid can be obtained. _The carbon source In particular, it may comprise, at least, a compound selected from the group consisting of a unit alcohol, a polysaccharide, a carbon dioxide, a fatty acid, a glycerol, a tri- and a di-indenyl group, and a mixture comprising these compounds. : The single: alcohol = methanol and ethanol. Appropriate diversity Alcohols include glycerin and sugar-based fatty acids or glycerides, which can be provided in the form of edible oils, and the car is a plant source. It is used because _, because sugars can usually be regenerated by biological means, S pays The agricultural product is preferably agricultural waste. Preferably, the saccharide used is selected from the group consisting of glucosinolate, glucosamine, fructose, sucrose, lactose, agarose, and sucrose. a vouching enzyme containing glucose, and a multi-enzyme containing glucose. If desired, an indoleamine is formed according to the present invention. This may have been: a formula of (1) an amine which can be known by a known method via a ring T The foundation is reached. For example, 46 201127961 6-ACA can be converted to form caprolactam, as described in us-A 6,194,572. The reaction conditions of any of the biocatalytic steps of the present invention can be selected depending on the conditions known for the biocatalyst, especially the enzymes disclosed herein and optionally used in some conventional experiments. In particular, the pH of the reaction medium used can be selected from a wide range of limitations as long as the biocatalyst is active under the pH conditions. Alkaline, neutral or acidic conditions can be used, depending on the type of biocatalyst and other factors. If the method involves the use of a microorganism, such as an enzyme that can be used to express a method of catalyzing the invention, the pH selected should be such that the microorganism can perform its intended function. The pH may be selected in particular from 4 pH units below neutral pH and 2 pH units above neutral pH, such as between pH 3 and pH 9 at 25 C. In the system. In a system, water is the sole or primary solvent (&gt;5〇wt.%, especially >9〇wt%, based on total liquid), where, for example, a small amount is dissolved (&lt;5〇 The wt%, especially &lt;chuan wt.%, based on the total liquid weight) of alcohol or another solvent (e.g., as a carbon source), and this concentration allows the microorganism to remain active, the system can be considered an aqueous solution. In particular, if the mother fungus and/or the money is preferably acidic, the pH may range from pH 3 to pH 8, based on a 25 t: substantially aqueous solution system. If desired, the pH can be adjusted using acid and/or assay, or buffered with a suitable acid and assay composition. In principle, the resting time can be selected in a wide range as long as the biocatalyst exhibits foot activity and/or growth filaments. This includes aerobic, microaerobic, oxygen limited and anaerobic conditions. 1 The anaerobic conditions described herein are defined as being free of any oxygen, or wherein 47 201127961 is essentially a biocatalyst, especially a microorganism, and the conditions of no oxygen consumption generally correspond to an oxygen consumption of less than 5 mmol/Lh, especially The gas, gas consumption, and the aerobic condition are those in which the amount of oxygen dissolved in the culture solution is sufficient, so that the length of the length loss is unlimited, and the consumption of the milk gas can be at least 1 〇h ' _ better than 20 mmol A.h, More preferably greater than 50 inm 〇 l / lh, optimally greater than 100 mmol / lh. The oxygen-restricted condition is defined as the condition that the oxygen consumption is limited to the case where oxygen is transported from the gas to the liquid, and the limit of the oxygen-restricted condition is determined according to the upper limit of the anaerobic condition, that is, usually at least 丨, especially at least 2.5 mmol/lh, or at least 5 mm〇l/lh. The upper limit of the oxygen-restricted condition is the lower limit of aerobic conditions, i.e., less than 1 〇〇 mm 〇 1 Ah, less than mmol / l.h, less than 20 mm 〇 l / l, h, or less than 10 mm 〇 1 / Lh. This condition is aerobic 'anaerobic or oxygen limited, depending on the conditions at which the process is carried out, in particular the amount and composition of the influent gas stream, the actual mixing/mass transfer characteristics of the device used, the type of microorganism used, and the microorganism density. In principle, the temperature used is not critical as long as the biocatalyst, in particular the enzyme, exhibits substantial activity. In general, the temperature may be at least o°c, especially at least 2, particularly preferably at least 2 (rc. The maximum temperature desired depends on the biocatalyst. In general, this maximum temperature is known in the art, such as commercial The information on the available biocatalyst product descriptions may be determined or may be determined by general techniques. The information is disclosed herein. The temperature is usually 9G ° C or lower, preferably scratched or lower, especially pits. 48 201127961 or lower, more preferably 40t or lower. Especially if the biocatalytic reaction is carried out outside the host organism, a high concentration of organic solvent-containing reaction medium can be used (eg greater than 50% or greater than 90 wt· %) 'If the enzyme used maintains sufficient activity in this medium. In a preferred method, the amine of formula (1) is a whole cell biotransformation method that will accept or form an intermediate of amines. Substance (A-form (2) (X-rigid, acid of formula (3), and amino acid of (4)) are converted into amines, wherein the package 3 U organisms can be manufactured - or a plurality of a biocatalyst (usually - or a plurality of enzymes) that catalyzes the conversion of the organism, The plurality of biocatalysts are selected from the group consisting of a biocatalyst capable of catalyzing the conversion of α-amino acid, a biocatalyst capable of catalyzing the conversion of α-amino acid to _, a biocatalyst capable of catalyzing, and catalyzing Amines = group of agents, group + preferred embodiment 2 = carboxylase, and / or at least one 醆 +, the initial carbon source contains mr de(four) and silk transferase. Polyol, pure, - oxidation f. The substance 'selected from the unit alcohol, these compounds, the W, the material, sighs include glycerides and ethanol. Suitable polyols include glycerol: and alcohols including sterols: yes, class, because _通;=plant source. Although the source is large, such as agricultural products, 斡π is derived from biological renewables and is selected from (four) agricultural waste. It is better to make powder, cellulose and hemicellulose q 1 ' Sugar, lactose, sucrose, spleen glucose, and polysaccharides containing, preferably, glucosinolate. It can be provided using two or two, preferably plant-derived glucose, including ε ^T~r 49 201127961 - cells , especially recombinant cells, which comprise one or more catalyzed methods A biocatalyst (usually one or more enzymes) for a certain reaction step can be constructed using molecular biotechnology, which is known in the art. For example, one or more biocatalysts are produced in a recombinant cell (which can be Heterologous system), the technique can be used to provide a vector (eg, a recombinant vector) comprising one or more genes encoding the one or more biocatalysts. One or more vectors can be used, each of which One or more such genes are included.The vector may comprise - or a plurality of regulatory elements, such as one or more promoters, operably linked to a gene encoding a biocatalyst. The term "operative linkage" is intended to create a functional relationship with a poly(tetra) acid region ements) (a vine-acid sequence) when the nucleic acid sequence is linked to another nucleus in a functional relationship. It is called "operating connection." For example, a promoter or an enhanced translation of an operability linkage to a coding sequence, if it affects the repertoire of the coding sequence, its function is: trans: = because of transcription 'it is located in the transcription of the gene The starting position: = two structurally - polymerase binding is limited, the m...dna phase, including but not limited to the conversion (four), the financial state and any other technically known to promote via the position of the direct protein ^ The nuclear (four) phase of the sub-touch. "The group of chicks have active activity under the environment and the hair piece. Promoters are promoters that promote the activity of the promoters under the control of environmental and developmental conditions. The term "homologous 50 201127961", when used to refer to a relationship between a particular (recombinant) nucleic acid or a nuclear acid molecule, a plaque host organism or a host cell, should be 昤铉v 羯 essentially the nucleic acid or The compound peptide/knife is made of a host cell of the same species or an organism, preferably the same variety or strain. It can be used to make the enzyme used in the present invention, /, which is the nucleic acid sequence of the aminotransferase, the amino acid dehydrogenase, or the decarboxylase gravel as described above, to achieve a predetermined amount of promoter. Same as the nucleic acid sequence encoding the enzyme to be spent

The source, or a nucleic acid sequence (coding sequence) operably linked thereto, is heterologous. Preferably, the promoter is homologous, i.e., host cell endogenous. X If a heterologous promoter is used (relative to a nucleic acid sequence encoding an enzyme of interest), the heterologous promoter preferably produces a higher stable transcript comprising the coding sequence (or more transcription can be made) Molecules, that is, mRNA molecules produced per unit time, are compared with the promoters of the money. Suitable promoters herein include both constitutive and inducible natural promoters, as well as genetically modified promoters, which are well known to those skilled in the art. A "strong constitutive promoter" is one that produces a high frequency of mRNA and is compared to the original host cell. In Gram's yang, this strong constitutive promoter of the towel includes SP01-26, SP01-15, veg, p; yc (pyruvate de-negotase promoter and izmyE. Gram-positive bacteria can induce and promote Sub-examples include the IPTG-inducible PsPac promoter and the wood-induced PxylA promoter. The constitutive and inducible promoters in Gram-negative bacteria include, but are not limited to, tac 'tet, trp-tet ' Ipp, Iac, ipp-iac, laclq, T7, 51% 201127961 Γ5, Γ3, fine, ara (P(10)), sp6, λ Pr, and χ Pl. (Filament) fungi promoters are known in the art, For example, glucose heart phosphate dehydrogenase gpi / A promoter 'protease promoters such as corpse 卬 eight, corpse B, corpse e /? C glucose; Diankou _; coffee a promoter, temple powder enzymes (four) eight, Promoter, catalase (3) 汲 or (3) recognize promoter, glucose oxidase exhaust; ^ promoter, β-galactosidase / acA promoter, α_glucosidase 叩丨8 promoter, translation elongation factor Ie/A promoter, xylanase promoter, such as, ▲ Β, xhC, x / nD, cellulase promoter such as coffee a, eg / B, c from Α, transcript For facilitators such as 'A, creA, x/nR, /?acC, sound T, or another promoter, refer to the NCBI webpage (http://www.ncbi.nlm.nih.gov/entrez/) terminology "heterologous" when used in a nucleic acid (pNA or RNA) or protein, is a protein or nucleic acid that is not naturally part of an organism, cell, gene, or DNA or RNA sequence. Exist or found in a cell' or at a genomic or DNA or RNA sequence position, unlike a naturally occurring one. A heterologous nucleic acid or protein is not an endogenous person to be introduced into a cell, but may be obtained from another cell or Manufactured recombinantly. Generally, although not necessarily, the nucleic acid encodes a protein which is normally transcribed or expressed by the cell. Typically, the exogenous RNA encodes an abnormal expression in the presence of the exogenous RNA. Proteins in cells. Heterologous nucleic acids and proteins may also be referred to as foreign nucleic acids or proteins. Any heterologous or foreign nucleic acid or protein well known to those skilled in the art is encompassed by respect to the cells in which they are expressed. Heterologous nucleic acid or protein The method of the present invention can be carried out in a host organism, which is a novel method. 52 201127961 Accordingly, the present invention is also related to a host cell, the cell packet, or a plurality of biocatalysts which catalyze at least one step of the method of the invention, Is at least the step of catalyzing the conversion of the alpha-keto acid of formula (2), (3) and (4), or the conversion of the amine of formula (1) with an amino acid. The invention is also related to a novel carrier or genes, encoding - Or a plurality of catalyzed agents: at least two enzymes, especially catalyzed, converted to amines to a stimuli, and associated with a novel host cell, or a plurality of bases' The method of the present invention at least: the step of converting the enzyme into a 6_ACA, in particular, a step of catalyzing the conversion of an acid to an amine (the - or a plurality of genes may be formed - or a plurality of carriers). Especially on

In a particular embodiment, the host cell of the invention is a recombinant cell comprising a nucleic acid sequence encoding a catalyzable amine-transfer reaction or a reductive amination reaction to produce an amine from the alpha keto acid. The biocatalyst of the basic acid can be a part of the vector, or can be inserted into the chromosome (10) A. In particular, the host cell of the present invention can comprise at least one nucleic acid sequence, in particular at least two nucleic acid sequences, selected from the coding a nucleic acid sequence of an enzyme for ketoacid dehydrogenase activity, a nucleic acid sequence encoding an enzyme having an aldehyde group (such as $methionine), and an amino acid sequence encoding a ct_-acid aminotransferase A nucleic acid sequence of an enzyme, a nuclear ruthenium sequence encoding an enzyme having ketoacid dehydrogenase activity, and a nuclear I sequence encoding an enzyme having a-amino acid decarboxylase activity. Of these sequences, one or more are particularly preferred - or both are recombinant sequences. 53 201127961 In a preferred embodiment, the host cell, generally a recombinant host cell, or a vector of the invention, may comprise a nucleic acid sequence encoding at least one organism having (X-keto acid decarboxylase activity) a catalyst, and/or at least one nucleic acid sequence selected from the group consisting of a biocatalyst encoding an aldehyde group (e.g., 5-mercaptovalerate) aminotransferase activity. In this embodiment, the coding device (X- A nucleic acid of an enzyme of keto acid decarboxylase activity, particularly comprising a nucleic acid sequence encoding SEQ ID NO: 31, SEQ ID NO: 34, SEQ ID NO: 37, SEQ ID NO: 40, SEQ ID NO: 43, SEQ ID NO: 46, SEQ ID NO: 143, SEQ ID NO: 146. Amino acid sequence of SEQ ID NO: 149, SEQ ID NO: 152, or a homologue of these sequences, and/or a nucleic acid sequence encoding an enzyme having aldehyde aminotransferase activity, particularly comprising a nucleic acid sequence. The amino acid sequence of SEQ ID NO: 2, SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 69 or a homolog thereof may be encoded. One or more of the nucleic acid sequences may form one or more recombinations. One part of the carrier. In another preferred embodiment Wherein the vector or host cell comprises a nucleic acid sequence encoding an enzyme having (X-keto acid aminotransferase activity, and/or a nucleic acid encoding an enzyme having a-amino acid decarboxylase activity. a nucleic acid sequence of an enzyme of ketoacid aminotransferase activity, particularly comprising a nucleic acid sequence encoding SEQ ID NO: 2, SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21. , SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, amino acid sequence of SEQ ID NO: 29 or a homolog thereof. One or more of the nucleic acid sequences may form part of one or more recombinant vectors. In one embodiment, the host cell of the present invention comprises a nucleic acid sequence encoding an enzyme having alpha-aminopimelate 2-dehydrogenase activity, and a core 54 201127961 acid sequence encoding Ot-aminopimelic acid An enzyme for decarboxylase activity. In a preferred embodiment, the host cell of the present invention comprises a nucleic acid sequence encoding an enzyme having 6-aminohexanoic acid 6-dehydrogenase activity, and a nucleic acid sequence encoding the same Enzyme of oc-ketopimelate decarboxylase activity. The invention One or more genes of the main cell or vector may be particularly selected from the gene encoding the above enzyme. In a particularly preferred embodiment, the host cell is a recombinant cell comprising at least one nucleic acid sequence selected from the sequence number. 1. Serial number 3, serial number 4, serial number 6, serial number 7, serial number 11, serial number 13, serial number 14, serial number 16, serial number 18, serial number 20, serial number 22, serial number 24, SEQ ID NO: 26, SEQ ID NO: 28, SEQ ID NO: 30, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 44 45. A population consisting of SEQ ID NO: 47, SEQ ID NO: 64, SEQ ID NO: 66, SEQ ID NO: 68, SEQ ID NO: 142, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 147, SEQ ID NO: 151, and functional analogs thereof. A nucleic acid sequence encoding an enzyme having an aldehyde (e.g., 5-FVA) aminotransferase activity, particularly selected from the group consisting of SEQ ID NO: 1 '3, 4'6, 7, 64, 66, 68, and the function of these sequences A group of analogues. A nucleic acid sequence encoding an enzyme having oc-keto acid decarboxylase activity, which is particularly selected from the group consisting of 30, 32, 33, 35, 36, 38, 39, 41, 42, 44, 45, 47, And a population of functional analogs of these sequences. In a preferred embodiment, the host cell comprises a nucleic acid sequence encoding an enzyme which catalyzes the conversion of the (4) oc-amino acid to the formula (2) 06-keto acid, the nucleic acid sequence being: : 1, 3, 7, 11, 13, 14, 16, 18, 20, 55 201127961 22, 24, 26, 28' or a function thereof - which may be a wild type or a non-wild type sequence. In a specific embodiment, the host cell comprises at least one nucleic acid sequence encoding a biocatalyst having OC-aminopimelate decarboxylase activity, which may be host cell homology or heterologous. In particular, the biocatalyst may be selected from the group consisting of de-faced (EC 4.1.1), especially her acid de-suppressing S^(EC 4.1.1.15) monoaminoheptanoic acid de-septase (Ec 4 1 1 20), glutamic acid 1-deactivating enzyme (EC 4.1.1.11) 'branches α__ de-enzyme, valeric acid decarboxylase, (X-ketoglutarate decarboxylase, pyruvate a group consisting of chymase (EC 4丄11:) and grasshopper acetic acid decarboxylase (E_C. 4.U.3). In a special case, the host cell contains one or more enzymes that catalyze The reaction of α-keto acid (e.g., AKp or AKS) is formed by AKG (as described above). An enzyme system which forms a partial pathway of α-aminoadipate which is synthesized from the amino acid can be used. The term 'enzyme system' is used. Herein, especially refers to a single enzyme or group of enzymes that catalyze a specific conversion reaction. The conversion may comprise one or more chemical reactions 'known or unknown intermediates, such as from AKG to AKA, from AKA to AKp, or Converted from Ακρ to AKS. This system can be isolated in cells or isolated from cells. It is known that aminotransferases have a wide range of U. If present, hopefully The activity of - or more of such enzymes in low host cells allows conversion of AKA or AKp to alpha-amino adipic acid (AAA) or (X-aminopimelic acid can be reduced while maintaining other amino acids Or an intracellular component, such as a catalytic function associated with the synthesis of a particular compound of interest. In addition, it is preferred that the pro-primor cell lacks other enzyme activities that cause AKA to be converted to an undesirable by-product. 56 201127961 Preferably, the organism The host or host cell can convert the fatty acid to the same acid and is carried out as described above. In a specific embodiment, the host cell encapsulates - or a plurality of enzymes which catalyze the formation of the formula (2) alpha-keto acid. An enzyme system that forms part of the amino acid degradation pathway can be used, as described by Atsumi S, Hanai T, Liao jc Nature. 2008 Jan 3; 451 (7174): 86-9, or in a suitable manner, such as with hydroxy And/or oxidative means, an enzyme system that modifies fatty acids. The enzyme system can be a natural producer of the host cell, or a heterologous alteration. For example, the system can be isolated from cells other than the host cell. The term &quot;enzyme system Used in In particular, it refers to a single enzyme or group of enzymes that catalyze a specific conversion reaction. The conversion may comprise one or more chemical reactions with known or unknown intermediates. It is suitable for the preparation of the formula (4) α_ by whole-cell biotransformation. Preferred host cells for amino acids, encoding one or more biocatalysts, catalyze at least one reaction step for the preparation of keto-peptimeic acid from alpha-ketoglutaric acid. Suitable biocatalysts are as described above for the preparation of alpha- The keto acid is the user β. For example, the host cell may be selected from bacteria, yeast or fungi. In particular, the host cell may be selected from the group consisting of Pasta, Phyllobacter (PemWZZ/Mm), yeast ("Sacc/ia/Omyces", Kluyveromyce, pichia, Candida, Candida (opening αηπηΜα·), Bacillus, Corynebacterium, Pseudomonas, glucosinolate fG/wccmo&amp; flcier), sputum bacillus /Mm), A. sinensis (like a group consisting of A. serrata and Escherichia. This usually colonizes and expresses one or more of the above-mentioned coding nucleic acid sequences. Especially 疋 '5 Hai A host strain suitable for biosynthesis of an amine of formula (1) and a host cell, which may be selected from Escherichia coli, Bacillus subtilis (fiac/Z/wi like), Bacillus amyloliquefaciens amVWgwe/aci'ew) , Corynebacterium glutamicum g/Miami'cwm), black fungus (Α·57^Γρ7/Μ&gt;5 m'ger), Penicillium chrysogenum, S. cerevisiae cereWdfle), Polymorphic Hanillella (HansenwZa /7〇/卿0叩/^, Candida albicans (Can also be MMc(10) hook, Kluyverella cerevisiae ( Along the M&gt;w/Om);caZiu: along), xylose-fermenting yeast (corpse; ^5 rushed out j, Basil yeast CPic/π'α household 3 pairs of gentlemen), smoldering thermophiles (Methanothermobacter thermoautotropicum/^W) ' ,i; f (Methanococcus maripaludis), Methanococcus (Methanosarcina vo/_), A. sinensis (4), _, Pasteurella methanei (Methanosarcina and the group of host cells composed of Mei/ianc^ardna rnazei'. The d host cell can in principle be a naturally occurring organism, or a genetically engineered organism. Genetically engineered, using a mutation screen or a strategy known in the art. In a particular embodiment, the host cell can naturally comprise (or can be made) one or more enzymes suitable for catalyzing the methods of the invention. a reaction step, such as a reaction step having a group selected from the group consisting of a degrading agent, an aminotransferase and an amino acid dehydrogenase, or a plurality of activities, which catalyze the method of the invention. For example, Escherichia coli can be used. Made naturally The fortune method t can be used to transfer the reverse-synthesis enzyme. It can also provide a recombinant host cell, which has (iv) a recombinant gene of the method of the present invention, which can be catalyzed by the invention. The method - the reaction step depends on both of the recombinant genes. For example, the host cell may be selected from the group consisting of Corynebacterium (Ci) r3; n6acid_ 咖 · · _) species 'especially Corynebacterium glutamicum (four) brain coffee _ chemical ·), intestinal bacteria, especially Escherichia coli ( Such as ~ café (9) ..., wearing cell cup halogen, especially Bacillus subtilis (Wan Xian Office · $) and Bacillus methanolicus (anti-mei / iimWcMi), and yeast, especially scorpion wine yeast production (tSacc / iGTOmycej cereWiiae Particularly suitable for C. glutamicum (C Wwiam/cMm) or Bacillus methanolicus (and mei/zawWci^) strains, which have been developed as an industrial process for the production of lysine. The present invention is more related to a microorganism. , which may be a wild-type microorganism isolated from the natural environment, or a recombinant microorganism, comprising a substrate selected from the group consisting of SEQ ID NO: 3, SEQ ID NO: 6, SEQ ID NO: 13, SEQ ID NO: 32, SEQ ID NO: 35, SEQ ID NO: 41 44. A nucleic acid sequence of SEQ ID NO: 47, and a DNA of a functional analog thereof. A functional analog of a nucleotide sequence, which refers to a nucleotide sequence encoding the same amino acid sequence, or encoding the nucleotide a homolog of a sequence. In particular, a preferred functional class A nucleotide sequence having similar, identical or preferred expression levels in a host cell of interest, referred to as a functional class 5 59 201127961. The invention is more related to a polynucleotide or carrier, A nucleic acid sequence comprising, for example, a SEQ ID NO: 3, SEQ ID NO: 6, SEQ ID NO: 13, SEQ ID NO: 32, SEQ ID NO: 35, SEQ ID NO: 41, SEQ ID NO: 44, SEQ ID NO: 47, and non-wild type functional analogs thereof. Preferably, the polynucleotide or vector is such that the host cell, particularly the E. coli host cell or another host cell, can catalyze at least one reaction step of converting AKP to 6-ACA in a high yield manner, corresponding to the corresponding wild-type gene phase. Optionally, the polynucleotide or vector comprises one or more nucleic acid sequences encoding one or more other biocatalysts suitable for catalyzing at least one step of the method of the invention, particularly one or more of the above-described biocatalysts. More particularly, the present invention relates to a process for the preparation of an alpha-amino acid comprising converting a 5-acid to an alpha-amino acid which is catalyzed by a biocatalyst. A biocatalyst having the above-described aminotransferase activity or reductive amination activity. 4 As described above, the oxime amino acid can be used to prepare an amine of the formula (1). Further, the α-amino acid can be used, for example, in chemistry. Or biochemical research or as this compound' as used in preparative or analytical grade separation techniques, such as liquid 'buffered capillary electrophoresis. *Objective chromatography or, in addition, the amino acid of formula (4) prepared according to the invention, one step to prepare another compound For example, further, for example, a cc-amino acid convertible internal guanamine compound such as an internal amine can be obtained, for example, by cyclization, (iv) an amine, a shape, a #', and the following formula 60 (5) ), 201127961

H-C-A I

H and the examples are exposed to high temperatures. Where A is as defined in equations (1) and (2). As mentioned above, it can be shown that AAP can be chemically converted to caprolactam, which is not limited by theory. It should be considered that in this reaction, ^shelamine 6-ACA can be shaped: CO〇H, which can also be short-lived. Intermediate product. Further, the amine of the formula (1) has a compound for the preparation of a cyclic compound. Further, the amine of the formula (1) can be used in a method for producing a polymer. The method comprises an amine or a mixture of compounds containing at least one amine of formula (1),

The polymerization reaction is carried out, optionally in the presence of one or more of the genus D-, and the presence of the precursor. In contrast, the amine of the formula (1) can be polymerized into a polyamine. For example, the formula (1) has R = Amines of NH2 (diamine), which can be polymerized into polyamines, such as by copolymerization of T-mono-acids. The formula (1) amines (amino acids) with bCOOH can be selectively combined with - or more The other monomers, such as other amino acids, are polymerized into polyamines. ~ The polymerization reaction can be introduced into host cells, such as the host cells described above, and the invention will be described in detail by the following examples. Examples of Amino Compounds 1.1 General Methods Molecular and Gene Technology Standards Genes and Molecular Biology are generally known in the art and have been previously described (Maniatis et al. 1982 "Molecular cloning: a laboratory £ 61 201127961 manual'' · Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Miller 1972 "Experiments in molecular genetics", Cold Spring Harbor Laboratory, Cold Spring Harbor; Sambrook and Russell 2001 &quot;Molecular cloning: a laboratory manual" (3 Rd edition), Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press; F. Ausubel et al, eds., &quot;Current protocols in molecular biology&quot;, Green Publishing and Wiley Interscience, New York 1987). plastids and colonies pBAD /Myc-His C was obtained from Invitrogen (Carlsbad, CA, USA). The plastid pBAD/M; yc-His-DEST was constructed as described in WO2005/068643 for protein expression. £.co//TOP 10 ( Invitrogen, Carlsbad, CA, USA) is used for all selection steps and is used to express the target gene. Culture medium LB medium (10 g/tryptone, 5 g/Ι yeast extract, 5 g/1 NaCl) ) is used to culture E. coli. Antibiotics (5 〇 pg/mi carbenicillin) were added to maintain the plastids. Under the control of the PBAD promoter of the pBAD/Myc-His-DEST-derived plastid, a final concentration of 0.2% (w/v) of L-arabinose was added to induce gene expression. Plasm identification The system with different genes is identified by genetic, biochemical and/or phenotypic techniques known in the art, such as resistance to antibiotics through transformation, PCR diagnostic analysis of transformations, or purification of plastid DNA. , restriction enzyme analysis of purified plastid DNA, or DNA sequence analysis. 62 201127961 HPLC-MS analysis for determining 5-FVA 5-FVA was measured using selective reaction monitoring (SRM)-MS detection to measure the shift of m/z 129+83. The 5-FVA concentration was calculated by measuring the area under the peak of the 5-FVA spike, which was rushed at about 6 minutes. It is calibrated using an external standard process. All LC-MS experiments were performed on an Agilent 1200 LC system consisting of a quaternary pump, an autosampler and a column oven, and combined with an Agilent 6410 QQQ triple quadrupole MS handle. LC conditions: Column 50 X 4.6 mm Nucleosil C18, 5 μηι (Machery &amp; / Nagel) pre-column, shank to 250 x 4.6 mm id· - Prevail Cl8, 5 μιη (Alltech) Column temperature: room temperature Lift: A: Water containing 0.1% formic acid B: acetonitrile with 0.1% formic acid gradient: time (minutes) 0 6 % of extract B 10 50 6.1 10 11 10 Flow rate: 1.2 ml/min, split before entering MS 1:3 Injection volume: 2 μΐ MS conditions: ε 63 201127961 Ionization: Negative ion shower

Ion source conditions: ion spray voltage: 5kV temperature: 350 °C Fragmenter voltage and collision energy optimized sweep mode: selective reaction mode: m/z 129 83 offset used to determine AAP HPLC-MS analysis AAP The protonated molecules of AAP with m/z 176 were measured by special ion monitoring (SIM)-MS spectrometry. The concentration of AAP was calculated by measuring the area under the peak of the AAP spike. The sample was flushed at a lag time of 2.7 minutes. It is calibrated using an external standard process. All LC-MS experiments were performed on an Agilent 1100 LC system 'which consisted of a quaternary pump, deaerator, autosampler and column oven' coupled with a 三 2000 triple quadrupole MS (Applied Biosystems). The LC conditions are as follows: Column: Eluent: Flow rate: Gradient: 50*4 Nucleosil C18, 5 μιη (Macherey-Nagel) + 250 x 4.6 Prevail Cl8, 5 μπι (Alltech), both room temperature (RT) A = ultrapure water with 0.1% (Wv) formic acid B = acetonitrile with 0.1% (v/v) formic acid (pa, Merck) 1.2 ml/min, split 1:3 gradient before entering MS. At =0 minutes, this is 90% (v/v) A and changes to 50% (v/v) A in 6 minutes. At 6.1 minutes, the gradient was adjusted back to the original condition. Injection volume: 2 μΐ 64 201127961 MS conditions: Ionization using positive ion shower Detection: SIM mode, m/z 176, dwell time 1 〇〇 msec. HPLC-MS analysis for determining 6-ACA Calibration: The calibration was performed with an external calibration line of 6-ACA (m/z 132 + m/z 114 'Rt 7.5 minutes). All LC-MS experiments were performed on an Agilent 1100 consisting of a quaternary pump, a deaerator, an autosampler and column oven, and a single quadrupole MS (Agilent, Waldbronn, Germany). The LC-MS conditions are as follows: Column: 50*4 Nucleosil (Mancherey-Nagel) + 250 X 4.6 Prevail C18 (Alltech), both room temperature (RT) Equip: A = 0.1% (v/v ) Ultrapure water for formic acid B = acetonitrile (pa, Merck) Flow rate: 1.0 ml/min, split 1:3 before entering MS Gradient: Gradient starts at t=0 minutes 'this time is 1〇〇% ( v/v) a, maintained for 15 minutes, changed to 8〇% (v/v) b (t=30 minutes) within 15 minutes. The gradient was maintained at 8〇% (v/v) B from 30 to 31 minutes.

Injection volume: 5 μΐ MS detection: ESI(+)-MS Electrospray ionization (ESI) is operated in positive scan mode with the following conditions: 50-500, 50 V fragmenter voltage, completion m 々 Step size, 350 ° C dry gas temperature, 1 〇 L N2 / min 65 201127961 Dry gas, 50 psig atomizer pressure and 2,5 kV capillary voltage. 1.2 Selection of the target gene The design position of the construct is added to the ribosome binding position of all genes and the upstream gene position of the initiation codon and the position downstream of the stop codon to facilitate colonization, using Gateway technology (Invitrogen) , Carlsbad, CA, USA). Gene synthesis and plastid construction The synthetic gene line was obtained from DNA2.0 and codon-optimized for expression in E. coli 'according to the DNA 2.0 standard procedure. Vibrio fluvialis/Zwvia along the JS17 aminotransferase gene [SEQ ID NO: 1] and B. velutii KBAB4 aminotransferase gene [SEQ ID NO: 4], encoding the ω-aminotransferase of Vibrio fluvialis JS17, respectively [SEQ ID NO: 2], amino acid sequence of aminotransferase (SEQ ID NO: 5) of B. velutii (9) phantom? (KB) KB4, which is codon-optimized, resulting in sequence I; No. 3] and [SEQ ID NO: 6] were synthesized from DNA. From Escherichia coli [SEQ ID NO: 30], Saccharomyces Cerevisiae (Sacc/wromyce·? cereWWize) [SEQ ID NO: 33], Gymnosporium

[SEQ ID NO: 36], Lactobacillus reuteri (Laciococcws / aci (4) [SEQ ID NO: 39], [SEQ ID NO: 42], and Mycohacieri Mm iMfeercM / im··? [SEQ ID NO: 45] de-reducing enzyme The gene is a pyruvic acid encoding the diaminopimelate decarboxylase LysA [SEQ ID NO: 31], Saccharomyces cerevisiae Gacac/iaromKW 66 201127961, respectively, of Escherichia coli (five yc/ieWc/z/a co/ί) Destroy enzyme Pdc [SEQ ID NO: 34], Pyruvate dehydrogenase pdcI472A [SEQ ID NO: 37], and Lactobacillus leucocephalus (Laciococcwj / this edge) branch a-ketoacid dehydrogenase KdcA [sequence No. 40] with α-ketoisovalerate de-suppressing enzyme jqvD [SEQ ID NO: 43] ' and tuberculosis divergence iMi?ercM/&lt;9&gt;sz··?) ot-_glutaric acid decarboxylase Kgd The amino acid sequence of [SEQ ID NO: 46] is also codon-optimized, and the resulting sequence [SEQ ID NO: 32], [SEQ ID NO: 35], [SEQ ID NO: 38], [SEQ ID NO: 41], [SEQ ID NO: 44] And [SEQ ID NO: 47] were obtained from DNA synthesis. The gene construct was cloned into the pBAD/M:yc-His-DEST expression vector using 'Gateway technology (Invitrogen), via introduction to (10) tolerance position, pDONR201 (Invitrogen) as the entry vector, as described in the usage procedure ( Www.invitrogen.com). Using this method, the expression vectors pBAD-V//__AT and pBAD-5we_AT can be obtained separately. The corresponding expression strains were obtained by transforming the chemical competence to £ τοριο (invitr0gen) with the pBAD-expression vector. PCR selection of each gene encoding a biocatalyst was carried out by genomic DNA doubling by PCR technique using pcr supermix High Fidelity (Invitrogen) according to the instructions for use, using the primers in the table below. s 67 201127961 Table 2 Gene-derived gene sequence number Enzyme serial number primer sequence number Pseudomonas aeruginosa 7 8 9&amp;10 Pseudomonas aeruginosa 26 27 60&amp;61 Pseudomonas aeruginosa ( Pseudomonas aeruginosa) 66 67 72&amp;73 Pseudomonas aeruginosa 68 69 74&amp;75 Bacillus subtilis 14 15 48&amp;49 Bacillus subtilis 16 17 50&amp;51 Bacillus subtilis (Bacillus subtilis) 64 65 70&amp;71 Rhodobacter sphaeroides 18 19 52&amp; 53 Legionella pneumophila 20 21 54&amp;55 Nitrosomonas europaea 22 23 56&amp;57 Neisseria gonorrhoeae 24 25 58&59 Rhodopseudomonas palustris 28 29 62&63 PCR reaction analysis by agarose electrophoresis, PCR products of the correct size are washed by gel It was proposed to use QIAquick PCR purification kit (Qiagen, Hilden, Germany). The purified PCR product was selected to be thinned onto the pBAD/Myc-His-DEST expression vector, using Gateway technology (Invitrogen), by introducing the αίίβ position, and using pDONR-zeo (Invitrogen) as the entry vector, as in the instructions for use. Described. The gene sequence replicated by PCR was identified by DNA sequencing. Thus 68 201127961 to obtain the performance vector pBAD-/^e-_gi9946143_AT, pBAO-Bsu_gil6078032_AT &gt; pBAD-Bsu_gil6080075_AT &gt; pBAO-Bsu_gi 16077991_AT, pBAO-Rsp„AT, pBAD-jL/m_AT, pBAD-A^ew-AT, pBAD-A^o_AT, pBAD-corporate ae_gi9951299_AT, pBAD-Pae_gi9951072_AT, pBAD-corpse ae_gi9951630_AT and pBAD-/^a_AT. Corresponding performance strains can be transformed by pBAD constructs by chemically competent TOPIO cells (Invitrogen) The culture of E. coli to express small amounts of protein was carried out in a 96-deep well plate using 940 μM medium containing 0.02% (w/v) L-arabinose. Transfer the cells from the frozen stock. Inoculate into a 96-well plate (Kiihner, Birsfelden, Switzerland). Place the plate in a cyclotron shaker (300 rpm, 5 cm amplitude) and incubate at 25 ° C for 48 h. Typically 〇〇 62 () 11111 To 2-4. Preparation of cell lysates Preparation of lysis buffer The lysis buffer contains the following components: Table 3 1M MOPS pH 7.5 5 ml DNAse I Grade II (Roche) 10 mg Lysozyme 200 mg MgS04.7H20 123.2 mg Disulfide Threitol (DTT) 154.2 mg H2 〇 (MilliQ) Equilibrated to 100 ml The solution was prepared freshly before use. 69 201127961 A small amount of cultured cells were harvested by centrifugation by lysis to prepare cell-free extracts (see previous paragraph), and the supernatant was discarded. The cell pellet formed during centrifugation was frozen at -20 ° C for at least 16 h, then thawed on ice. Add 50 μl of freshly prepared lysis buffer to each well, and the cells were shaken vigorously with the plate 2-5 Suspension in minutes. In order to achieve lysis, the plate was placed at room temperature for 30 minutes. In order to remove the cell residue, the plate was centrifuged at 6000 g for 20 minutes at 4 ° C. The supernatant was transferred to a new plate. 'Save on ice until the next step. Prepare the cells in the cell-free extract by ultrasonic shock (see the previous paragraph) and harvest by centrifugation, discard the supernatant. Add 1 mi, PH7 phosphoric acid Potassium buffer to 0'5 g wet cell pellets' cells were suspended by strong shock. In order to achieve lysis, the cells were subjected to ultrasonic shock for 20 minutes. To remove cell debris, the plate was incubated at 4. (:, centrifuge at 6000 g for 20 minutes. The supernatant is transferred to a clean tube and cooled at -20 ° C; east, until the next use. U chemically hydrolyzes methyl S and methotiric acid. The preparation of the 5-A shiidy 4-aminotransferase reaction, ie, 5-mercaptovaleric acid, is prepared by chemically hydrolyzing the fluorenyl 5-mannyl pentadecyl ester by the following steps: 1〇 % (W/V) of sulfhydryl group 5, decyl valeric acid aqueous solution, adjusted to pH 14J with NaOH. After standing at 24 hours for 20 hours, the pH value was adjusted to 7.1,6 S-branched with HC1. Enzymatic reaction of conversion of valeric acid to 6_ACA Unless otherwise indicated, a preparation of 10 mM 5-mercaptoleic acid, 20 m]Vi, racemic &amp; methylbenzylamine, and 200 μM pyridaldehyde 5 was prepared. ,-phosphate 70 201127961 50 mM acid clock buffer, pH 7.0 mixture. 100 μΐ of the reaction mixture was dispensed into each well of the porous disk. To initiate the reaction, add 20 μM per well. Cell extract. The reaction mixture was incubated for 24 h at 37 ° C. In addition, the chemical blank mixture (without cell-free extract), and the biological blank group (with pBAD) /Myc-His C co/!· TOP10), cultured under the same conditions. The samples were analyzed by HPLC-MS. The results are summarized in the table below. Table 4: Conversion from 5-FVA to 6-ACA, in the amine transfer Biocatalyst 6-AC A concentration in the presence of enzyme [mg/kg] E. co/iTOP10/pBAD-V/?_AT 43* E. co/i TOP 10/pBAD-Pae_AT 930 E. coli TOPlO/pBAD-Pa^ AT 25* E. co/i TOP 10/pBAD-fiwe_AT 24* E. coli TOP10/pBAD-fijM_gil6077991_AT 288 E. coli TOP10/pBAD-Pae_gi9951072_AT 1087 E. coli TOP10/pBAD-Pae_gi9951630_AT 92 ^.^-pBAD/MK -His C^E. coli TOP10 (k blank group) 0.6 &quot;^ no (chemical blank group) ndnd: not detected * method differs in the use of 10 μΐ without cell extract, not 20 μΐ, The ratio of _5'_ phosphate is 50 μΜ instead of 200 μΜ, and the volume of the reaction mixture in each well is 190 μΐ instead of 100 μΐ. It is shown that 6-ACA can be formed from 5-FVA at the amine group. In the presence of a transferase, the enzyme reaction of 1.7 ΑΚΡ to be converted to 5 valeric acid consists of 50 mM hydrazine, 5 mM gasification, 100 μΜ ratio of multi-chain 5, _ sate (for LysA), Or 1 mM sulfur A reaction mixture of 100 mM potassium phosphate buffer, pH 6.5, of an amine dibasic acid salt (for all other enzymes). The reaction mixture of 4 ml 71 ε 201127961 was partitioned into a reaction tube. To initiate the reaction, 1 ml of cell-free extract prepared by ultrasonic shock was added to each well. For the commercially available grass-wound acetate de-slow enzyme (Sigma-Aldrich, Cat. No. 04878), 50 U was added. The reaction mixture was at 37. (:, stirring with magnetic stirrer for 48 h. In addition, chemical blank mixture (without cell extract), and biological blank group (with pBAD/AO^-His C5. TOP10) under the same conditions The samples collected from different time points in the reaction were analyzed by HPLC-MS. The results are summarized in the table below.

Table 5: 5-FVA biocatalyst formed by AKP in the presence of decarboxylase 5-FVA Concentration [mg/kg] 3h 18h 48h E. co/i TOP 10/pBAD-LysA 150 590 720 E. co/i TOPlO /pBAD-Pdc 1600 1700 1300 E. coli TOP10/pBAD-PdcI472A 2000 2000 1600 E. coli TOPlO/pBAD-KdcA 3300 2300 2200 E. co/iTOPlO/pBAD-KivD 820 1400 1500 Grasshopper acetic acid decarboxylase nd 6 10 pBAD/Myc-His C Ε·α&gt;ίί TOP10 (biological blank group) ndndnd none (chemical blank group) ndndndnd: No detection was found to be formed by AKP in the presence of decarboxylase. 1.8 long-term conversion to 6-long enzyme reaction, prepared in the presence of recombinant decarboxylase containing 50 mM AKP, 5 mM magnesium hydride, 100 μΜ. A reaction mixture of 100 mM potassium phosphate buffer, pH 6.5, than polyacid 5, _ phosphate (for LysA), or 1 mM thiamine bisphosphate (for all other enzymes). The reaction mixture of d ^ ml 72 201127961 is gi in the reaction tube. To initiate the reaction, 1 ml of cell-free extract prepared by ultrasonic shock was added to each well. The reaction mixture was stirred at 37 C with a magnetic stirrer 481^, a chemical blank mixture (without cell-free extract), and a biological blank group (with pBAD/MK_ms C5·c-ΤΟΡΙΟ), under the same conditions Under cultivation. Samples taken at different time points from the reaction were analyzed by HPLC-MS. The results are summarized in the table below.

Table 6: Formation of 6-ACA biocatalyst from AKP in the presence of decarboxylase] 6-AC A Concentration [mg/kg] 3h 18h 48h E. co/i TOPlO/pBAD-LysA na 0.01 0 E. coZi TOPI0/ pBAD-Pdc 0.1 0.3 na E. co/z TOPI0/pBAD-PdcI472A 0.03 0.1 0.2 E. co/ί TOP 10/pBAD-KdcA 0.04 0.1 0.3 E. co/i'TOPlO/pBAD-KivD na 0.3 0.6 Λ-pBAD /A/jc-His C^E. coli iopio (biological blank group) ndndnd no (chemical blank group) η η _ soil group B ndndndnd = no detected results show that in the presence of decarboxylase, 6_ACA can be formed by AKp . It should be considered that E. coli itself contains natural 5 _ F VA aminotransferase activity. 1.9 In the presence of recombinant decarboxylase and recombinant aminotransferase, Ακρ is converted to the heart of the sputum and shoulder preparation contains 50 mM AKP, 5 mM magnesium chloride, 1 〇〇 μΜ pyridoxal 5'-phosphate, 1 mM Thiamine diphosphate, 100 mM potassium phosphate buffer solution of 5 mM racemic alpha methylbenzylamine, reaction mixture of ρ Η 6 5 . i 6 73 % 201127961 ml The reaction mixture was dispensed into a reaction tube. To initiate the reaction, a cell-free extract containing 0.2 ml of decarboxylase and a cell-free extract containing 2 ml of aminotransferase were added to each reaction tube. The reaction mixture was stirred at 37 for 48 h with a magnetic stirrer. In addition, the chemical blank mixture (without cell-free extract and bio-blank group (with pBAD/Myc-His C:: co/Z TOP10) was cultured under the same conditions. Samples taken at different time points from the reaction were HPLC-MS analysis. The results are summarized in the table below. Table 7: Formation by AKp in the presence of recombinant decarboxylase and recombinant aminotransferase

6-ACA concentration of 6-ACA after 48 hours [mg/kg] E. co/i TOPIO/ pBAD-Vfl-AT E· coli TOPIO/ pBAD-Bwe-AT E. coli TOP 10/ pBAD-PAE gi9946143_AT E. co /iTOPl〇/pBAD-Pdc 183.4 248.9 117.9 E. coli TOP 10/ pBAD-PdcI472A 458.5 471.6 170.3 E. co/iTOPl〇/pBAD-KdcA 497.8 497.8 275.1 E. coli TOPIO/ pBAD-KivD 510.9 510.9 314.4 AT=Amine Transferase DC = decarboxylase did not detect 6-ACA in the chemical blank group and the biological blank group. Furthermore, the results show that the yield of conversion to 6-ACA is enhanced as compared to host cells having only recombinant decarboxylase (and no recombinant aminotransferase). 74 201127961 1.10 Construction of aminotransferase and devastatase plastids in S. cerevisiae The aminotransferase gene from Vibrio fluvialis (//wWa/) JS17, which encodes a river arc Bacterial (K/ZMvk/b) JS17 ω-aminotransferase amino acid sequence [SEQ ID NO: 2], PCR-multiplied, doubling from pBAD-Vfl_AT [SEQ ID NO: 3], using Phusion DNA polymerase (Finnzymes) ), according to the instructions for use, and use a specific primer [serial number 76 &amp; 77]. Pseudomonas aeruginosa (TieMiiomo/ia? aminotransferase gene [SEQ ID NO: 7], its aminotransferase of Pseudomonas aeruginosa [SEQ ID NO: 8], multiplied by pBAD using PCR technology -Pae_AT, using Phusion DNA polymerase (Finnzymes), according to the manual, and using a specific primer [SEQ ID NO: 78 &amp; 79]. The resulting PCR product was cloned onto the vector pAKP-41 using 5 &lt;el and mHI The restriction enzymes, which produce the vectors pAKP-79 and pAKP-80, respectively, thus contain the aminotransferase gene in the presence of the S. cerevisiae cerev/ike promoter and the S. cerevisiae terminator. Decarboxylase gene from S. cerevisiae cereWhae [SEQ ID NO: 33], the pyruvate decarboxylase Pdc [SEQ ID NO: 34] encoding Saccharomyces cerevisiae (Sctcc/mramyce·? cerevbke) was multiplied using PCR technology. pBAD-Pdc, using Phusion DNA polymerase (Finnzymes), according to the manual, and using specific primers [SEQ ID NO: 80 &amp; 81]. Dehydroenzyme gene from Z. aciococc Mi Zilciij [SEQ ID NO: 39] 'which encodes the Lactobacillus cerevisiae branch. α-keto acid decarboxylase KdcA [SEQ ID NO: 40], using PCR Technology was multiplied from 75 S—201127961 pBAD-KdcA using Phusion DNA Polymerase (Finnzymes) according to the instruction manual and using specific primers [SEQ ID NO: 82 &amp; 83]. The resulting PCR product was cloned into the vector pAKP-44, and the vector pAKP-81 and pAKP-82 were generated using AscI and a restriction enzyme, respectively, and thus the de-septase gene was contained in Saccharomyces cerevisiae (*S. cerevzWae) ga /2 promoter and Saccharomyces cerevisiae 〇 5. in the presence of a terminator. The plastids pAKP-79 and pAKP-80 were subjected to ~cl and ikl restriction enzyme digestion, and the plastids pAKP-81 and pAKP-82 were restriction enzymes; 0^1 was cut. An aminotransferase fragment, which is ligated to a low copy number gene-containing vector PRS414, such as a Ι/Ι^αΙ decarboxylase fragment, which has been cleaved by SacI restriction enzymes, S. cere WWae) . The resulting plasmid was as follows: pAKP-85: PgallO-Pae_AT-Tadh2 Pgal2-Pdc_DC-Tpmal pAKP-86: PgallO-Pae_AT-Tadh2 Pgal2-KdcA_DC-Tpmal pAKP-87: PgallO-Vfl_AT-Tadh2 Pgal2-Pdc_DC-Tpmal pAKP-88 : Pgall〇-Vfl_AT-Tadh2 Pgal2-KdcA-DC-Tpmal 1.11 Transformation of Saccharomyces Cerevisiae (s. cerevisiae) S. cerevisiae CEN.PK113-3C, transformed by 1 pg of plastid DNA, According to the method described by Gietz and Woods (Gietz, RD and Woods, RA (2002)). Yeast strains were transformed by Liac/SS vector DNA/PEG method (Methods in Enzymology 350: 87-96). The cells were seeded on a plastic dish containing lx Yeast Nitrogen Base, which was free of amino acids and 2% glucose. The resulting strain was grown under aerobic conditions, cultured at 3 ° C for 48 hours, and cultured in a Verduyn minimal medium containing 76% glucose and 4% galactose at 76 201127961. Preparation of Cell-Free Extract 1 ml of acid-filled clock buffer (pH 7) was added to 0.5 g of cell sputum. This mixture was added to a 2 ml eppendorf tube containing 0.5 g of glass beads having a diameter of 0.4-0.5 mM. The sample was violently shaken by the eppendorf shaker (IKA VIBRAX-VXR) for 20 s. The resulting cell-free extract was centrifuged at 14,000 rpm ' 4 ° C for 5 minutes. The supernatant was used in the enzyme activity assay. 1.13 AKP conversion to 6-ACA enzyme reaction, prepared in the presence of deoxylase and aminotransferase co-expressed by Saccharomyces cerevisiae (β. cerevisiae) containing 50 mM AKP, 5 mM magnesium sulphate, 1 〇〇 A reaction mixture of 100 mM potassium sulphate buffer pH 6.5 with 50 mM racemic α-mercapto benzylamine. 1.6 mi The reaction mixture was partitioned into a reaction tube. To initiate the reaction, 0-4 ml of Saccharomyces cerevisiae was added to each reaction tube containing no cell extract containing decarboxylase and aminotransferase. The reaction mixture was stirred at 37 ° C with a magnetic stir bar. In addition, chemical blank mixtures (without cell extracts) and biological blanks were cultured under the same conditions. The sample after 19 hours of cultivation was analyzed by HPLC-MS. The results are summarized in the table below. 77 201127961 Table 8: Formation of 6-ACA from AKP, using microbes as biocatalyst biocatalyst 6-ACA concentration [mg/kg] Letting Saccharomyces Cerevisiae ("S. cereWitVie" pAKP-85 63 Saccharomyces Cerevisiae (Fork 226 Saccharomyces Cerevisiae Bacteria (brothers cereWWae) ρΑΚΡ-87 1072 Saccharomyces cerevisiae ("S. cerevisiae" ρΑΚΡ-88 4783 Saccharomyces Cerevisiae (&amp; cerevidae) (biological blank group) 3.9 None (chemical blank group) 1.3 Ι·14 α-keto Glycan Preparation of 50 mM potassium phosphate containing 10 mM α-ketopimelic acid, 2 mM L-alanine, and 50 μM pyridoxal 5'-phosphate by an enzyme reaction of diacid to α-aminopimelic acid Buffer, reaction mixture at pH 7.0. 800 μΐ of the reaction mixture was dispensed into each well of a multiwell plate. To initiate the reaction, 200 μM of cell lysate was added to each well. The reaction mixture was shaken at 37 ° C. In the device for 24 h. In addition, the chemical blank mixture (without cell-free extract), and the biological blank group (with pBAD/Myc-His C:: TOP10) were placed under the same conditions. The sample was analyzed by HPLC-MS. The results are summarized in the table below. 78 201127961 Table 9: Aminotransfer AAP biocatalyst AAP concentration [mg/kg] formed by AKP in the presence of enzyme (after 24 hours) E. co//TOP10/pBAD-V/Z_AT 3.7 E. coli TOPlO/pBAD-Psy_KT 15.8 E. co/i TOPIO /pBAD-β^Μ_gil6078032_AT 11.2 E. coZt TOPI0/ρΒΑϋ-/?5·ρ_ΑΤ 9.8 E. coli TOPl0/pBAO-Bsu_gil6080075_KT 4.6 E. coZi TOPI0/pBAD-Lpn_AT 5.4 E. c〇m〇P10/pBAO-Neu_KT 7.7 E. coli ΎΟΡΙΟ/ρΒAD-Ngo_KT 5.1 E. coli TOP10/pBAD-Pae_gi9951299_AT 5.6 E. coliTOPlO/pBAO-Rpa^AT 5.4 with pBAD/M; yc-His C £. TOPIO (bio blank group) 1.4 None (chemistry Blank group) 0 The results show that AKP can be catalyzed by biocatalyst to form AAP. II. Examples of preparation of CX-keto acid Π.1 General methods Molecular and genetic technology Standard genes and molecular organisms are generally known in the art, and It has been previously described (Maniatis et al. 1982 "Molecular cloning: a laboratory manual". Cold Spring Harbor Laboratory, Cold Spring Harbor, NY; Miller 1972 "Experiments in molecular genetics", Cold Spring Harbor Laboratory, Cold Spring Harbor; Sambrook and Russell 2001 "Molecular cloning: a laboratory manual" (3rd edition), Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press; F. Ausubel et al, eds·, "Current £ 79 201127961 protocols in molecular biology,, 5 Green Publishing and Wiley Interscience , New York 1987). Plastids and plants pMS470 (Balzer, D.; Ziegelin, G.; Pansegrau, W.; Kruft, V.; Lanka, E. Nucleic Acids Research 1992, 20(8), 1851-1858.) and pBBRIMCS (Kovach ME, Phillips RW, Elzer PH, Roop RM 2nd, Peterson KM. Biotechniques. 1994 May; 16(5): 800-2. pBBRIMCS: a broad-host-range cloning vector) has been previously described. E. coli strains TOPIO and DH10B (Invitrogen, Carlsbad, CA, USA) were used in all selection procedures. E. coli strains BL21 A1 (Invitrogen, Carlsbad, CA, USA) and BL21 (Novagen (EMD/Merck), Nottingham, UK) were used for protein expression. Culture medium; 2xTY culture solution (16 g/1 tryptopeptone '10 g/1 yeast extract, 5 g/1 NaCl) was used to culture E. coli. Antibiotics (loo pg/mi ampicillin, 50-100 pg/ml neomycin) were added to maintain the plastids in E. coli. In order to induce the expression of the gene in E. coli, arabinose (for BL21-AI-derived strains) and IPTG (for pMS470, PBBR1MCS-derived strains) were used, with a final concentration of 0.02% (L-arabinose) and 0.2. mM (IPTG). AKP manufactured by E. coli is cultured in M9 minimal medium (12.8 g/L Na2HP04.7H20, 3 g/L KH2P〇4, 0.5 g/L NaCl, 1 g/L NH4C1, 2 mM MgS04, 0.1 mM CaCl2) 'Added 1% glucose. Plastid identification 80 201127961 Gene, biochemical and/or phenotypic rhythm known to the technical system of different genes, such as the resistance to antibiotics, PCR5 to the analysis of the stalks Purification of bulk DNa, restriction enzyme analysis of purified plastids, or DNA paste analysis. The integrity of all new constructs is confirmed by restriction enzyme cleavage, and if a PCR step is involved, additional sequencing confirmation is performed. UPLC-MS/MS analysis for determining α-keto acid

Waters HSS Τ3 column, 1&gt;8, 1 〇〇*2a, is used to prepare α-keto acid, and the elution gradient is shown in Table (1). The extract A consisted of 0.1% citric acid, and the extract B consisted of 〇1% formic acid. The flow rate was 0.25 ml/min and the column was 4 Torr. (: Thermal equilibrium state. 5 in 0: Ϊ separation &amp; keto acid, 6 fu, 5_FVA and the same (4) citrate time (minutes) 0 5.0 5.5 10 10.5 15 % A 100 85 20 20 100 100 % B 0 15 80 Wide 80 0 0

The Waters micromass Quattro micro API is used in the electrospraying step, whether in positive or negative ionization mode, depending on the compound to be analyzed, using multiple reaction monitoring (MRM). The ion source temperature was maintained at 13 Torr. Helium, wherein the solvent desorption temperature was 350 ° C and the flow rate was 500 L / hr. In the case of AKP, deprotonated molecules are fragmented by 1〇_14eV# to produce specific fragments such as H2〇, CO and co2. To determine the concentration, a standard curve for the synthetic preparation of the compounds is established to calculate the reaction factors for each ion. This is used to calculate the concentration of an unknown sample. £ 81 201127961 //.2 Synthesis of 2-hydroxypimelic acid (a method for illustrating how AKP is used for testing purposes) 2-hydroxypimelic acid used as a substrate for the biocatalytic reaction of ruthenium Synthesis by hydrogenation (from Syncom). AKP (2.2 g, 12.6 mmol) was dissolved in methanol (50 mL), and 3 〇mg of 卩〇1 (?(1/(:, 5%) attached to the coke and sputum was added and placed in an autoclave </ RTI> </ RTI> </ RTI> <RTI ID=0.0></RTI> </RTI> <RTIgt; The product was identified by ^-NMR and 13C-NMR as W-NMR (300 MHz, DMSO): δ 4.02-3.98 and 3.92-3.89 (dd, V = 7.6 Hz, V = 4.8 Hz, 1 Η), 2.28 and 2.18 ( , 3J = 7.2 Ηζ, 2H), 1.66-1.28 (m, 6 H) 13C-NMR (75 MHz, DMSO): δ 174.9, 173.6, 70.0, 51.6, 34.0, 33.6, 24.6 U.3 Preparation with heterogeneous The hydroxy-acid oxidase of ρΒΑΌ-DEST ToplO cells HAOX5B and LAOX8C are derived from DNA synthesis. The positional line is added to the ribosome binding position of all genes and upstream of the start codon, and downstream of the stop codon to help select Colonization, using Gateway technology (Invitrogen, Carlsbad, CA, USA). The genetic construct was cloned onto the pBAD/MK-His-DEST expression vector using Gateway technology (Invitrogen) via introduction to the αίίβ position, and Use pDONR201 (Invitrogen) as the entry vector, as described in the usage procedure (\^¥评.丨11¥如(^611.00111). Using this method, the expression vectors PBAD-HAOX5B and pBAD-LAOX8C can be obtained respectively. The expression strains were obtained by pBAD-expression vector, which was chemically competent to transform Wu.cw 82 201127961 TOPIO (Invitrogen). U.4 cultured E. coli to express protein as small growth of cells prepared in Example II.3, at 96 - Perform in a deep well plate using 940 μΐ medium containing 0.02% (w/v) L-arabinose. Transfer the cells from the frozen stock to a 96-well plate (Kiihner, Birsfelden, Switzerland) for inoculation. The culture plate was placed in a cyclotron shaker (300 rpm, 5 cm amplitude) and incubated at 25 ° C for 48 h. Generally, 〇D62Gnm to 2-4 was obtained. Preparation of U5 cell lysate The lysis buffer contains the following components: Table 11 Lysis Buffer Composition 1M MOPS pH 7.5 5 ml DNAse I Grade II (Roche) 10 mg Lysozyme 200 mg MgS04.7H20 123.2 mg Dithiothreitol (DTT) 154.2 mg H20 (MilliQ) 1 Balance to 100 ml Solution in use Freshly prepared before. Small amounts of cultured cells were harvested by centrifugation (see Example 2) and the supernatant was discarded. The cell pellet formed during centrifugation was chilled at -2 ° C for at least 16 h and then thawed on ice. 500 μl of freshly prepared lysis buffer was added to each well and the cells were suspended in a vigorous shaking plate for 2-5 minutes. In order to achieve lysis, the plate was left to stand at room temperature for 3 minutes. To remove cell debris, the plates were centrifuged at 6000 g for 20 minutes at 4 °C. The supernatant (containing %', acid oxidase, whether HA0X 5B or LA0X 8C) was transferred to the new: medium: Store on ice until the next step. , 1 83 201127961 JJ. 6 Preparation of ΑΚΡ 2-hydroxypimelic acid (final concentration 5 mM, &gt; 95% purity, prepared as described above) with an enzyme is the hydroxy acid oxidase prepared as described above 3 (regardless of It is contacted with haOX 5B or LAOX8C) in a buffer solution containing the following ingredients. 4-amino-amino-n-peptin (4_aminoanj; ipyrine) (1 m]yi) -3'5-dioxa-2-hydroxybenzenesulfonic acid (DCHBS) (10 mM) -50 mM potassium phosphate buffer, pH 7.5. - Horseradish peroxidase (200 U/ml) The reaction was allowed to stand at 37 ° C for 20 h. The sample was cooled; East and heated to 95 ° C for 2 minutes prior to analysis to precipitate the protein. After centrifugation, the supernatant was analyzed by UPLC-MS. The AKP concentration measured in the HAOX 5B sample was 59 mg/b and the AKP concentration measured in the LAOX 8C sample was 58 mgA.

Example 111 Preparation of 5-AVA IU. 1 gene in Escherichia coli for Z. jejuni (Mei/iimococcM·? aconitase (high aconitase) small subunit (AksE, MJ1271, [serial number 117]), high aconitase major unit (AksD, MJ1003, [SEQ ID NO: 114]), and high isocitrate dehydrogenase (AksF, MJ1596, [SEQ ID NO: 120]), from A. caryobacter (Mei/ianococcws SB high homogenate-like acid dehydrogenase (AksF, Mevan_0040, [SEQ ID NO: 123) homologue, from Giobococcus (Mei/ianococc Mi1 Nankai 3 high aconitase small subunit (AksE, Maeo_0652, [SEQ ID NO: 135]), aconitase large subunit (AksD, Maeo_0311, [SEQ ID NO: 138] homologue, from M. sphaeroides (Mei/iimococcwi·, Gaowutou 84 201127961, small acidase) Subunit (AksE, MMP0381 '[SEQ ID NO: 129]), homolog of the high aconitase major unit (AksD, MMP1480, [SEQ ID NO: 132]), high isocitrate dehydrogenase (AksF, MMP0880 [sequence] No. 126),]), brown nitrogen-fixing bacteria v-you) high citrate synthase (Nifv, [serial number 141]) 'from Vibrio fluvialis JS1? Aminotransferase protein [SEQ ID NO: 2], and the protein sequence of the branched alpha-keto acid decarboxylase KdcA [SEQ ID NO: 40] from L. reuteri, can be retrieved from the database. All genes except for brown nitrogen-fixing bacteria (A. Wne/imi/h·) high citrate synthase nifV (SEQ ID NO: 140), all of which are codon pairs optimized in E. coli, as described in WO 2008/000632 (Table 13) In the described method, the construct is prepared synthetically (Geneart, Regensburg, Germany). In the optimization process, 'avoiding the restriction enzyme cleavage position, the shared restriction enzyme cleavage position is added to the start and stop positions' Colonization onto the expression vector. Codon-optimized aminotransferase gene (SEQ ID NO: 3) from Vibrio fluvialis (ν^η·ο /Zmvk/) JS17, PCR-multiplied, polymerized using Phusion DNA Enzyme, according to the manual, use primer pair AT-Vfl_for_Ec (AAATTT GGTACC GCTAGGAGGAATTAACCATG) + AT-Vfl_rev_Ec (AAATTT ACTAGT AAGCTGGGTTTACGCGACTTC). Codon-optimized de-reducing enzyme from Lactobacillus

The gene encodes the L-lactobacillus branched α-decanoate decarboxylase KdcA (SEQ ID NO: 41), which is multiplied using Phusion DNA polymerase. According to the manual, the primer Kdc_for_Ec (AAATTT ACTAGT GGCTAGGAGGAATTACATATG) and Kdc_rev_Ec (AAATTT e-7) are used. · 85 201127961 AAGCTT ATTACTTGTTCTGCTCCGCAAAC). The aminotransferase fragment was cleaved by KpnI/Spel and the decarboxylase fragment was cleaved by Spel/Hindlll. Both fragments were ligated with ppnBR-lac cleaved with Kpnl/Hindlll to obtain pAKP-96. Small aconitase from J. japonicus (M. 々mncLsc/n'i), M. sphaeroides (de-stained, E. aeruginosa (M. marxianus marba/wA·5) The subunit (AksE), the high aconitase major unit (AksD) and the gene of the high isocitrate dehydrogenase (AksF) were optimized by E. coli codons (using W008000632; Table 13). The sub-pair optimized gene sequences are SEQ ID NO: 115, 118, m, 124, 127, 130, 133, 136 and 139. The construct is synthetically prepared (Geneart, Regensburg, Germany) containing the wild-type nifv gene. The optimized gene (SEQ ID NO: 140), also see Table 12, listing the genes for each construct. In the optimization process, 'avoiding the restriction enzyme cutting position, the shared restriction enzyme cutting position is added. The start and stop positions are selected for expression on the expression vector. In addition, the tac promoter sequence upstream of the AksD sequence from pMS470 is added. Each ORF is preceded by a consensus ribosome binding site, and a leader sequence to drive transcription of E. coli Translation. The synthetic AksA/AksF cassette sequence is via Ndel/ Xbal cut, the synthetic AksD/AksE cassette sequence was cut by Xbal/Hindlll. The fragment containing Aks gene was inserted into the Ndel/Hindlll position of pMS470 to obtain each vector. These quality systems and plastid pAKP96, one from the river The bacterium (V7Z?n'o / Ζϋνί·α/), the carrier containing the aminotransfer Sf: gene (AT) and the de-slowing enzyme gene (DC) from lactic acid bacteria/acibj, together transform BL21 The strains listed in Table 12, 201127961 are listed in Table 12. Table 12 Each gene sequence strain present in eAKP236, eAKP-489 and eAKP-491 provides the organism NifV Seq ID AksD Seq ID AksE Seq ID AksF Seq ID AT DC eAKP236 M.jarwmchii &amp; Azotobacter vinelandii (NifV) &amp; y. fluvialis ί L. lactis 140 115 118 121 3 41 eAKP489 Markov M.maripaludis &amp; M. mnnielii &amp; Azotobacter vinelandii (NijV) &amp; Vibrio fluvialis &amp; Ritter Lactobacillus (L. lactis) 140 133 130 124 3 41 eAKP491 Espresso (M, aeolicus) Nankai 3 &amp; M.maripaludis &amp; Azotobacter vinelandii (NifV) &amp; V. fluvialis ί雷特 k lactic acid bacteria (L Lactis} 140 139 136 127 3 41 (ΑΤ=aminotransferase, DC=decarboxylase) U1.2 Protein expression and metabolite production in E. coli eAKP236, eAKP-489 and eAKP-491 (see Table 12) The culture was incubated in a test tube and cultured overnight in 10 ml of 2*TY medium. The 200 μl culture was transferred to an Erlenmeyer flask containing 20 ml of 2 χΤΥ medium. The Erlenmeyer flask was placed on a 30 C, 280 rpm orbital shaker. After 4 h, IPTG was added at a final concentration of O.lmM, and the Erlenmeyer flask was placed at 3 Torr. (:, l6h at 12 rpm. The cells in 20 ml culture were collected by centrifugation, suspended in 4 miM9 medium, containing 1% glycerol, and cultured in a 24-well plate at 30 ° C, 210 rpm. After 24 hours, the supernatant was collected by centrifugation and stored in _2 , until analysis. UL3-detected 5-AVA in the culture sample was used to determine the HPLC-MS analysis of 5-AVA using the external 6-AVA Calibration line (m/z 116, hysteresis time 4.1 minutes) was calibrated. All LC-MS experiments were performed on an Agilent 1200 LC system with a quaternary pump, deaerator, autosampler, column oven, and

S 87 201127961 Triple-quadrupole 6410 LC-MS (Agilent, Waldbronn, Germany). The LC-MS conditions were as follows: Column: 50*4 mm Prevail Cl8 (Alltech) + 250 * 4.6 mm Prevail C18 (Alltech), both room temperature (RT) Eluent: A = 0.1% (v/ v) Ultrapure water of citric acid B. Acetonitrile with 0.1% (v/v) formic acid (Lichrosolv, Merck) Flow rate: 1.2 ml/min, split 1:3 before entering MS Gradient: Gradient starts at t= At 0 minutes, this time is 1〇〇% (v/v)A, which changes to 15% (v/v)B in 8 minutes, and changes to 50% (v/v) B from 8-9 minutes. The gradient was maintained constant at 50% (v/v) B from 9 to 12 minutes.

Injection volume: 5 μΐ MS detection: ESI(+)-MS Electrospray ionization (ESI) was performed in negative ion SIM mode using the following conditions: MS2 SIM at m/z 116, residence time 100 msec, 70 V Fragmenter voltage," (^ dry gas fox' I2 L IVmin dry gas, 5 psig atomizer pressure, and 3 kV capillary voltage. Table 13 Results Strain C-Source Resting Time - Culture Medium Culture Conditions 5-AVA [mM/1] BL-21 Glycerin 24h M9 24 ΜΤΡ nd eAKP-236 Glycerin 24h M9 24 ΜΤΡ 0,18 eAKP-489 Glycerin 24h M9 ~~~—— 24 hole ΜΤΡ 0,11 eAKP-491 Glycerin 24h ~ M9 —--- — 24-well ΜΤΡ 0,13 ----- 88 201127961 The results clearly show that 5-AVA is present in the recombinant strain, but not observed on the untransformed BL-21 strain. IV 5-FVA aminotransferase activity for shorter receptor analogs W.1 Preparation 4-methylmercaptobutyrate 4-mercaptobutyric acid (4-FBA) system thiol 4-methionine The acid ester is prepared by chemical hydrolysis (Syncom, Groningen, The Netherlands). The preparation method is as follows. 10% (w/v) aqueous solution of methyl 4-mercaptobutyrate, NaOH Adjusted to pH 14.1. After standing at 2 ° C for 24 hours, the pH was adjusted to 7.1 with HC1. The thus obtained 4-FBA solution was appropriately diluted and used. IV.2 S-FVA-ATs for shorter Activity of the Amino Analogs The cell-free extract (CFE) containing the aminotransferase was prepared as above and tested for its activity on 4-FBA and 3-methylmercaptopropionic acid (3-FPA), respectively, in spectroscopy. In the test, 4-FBA was prepared as above, and 3-FPA was obtained from Sigma-Aldrich (Schnelldorf, Germany). The activity test was based on α-mercaptobenzylamine (MBA) as an amine group and detected by-product benzene. Ethyl ketone on the Perkin Elmer Lambda35 UV/VIS spectrometer 'thermal equilibrium at 30 ° C' wavelength of 300 nm. The final reaction volume is 1 m containing 50 μM of appropriately diluted CFE, which contains the aminotransferase, which is discarded. Plastic UV tube, mixed with 25mM MBA and 10mM 5-FVA, 4-FBA or 3-FPA, in 50 mM potassium phosphate buffer, pH 7.5, containing 0.1 mM pyridoxal 5'-phosphate (PLP) . The reaction was initiated by the addition of 1 μM of formic acid to other test components, and the other components were pre-placed in a spectrometer at 30 ° C for 5 minutes. After adding formic acid, the absorbance at 300 nm was recorded, and the aminotransferase activity in the CFE sample was calculated after three measurements, 89 201127961 according to Lambert-Beer's law. The molar concentration of acetophenone extinction coefficient ε = 0.28

Cm2^mol. One unit (U) of the aminotransferase activity is defined as from 25 mM

1 Pmol of acetophenone was formed in MBA with 10 mM formic acid and contained '0.1 mM PLP' per minute in 50 mM potassium phosphate buffer pH 7.5 at 30 °C. Aminotransferase V//_AT (SEQ ID NO: 2), Pae-AT (SEQ ID NO: 8), 5we_AT (SEQ ID NO: 5), and P-like-AT_gi9951072 (SEQ ID NO: 67) volume activity in U per ml of CFE 'Listed in Table 14. Table 14: Volumetric activity of CFE containing 5-FVA aminotransferases Vfl_AT, Pae_AT, Bwe_AT and Pae_AT_gi9951072, respectively containing 1 mM mM 5-mercapto valeric acid (5-FVA), 4-mercaptobutyric acid (4-FBA), or 3-mercaptopropionic acid (3-FPA). Biocatalyst activity 5-FVA [U/ml] Activity 4-FBA [U/ml] Activity 3-FPA [U/ml] Vfl_AT 59.5 33.4 144 Pae-AT: 18.2 15.7 48.1 Bwe_KT 0.42 0.38 0.95 Pae_AT_gi9951072 4.94 2.59 4.62 These results Showing aminotransferases V/IAT (SEQ ID NO: 2), i^e_AT (SEQ ID NO: 8), fiwe_AT (SEQ ID NO: 5), and ~β-AT-gi9951072 (SEQ ID NO: 67), also applicable 4-曱Mercaptoic acid (4-FBA) aminotransferase and 3-mercaptopropionic acid (3-FPA) aminotransferase. Example V AK? -DCs for shorter substrates

Cell-free extracts containing AKP-decarboxylase KdcA (SEQ ID NO: 40), KivD (SEQ ID NO: 43) and PDC I472A (SEQ ID NO: 37) were freshly prepared as above and tested for AKP and alpha-ketoadipate, respectively. (AKA) to de-rebase. The total reaction volume was 2.5 ml, and the PDC 90 201127961 variant 1472 (undiluted), KivD (undiluted) and KdcA (1:10 dilution) containing 5% CFE were added to the total reaction volume, respectively, with 25 mM AKP. Or AKA reaction, in 100 mM potassium phosphate buffer, pH 6.5, containing 1 mM thiamine diphosphate and 5 mM MgCl2 at 37 ° C and 400 rpm. After 5 minutes of reaction, 0.25 ml of the sample was taken and the reaction was terminated by the addition of 〇·75 ml of a 1:1 acetonitrile/water mixture and centrifuged (20 min, 5000 x g). The sample was analyzed by HPLC-MS using decarboxylation of a general AKP to 5-nonylvaleric acid (5-FVA) and decarboxylation of AKA to 4-mercaptobutyric acid (4-FBA). As a result, it was found that some 5-FVA or 4-FVA have been converted to adipic acid or glutaric acid by the enzyme in the cell-free extract. Therefore, the activity of AKP decarboxylase and AKA decarboxylase is calculated by summing the concentration of 5-FVA plus adipic acid formed and the concentration of 4-FBA plus glutaric acid. One unit of AKP decarboxylase activity is defined as the sum of 5-FVA and adipic acid in μπιοί 'formed from 25 mM AKP per minute at 37 ° C in 100 mM potassium phosphate buffer, pH 6.5, containing 1 mM Thiamine diphosphate in the presence of 5 mM MgCl2. One unit of AKA decarboxylase activity is defined as the sum of 4-FBA and glutaric acid, in units of μιηοΐ, formed from 25 mM AKA per minute, at 37 ° C 'in 100 mM potassium acetate buffer, pH 6.5 Containing 1 mM thiamine diphosphate in the presence of 5 mM MgCl2. The results are shown in Table 15. 91 201127961 Table 15: Activity of AKP deficient enzymes PDC I472A, KdcA and KivD for pimelic acid (ΑΚΡ) and α-ketoadipate (AKA), with a concentration of 50 mM long-term biocatalyst for ΑΚΡ activity [U /ml] Activity for AKA [u/ml] 卩0匸 Variant 1472-8 0.06 0.46 KdcA 23.4 8.07 KivD 0.18 0.73 These results show decarboxylase KdcA (SEQ ID NO: 4), KivD (SEQ ID NO: 43) and PDC I472A (SEQ ID NO: 37), also a suitable alpha-ketoadipate decarboxylase. [Simple diagram:! (none) [Explanation of main component symbols] (none) 92 201127961 Sequence Listing &lt;210&gt; 1 &lt;211&gt; 1362 &lt;212&gt; DNA &lt;213&gt; Vibrio fluvialis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1)..(1362) &lt;400&gt; 1 atg aac aaa ccg caa age tgg gaa gee egg gee gag acc tat teg etc Met Asn Lys Pro Gin Ser Trp Glu Ala Arg Ala Glu Thr Tyr Ser Leu 15 10 15 tat ggt ttc acc gac atg cct teg ctg cat cag ege ggc aeg gtc gtc Tyr Gly Phe Thr Asp Met Pro Ser Leu His Gin Arg Gly Thr Val Val 20 25 30 gtg acc cat ggc gag gga ccc tat ate gtc gat gtg Aat ggc egg cgt Val Thr His Gly Glu Gly Pro Tyr lie Val Asp Val Asn Gly Arg Arg 35 40 45 tat ctg gac gee aac teg ggc ctg tgg aac atg gtc geg ggc ttt gac Tyr Leu Asp Ala Asn Ser Gly Leu Trp Asn Met Val Ala Gly Phe Asp 50 55 60 cac aag ggg ctg ate gac gee gee aag gee caa tac gag cgt ttt ccc His Lys Gly Leu lie Asp Ala Ala Lys Ala Gin Tyr Glu Arg Phe Pro 65 70 75 80 ggt tat cac gee ttt ttc Ggc ege atg tee gat cag aeg gta atg ctg Gly Tyr His Ala Phe Phe Gly Arg Met Ser Asp Gin Thr Val Met Leu 85 90 95 teg gaa aag ctg gtc gag gtg teg ccc ttt gat teg ggc egg gtg ttc Ser Glu Lys Leu Val Glu Val Ser Pro Phe Asp Ser Gly Arg Val Phe 100 105 110 tat Aca aac teg ggg tee gag geg aat gac acc atg gtc aag atg eta Tyr Thr Asn Ser Gly Ser Glu Ala Asn Asp Thr Met Val Lys Met Leu 115 120 125 tgg ttc ctg cat gca gee gag ggc aaa ccg caa aag ege aag ate ctg Trp Phe Leu His Ala Ala Glu Gly Lys Pro Gin Lys Arg Lys lie Leu 130 135 140 acc ege tgg aac gee tat cac ggc gtg acc gee gtt teg gee age atg Thr Arg Trp Asn Ala Tyr His Gly Val Thr Ala Val Ser Ala Ser Met 145 150 155 160 acc ggc aag ccc tat aat teg gtc ttt ggc ctg ccg ctg ccg ggc ttt Thr Gly Lys Pro Tyr Asn Ser Val Phe Gly Leu Pro Leu Pro Gly Phe 165 170 175 gtg cat ctg acc tgc ccg cat tac tgg ege Tat ggc gaa gag ggc gaa 48 96 144 192 240 288 336 384 432 480 528 576 1 201127961

Val His Leu Thr Cys Pro His Tyr Trp Arg Tyr Gly Glu Glu Gly Glu 180 185 190 acc gaa gag cag ttc gtc gcc cgc etc gee ege gag ctg gag gaa aeg 624

Thr Glu Glu Gin Phe Val Ala Arg Leu Ala Arg Glu Leu Glu Glu Thr 195 200 205 ate cag cgc gag ggc gcc gac acc ate gcc ggt ttc ttt gcc gaa ccg 672 lie Gin Arg Glu Gly Ala Asp Thr lie Ala Gly Phe Phe Ala Glu Pro 210 2X5 220 gtg atg ggc geg ggc ggc gtg att ccc ccg gcc aag ggc tat ttc cag 720

Val Met Gly Ala Gly Gly Val lie Pro Pro Ala Lys Gly Tyr Phe Gin 225 230 235 240 geg ate ctg cca ate ctg cgc aaa tat gac ate ccg gtc ate teg gac 768

Ala lie Leu Pro lie Leu Arg Lys Tyr Asp lie Pro Val lie Ser Asp 245 250 255 gag gtg ate tgc ggt ttc gga cgc acc ggt aac acc tgg ggc tgc gtg 816

Glu Val lie Cys Gly Phe Gly Arg Thr Gly Asn Thr Trp Gly Cys Val 260 265 270 acc tat gac ttt aca ccc gat gca ate ate teg tee aag aat ett aca 864

Thr Tyr Asp Phe Thr Pro Asp Ala lie lie Ser Ser Lys Asn Leu Thr 275 280 285 geg ggc ttt ttc ccc atg ggg geg gtg ate ett ggc ccg gaa ett tee 912

Ala Gly Phe Phe Pro Met Gly Ala Val lie Leu Gly Pro Glu Leu Ser 290 295 300 aaa egg ctg gaa acc gca ate gag geg ate gag gaa ttc ccc cat ggc 960

Lys Arg Leu Glu Thr Ala lie Glu Ala lie Glu Glu Phe Pro His Gly 305 310 315 320 ttt acc gcc teg ggc cat ccg gtc ggc tgt get att geg ctg aaa gca 1008

Phe Thr Ala Ser Gly His Pro Val Gly Cys Ala lie Ala Leu Lys Ala 325 330 335 ate gac gtg gtg atg aat gaa ggg ctg get gag aac gtc cgc cgc ett 1056 lie Asp Val Val Met Asn Glu Gly Leu Ala Glu Asn Val Arg Arg Leu 340 345 350 gcc ccc cgt ttc gag gaa agg ctg aaa cat ate gcc gag cgc ccg aac 1104

Ala Pro Arg Phe Glu Glu Arg Leu Lys His lie Ala Glu Arg Pro Asn 355 360 365 ate ggt gaa tat cgc ggc ate ggc ttc atg tgg geg ctg gag get gtc 1152 lie Gly Glu Tyr Arg Gly lie Gly Phe Met Trp Ala Leu Glu Ala Val 370 375 380 aag gac aag gca age aag aeg ccg ttc gac ggc aac ctg teg gtc age 1200

Lys Asp Lys Ala Ser Lys Thr Pro Phe Asp Gly Asn Leu Ser Val Ser 385 390 395 400 gag cgt ate gcc aat acc tgc acc gat ctg ggg ctg att tgc egg ccg 1248

Glu Arg lie Ala Asn Thr Cys Thr Asp Leu Gly Leu He Cys Arg Pro 405 410 415 2 201127961 ctt ggt cag tcc gtc gtc ctt tgt ccg ccc ttt ate ctg acc gag geg 1296

Leu Gly Gin Ser Val Val Leu Cys Pro Pro Phe lie Leu Thr Glu Ala 420 425 430 cag atg gat gag atg ttc gat aaa etc gaa aaa gcc ctt gat aag gtc 1344

Gin Met Asp Glu Met Phe Asp Lys Leu Glu Lys Ala Leu Asp Lys Val 435 440 445 ttt gcc gag gtt gcc tga 1362

Phe Ala Glu Val Ala 450 &lt;210&gt; 2 &lt;211&gt; 453

&lt;212&gt; PRT &lt;213&gt; Vibrio fluvialis &lt;400&gt; 2

Met Asn Lys Pro Gin Ser Trp Glu Ala Arg Ala Glu Thr Tyr Ser Leu 15 10 15

Tyr Gly Phe Thr Asp Met Pro Ser Leu His Gin Arg Gly Thr Val Val 20 25 30

Val Thr His Gly Glu Gly Pro Tyr lie Val Asp Val Asn Gly Arg Arg 35 40 45

Tyr Leu Asp Ala Asn Ser Gly Leu Trp Asn Met Val Ala Gly Phe Asp 50 55 60

His Lys Gly Leu lie Asp Ala Ala Lys Ala Gin Tyr Glu Arg Phe Pro 65 70 75 80

Gly Tyr His Ala Phe Phe Gly Arg Met Ser Asp Gin Thr Val Met Leu 85 90 95

Ser Glu Lys Leu Val Glu Val Ser Pro Phe Asp Ser Gly Arg Val Phe 100 105 110

Tyr Thr Asn Ser Gly Ser Glu Ala Asn Asp Thr Met Val Lys Met Leu 115 120 125

Trp Phe Leu His Ala Ala Glu Gly Lys Pro Gin Lys Arg Lys Le Leu 130 135 140

Thr Arg Trp Asn Ala Tyr His Gly Val Thr Ala Val Ser Ala Ser Met 145 150 155 160 3 s: 201127961

Thr Gly Lys Pro Tyr Asn Ser Val Phe Gly Leu Pro Leu Pro Gly Phe 165 170 175

Val His Leu Thr Cys Pro His Tyr Trp Arg Tyr Gly Glu Glu Gly Glu 180 185 190

Thr Glu Glu Gin Phe Val Ala Arg Leu Ala Arg Glu Leu Glu Glu Thr 195 200 205 lie Gin Arg Glu Gly Ala Asp Thr lie Ala Gly Phe Phe Ala Glu Pro 210 215 220

Val Met Gly Ala Gly Gly Val lie Pro Pro Ala Lys Gly Tyr Phe Gin 225 230 235 240

Ala lie Leu Pro lie Leu Arg Lys Tyr Asp lie Pro Val lie Ser Asp 245 250 255

Glu Val lie Cys Gly Phe Gly Arg Thr Gly Asn Thr Trp Gly Cys Val 260 265 270

Thr Tyr Asp Phe Thr Pro Asp Ala lie He Ser Ser Lys Asn Leu Thr 275 280 285

Ala Gly Phe Phe Pro Met Gly Ala Val lie Leu Gly Pro Glu Leu Ser 290 295 300

Lys Arg Leu Glu Thr Ala lie Glu Ala He Glu Glu Phe Pro His Gly 305 310 315 320

Phe Thr Ala Ser Gly His Pro Val Gly Cys Ala lie Ala Leu Lys Ala 325 330 335 lie Asp Val Val Met Asn Glu Gly Leu Ala Glu Asn Val Arg Arg Leu 340 345 350

Ala Pro Arg Phe Glu Glu Arg Leu Lys His lie Ala Glu Arg Pro Asn 355 360 365 lie Gly Glu Tyr Arg Gly lie Gly Phe Met Trp Ala Leu Glu Ala Val 370 375 380

Lys Asp Lys Ala Ser Lys Thr Pro Phe Asp Gly Asn Leu Ser Val Ser 385 390 395 400 4 201127961

Glu Arg lie Ala Asn Thr Cys Thr 405 Leu Gly Gin Ser Val Val Leu Cys 420 Gin Met Asp Glu Met Phe Asp Lys 435 440 Phe Ala Glu Val Ala 450

Asp Leu Gly Leu lie Cys Arg Pro 410 415

Pro Pro Phe He Leu Thr Glu Ala 425 430

Leu Glu Lys Ala Leu Asp Lys Val 445 &lt;210&gt; 3 &lt;211&gt; 1362 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Vibrio fluvialis JSI7 ω-amine FTase codonoptimized gene &lt; 400 &gt; 3 atgaataaac cacagtcttg ggaagctcgt gctgaaacct atagcctgta cggctttacc 60 gatatgccgt ctctgcacca gcgtggtact gtagtggtaa cgcacggtga gggcccgtac 120 atcgtggacg ttaatggccg ccgttacctg gatgcaaaca gcggcctgtg gaacatggtt 180 gcgggcttcg accacaaagg cctgatcgat gccgcaaaag cgcagtacga acgcttcccg 240 ggttatcacg cgttctttgg ccgtatgagc gaccagactg tgatgctgag cgaaaaactg 300 gttgaagtgt ccccgttcga tagcggtcgt gtcttttaca ctaactctgg cagcgaggct 360 aacgatacca tggttaagat gctgtggttc ctgcacgcag cggaaggcaa acctcagaaa 420 cgtaaaattc tgacccgttg gaacgcttat cacggtgtga ctgctgtttc cgcatctatg 480 accggtaaac cgtataacag cgtgttcggt ctgccgctgc ctggcttcgt gcatctgacc 540 tgcccgcact actggcgtta tggtgaggaa ggcgaaactg aggaacagtt cgtggcgcgt 600 ctggctcgtg aactggaaga aaccattcaa cgcgaaggtg cagatactat cgcgggc ttc 660 tttgcggagc ctgttatggg tgccggcggt gtgattccgc cggcgaaggg ctatttccag 720 gcaatcctgc cgatcctgcg caagtacgac attccggtta tttctgacga agtgatctgc 780 ggcttcggcc gcaccggtaa cacctggggc tgcgtgacgt atgacttcac tccggacgca 840 atcattagct ctaaaaacct gactgcgggt ttcttcccta tgggcgccgt aatcctgggc 900 ccagaactgt ctaagcgcct ggaaaccgcc atcgaggcaa tcgaagagtt cccgcacggt 960 1020 201127961 ttcactgcta gcggccatcc ggtaggctgc gcaatcgcgc tgaaggcgat cgatgttgtc atgaacgagg gcctggcgga aaacgtgcgc cgcctggcgc cgcgttttga agaacgtctg aaacacattg ctgagcgccc gaacattggc gaatatcgcg gcatcggttt catgtgggcc ctggaagcag ttaaagataa agctagcaag accccgttcg acggcaacct gtccgtgagc gaacgtatcg ctaatacctg tacggacctg ggtctgatct gccgtccgct gggtcagtcc gtagttctgt gcccaccatt tatcctgacc gaagcgcaga tggatgaaat gttcgataaa ctggagaaag ctctggataa agtgttcgct gaagtcgcgt aa &lt; 210 &gt; 4 &lt; 211 &gt; 1350

&lt;212&gt; DNA &lt;213&gt; Bacillus weihenstephanensis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) . . (1350) &lt;400&gt; 4 gtg caa gcg acg gag caa aca caa Agt ttg aaa aaa aca gat gaa aag

Val Gin Ala Thr Glu Gin Thr Gin Ser Leu Lys Lys Thr Asp Glu Lys 15 10 15 tac ctt tgg cat gcg atg aga gga gca gcc cct agt cca acg aat tta

Tyr Leu Trp His Ala Met Arg Gly Ala Ala Pro Ser Pro Thr Asn Leu 20 25 30 att ate aca aaa gca gaa ggg gca tgg gtg acg gat att gat gga aac lie lie Thr.Lys Ala Glu Gly Ala Trp Val Thr Asp lie Asp Gly Asn 35 40 45 cgt tat tta gac ggt atg tcc ggt ctt tgg tgc gtg aat gtt ggg tat

Arg Tyr Leu Asp Gly Met Ser Gly Leu Trp Cys Val Asn Val Gly Tyr 50 55 60 ggt ega aaa gaa ctt gca aga gcg gcg ttt gaa cag ctt gaa gaa atg

Gly Arg Lys Glu Leu Ala Arg Ala Ala Phe Glu Gin Leu Glu Glu Met 65 70 75 80 ccg tat ttc cct ctg act caa agt cat gtt cct get att aaa tta gca

Pro Tyr Phe Pro Leu Thr Gin Ser His Val Pro Ala lie Lys Leu Ala 85 90 95 gaa aaa ttg aat gaa tgg ctt gat gat gaa tac gtc att ttc ttt tet

Glu Lys Leu Asn Glu Trp Leu Asp Asp Glu Tyr Val lie Phe Phe Ser 100 105 110 aac agt gga teg gaa gcg aat gaa aca gca ttt aaa att get cgt caa

Asn Ser Gly Ser Glu Ala Asn Glu Thr Ala Phe Lys lie Ala Arg Gin 115 120 125 tat cat caa caa aaa ggt gat cat gga ege tat aag ttt att tcc ege 1080 1140 1200 1260 1320 1362 48 96 144 192 240 288 336 384 432 6 201127961

Tyr His Gin Gin Lys Gly Asp His Gly Arg Tyr Lys Phe lie Ser Arg 130 135 140 tac cgc get tat cac ggt aac tea atg gga get ett gca gca aca ggt 480

Tyr Arg Ala Tyr His Gly Asn Ser Met Gly Ala Leu Ala Ala Thr Gly 145 150 155 160 caa gca cag ega aag tat aaa tat gaa cca etc ggg caa gga ttc ctg 528

Gin Ala Gin Arg Lys Tyr Lys Tyr Glu Pro Leu Gly Gin Gly Phe Leu 165 170 175 cat gta gca ccg cct gat aeg tat ega aat cca gag gat gtt cat aca 576

His Val Ala Pro Pro Asp Thr Tyr Arg Asn Pro Glu Asp Val His Thr 180 185 190 ctg gca agt get gag gaa ate gat cgt gtc atg aca tgg gag tta age 624

Leu Ala Ser Ala Glu Glu lie Asp Arg Val Met Thr Trp Glu Leu Ser 195 200 205 caa aca gta gee ggt gtg att gag cca ate att act ggg ggc gga 672

Gin Thr Val Ala Gly Val lie Met Glu Pro lie lie Thr Gly Gly Gly 210 215 220 att tta atg cct cct gat gga tat atg gga aaa gta aaa gaa att tgc 720 lie Leu Met Pro Pro Asp Gly Tyr Met Gly Lys Val Lys Glu Lie Cys 225 230 235 240 gag aag cac ggt geg ttg etc att tgt gat gaa gtt ata tgt gga ttt 768

Glu Lys His Gly Ala Leu Leu lie Cys Asp Glu Val lie Cys Gly Phe 245 250 255 ggc egg aca ggg aag cca ttt gga ttt atg aat tat ggc gtc aaa cca 816

Gly Arg Thr Gly Lys Pro Phe Gly Phe Met Asn Tyr Gly Val Lys Pro 260 265 270 gat ate att aca atg gca aaa ggt att aca agt geg tat ett cct ttg 864

Asp lie lie Thr Met Ala Lys Gly lie Thr Ser Ala Tyr Leu Pro Leu 275 280 285 tea gca aca gca gtt aga ega gag gtt tat gag gca ttc gta ggt agt 912

Ser Ala Thr Ala Val Arg Arg Glu Val Tyr Glu Ala Phe Val Gly Ser 290 295 300 gat gat tat gat cgc ttc cgc cat gta aat aeg ttc gga ggg aat cct 960

Asp Asp Tyr Asp Arg Phe Arg His Val Asn Thr Phe Gly Gly Asn Pro 305 310 315 320 get get tgc get tta get ttg aag aat tta gaa att atg gag aat gag 1008

Ala Ala Cys Ala Leu Ala Leu Lys Asn Leu Glu lie Met Glu Asn Glu 325 330 335 aaa etc att gaa cgt tee aaa gaa ttg ggt gaa ega ctg tta tat gag 1056

Lys Leu lie Glu Arg Ser Lys Glu Leu Gly Glu Arg Leu Leu Tyr Glu 340 345 350 eta gag gat gta aaa gag cat cca aac gta ggg gat gtt cgc gga aag 1104

Leu Glu Asp Val Lys Glu His Pro Asn Val Gly Asp Val Arg Gly Lys 355 360 365 7 ^ 1152 201127961 ggc ctt ctt tta ggc att gaa eta gtg gaa gat aag caa aca aaa gaa Gly Leu Leu Leu Gly lie Glu Leu Val Glu Asp Lys Gin Thr Lys Glu 370 375 380 ccg get tcc att gaa aag atg aac aaa gtc ate aat get tgt aaa gaa Pro Ala Ser lie Glu Lys Met Asn Lys Val lie Asn Ala Cys Lys Glu 385 390 395 400 aaa ggt eta att att ggt Aaa aat ggt gac act gtc gca ggt tac aat Lys Gly Leu lie lie Gly Lys Asn Gly Asp Thr Val Ala Gly Tyr Asn 405 410 415 aat att ttg cag ctt gca cct cca tta age ate aca gag gaa gac ttt Asn lie Leu Gin Leu Ala Pro Pro Leu Ser lie Thr Glu Glu Asp Phe 420 425 430 act ttt ate gtt aaa aca atg aaa gaa tgt tta tcc ege att aac ggg Thr Phe lie Val Lys Thr Met Lys Glu Cys Leu Ser Arg lie Asn Gly 435 440 445 cag Taa Gin 1200 1248 1296 1344 1350 &lt;210&gt; 5 &lt;211&gt; 449

&lt;212&gt; PRT &lt;213&gt; Bacillus weihenstephanensis &lt;400&gt; 5

Val Gin Ala Thr Glu Gin Thr Gin Ser Leu Lys Lys Thr Asp Glu Lys 15 10 15

Tyr Leu Trp His Ala Met Arg Gly Ala Ala Pro Ser Pro Thr Asn Leu 20 25 30 lie lie Thr Lys Ala Glu Gly Ala Trp Val Thr Asp lie Asp Gly Asn 35 40 45

Arg Tyr Leu Asp Gly Met Ser Gly Leu Trp Cys Val Asn Val Gly Tyr 50 55 60

Gly Arg Lys Glu Leu Ala Arg Ala Ala Phe Glu Gin Leu Glu Glu Met 65 70 75 80

Pro Tyr Phe Pro Leu Thr Gin Ser His Val Pro Ala lie Lys Leu Ala 85 90 95

Glu Lys Leu Asn Glu Trp Leu Asp Asp Glu Tyr Val lie Phe Phe Ser 100 105 110 8 201127961

Asn Ser Gly Ser Glu Ala Asn Glu Thr Ala Phe Lys lie Ala Arg Gin 115 120 125

Tyr His Gin Gin Lys Gly Asp His Gly Arg Tyr Lys Phe lie Ser Arg 130 135 140

Tyr Arg Ala Tyr His Gly Asn Ser Met Gly Ala Leu Ala Ala Thr Gly 145 150 155 160

Gin Ala Gin Arg Lys Tyr Lys Tyr Glu Pro Leu Gly Gin Gly Phe Leu 165 170 175

His Val Ala Pro Pro Asp Thr Tyr Arg Asn Pro Glu Asp Val His Thr 180 185 190

Leu Ala Ser Ala Glu Glu lie Asp Arg Val Met Thr Trp Glu Leu Ser 195 200 205

Gin Thr Val Ala Gly Val lie Met Glu Pro lie lie Thr Gly Gly Gly 210 215 220 lie Leu Met Pro Pro Asp Gly Tyr Met Gly Lys Val Lys Glu lie Cys 225 230 235 240

Glu Lys His Gly Ala Leu Leu lie Cys Asp Glu Val lie Cys Gly Phe 245 250 255

Gly Arg Thr Gly Lys Pro Phe Gly Phe Met Asn Tyr Gly Val Lys Pro 260 265 270

Asp lie lie Thr Met Ala Lys Gly lie Thr Ser Ala Tyr Leu Pro Leu 275 280 285

Ser Ala Thr Ala Val Arg Arg Glu Val Tyr Glu Ala Phe Val Gly Ser 290 295 300

Asp Asp Tyr Asp Arg Phe Arg His Val Asn Thr Phe Gly Gly Asn Pro 305 310 315 320

Ala Ala Cys Ala Leu Ala Leu Lys Asn Leu Glu lie Met Glu Asn Glu 325 330 335

Lys Leu lie Glu Arg Ser Lys Glu Leu Gly Glu Arg Leu Leu Tyr Glu 340 345 350 9 201127961

Leu Glu Asp Val Lys Glu His Pro Asn Val Gly Asp Val Arg Gly Lys 355 360 365

Gly Leu Leu Leu Gly He Glu Leu Val Glu Asp Lys Gin Thr Lys Glu 370 375 380

Pro Ala Ser He Glu Lys Met Asn Lys Val He Asn Ala Cys Lys Glu 385 390 395 400

Lys Gly Leu He He Gly Lys Asn Gly Asp Thr Val Ala Gly Tyr Asn 405 410 415

Asn lie Leu Gin Leu Ala Pro Pro Leu Ser lie Thr Glu Glu Asp Phe 420 425 430

Thr Phe lie Val Lys Thr Met Lys Glu Cys Leu Ser Arg lie Asn Gly 435 440 445

Gin &lt;210&gt; 6 &lt;211&gt; 1350 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; B. weihenstephanensis KBAB4 Aminotransferase Codon Optimization Gene &lt; 400 &gt; 6 atgcaggcta ccgaacaaac ccaatctctg aaaaagactg acgaaaaata tctgtggcac 60 gcgatgcgcg gtgcagctcc gtctccgacc aacctgatta ttaccaaagc tgaaggcgcg 120 tgggtgaccg acattgacgg taaccgttat ctggatggca tgagcggcct gtggtgtgtt 180 aatgtcggtt atggccgtaa ggagctggcg cgcgcggcat ttgaacaact ggaagaaatg 240 ccgtacttcc cgctgactca aagccatgtg ccggctatca aactggcgga aaaactgaac 300 gaatggctgg acgacgaata cgtgattttc ttctctaatt ctggctccga agcaaacgaa 360 accgcattca aaatcgcccg tcaatatcac cagcagaaag gtgaccacgg ccgctataaa 420 ttcatcagcc gttatcgtgc ataccatggt aattctatgg gtgcgctggc tgctaccggt 480 caggctcagc gcaaatacaa gtacgaaccg ctgggtcagg gttttctgca cgttgcacca 540 ccggatacct accgtaaccc ggaagacgtc cacaccctgg cttctgccga agaaatcgat 600 10 660 201127961 cgtgttatga cctgggagct gtcccagact gttgcgggtg ttatcatgga acctattatt accggtggtg g cattctgat gccgccggac ggttatatgg gtaaagtcaa ggaaatctgc gaaaaacacg gcgcgctgct gatctgcgat gaagttatct gtggcttcgg tcgcaccggc aaaccatttg gcttcatgaa ttatggcgta aaacctgaca ttattaccat ggctaaaggc attacttccg cttatctgcc gctgagcgcg accgcagttc gccgcgaagt ttatgaagcg tttgttggtt ctgatgatta cgaccgtttc cgtcatgtaa acacgtttgg cggtaaccca gcggcatgtg cgctggcgct gaaaaacctg gaaatcatgg aaaacgaaaa gctgatcgaa cgtagcaaag aactgggtga acgtctgctg tacgaactgg aagatgtcaa agaacacccg aacgtgggcg atgttcgcgg taaaggcctg ctgctgggta ttgaactggt tgaagacaaa cagaccaagg aaccggcttc cattgaaaag Atgaacaaag tgattaacgc gtgcaaagag aaaggcctga tcattggtaa gaacggtgat accgtggcag gttataacaa cattctgcag ctggcgccgc ctctgagcat cactgaagaa gatttcacct tcatcgtcaa aactatgaag gagtgcctga gccgcatcaa tggtcagtaa 1350 &lt;210&gt; 7 &lt;211&gt; 13 71

&lt;212&gt; DNA &lt;213&gt; Pseudomonas aeruginosa &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1)..(1371) &lt;400&gt; 7 atg aac age caa ate acc Aac gee aag acc cgt gag tgg cag geg ttg

Met Asn Ser Gin lie Thr Asn Ala Lys Thr Arg Glu Trp Gin Ala Leu 15 10 15 age ege gac cac cat ctg ccg ccg ttc acc gac tac aag cag ttg aac

Ser Arg Asp His His Leu Pro Pro Phe Thr Asp Tyr Lys Gin Leu Asn 20 25 30 gag aag ggc geg egg ate ate acc aag gee gaa ggc gtc tat ate tgg

Glu Lys Gly Ala Arg lie lie Thr Lys Ala Glu Gly Val Tyr lie Trp 35 40 45 gac age gag ggc aac aag ate etc gat geg atg gee ggc etc tgg tgc

Asp Ser Glu Gly Asn Lys lie Leu Asp Ala Met Ala Gly Leu Trp Cys 50 55 60 gtc aac gtc ggc tac ggc ege gag gag ctg gtc cag gee gee acc egg

Val Asn Val Gly Tyr Gly Arg Glu Glu Leu Val Gin Ala Ala Thr Arg 65 70 75 80 cag atg ege gag ttg ccg ttc tac aac ctg ttc ttc cag acc gee cac 720 780 840 900 960 1020 1080 1140 1200 1260 1320 48 96 144 192 240 288 11 £ 201127961

Gin Met Arg Glu Leu Pro Phe Tyr Asn Leu Phe Phe Gin Thr Ala His 85 90 95 ccg ccg gtg gtc gag ctg gcc aag geg ate gee gac gtc get ccg gaa 336 Pro Pro Val Val Glu Leu Ala Lys Ala lie Ala Asp Val Ala Pro Glu 100 105 110 ggc atg aac cac gtg ttc ttc acc ggc tee ggc tee gag gcc aac gac 384 Gly Met Asn His Val Phe Phe Thr Gly Ser Gly Ser Glu Ala Asn Asp 115 120 125 acc gtg ctg cgt atg gtc ege cac tat Tgg geg acc aag ggc cag ccg 432 Thr Val Leu Arg Met Val Arg His Tyr Trp Ala Thr Lys Gly Gin Pro 130 135 140 cag aag aaa gtg gtg ate ggc ege tgg aac ggc tac cac 99C tee acc 480 Gin Lys Lys Val val lie Gly Arg Trp Asn Gly Tyr His Gly Ser Thr 145 150 155 160 gtc gcc ggc gtc age ctg ggc ggc atg aag geg ttg cat gag cag ggt 528 Val Ala Gly Val Ser Leu Gly Gly Met Lys Ala Leu His Glu Gin Gly 165 170 175 Gat ttc ccc ate ccg ggc ate gtc cac ate gcc cag ccc tac tgg tac 576 Asp Phe Pro lie Pro Gly lie Val His lie Ala Gin Pro Tyr Trp Tyr 180 185 190 ggc gag ggc ggc gac atg teg ccg gac gag ttc ggc gtc tgg gcc gcc 624 Gly Glu Gly Gly Asp Met Ser Pro Asp Glu Phe Gly Val Trp Ala Ala 195 200 205 gag cag Ttg gag aag aag att etc gaa gtg ggc gag gaa aac gtc gcc 672 Glu Gin Leu Glu Lys Lys lie Leu Glu Val Gly Glu Glu Asn Val Ala 210 215 220 gcc ttc ate gcc gag ccg ate cag ggc gee ggc ggc gtg ate gtc ccg 720 Ala Phe lie Ala Glu Pro lie Gin Gly Ala Gly Gly Val lie Val Pro 225 230 235 240 ccg gac acc tac tgg ccg aag ate ege gag ate etc gcc aag tac gac 768 Pro Asp Thr Tyr Trp Pro Lys He Arg Glu lie Leu Ala Lys Tyr Asp 245 250 255 ate ctg ttc ate gcc gac gaa gtg ate tgc ggc ttc ggc cgt acc ggc 816 lie Leu Phe lie Ala Asp Glu Val He Cys Gly Phe Gly Arg Thr Gly 260 265 270 gag tgg ttc ggc age cag tac Tac ggc aac gcc ccg gac ctg atg ccg 864 Glu Trp Phe Gly Ser Gin Tyr Tyr Gly Asn Ala Pro Asp Leu Met Pro 275 280 285 ate gcc aag ggc etc acc tee ggc tac ate ccc atg ggc ggg gtg gtg 912 lie Ala Lys Gly Leu Thr Ser Gly Tyr lie Pro Met Gly Gly Val Val 290 295 300 gtg ege gac gag ate gtc gaa gtg etc Aac cag ggc ggc gag ttc tac 960 Val Arg Asp Glu lie Val Glu Val Leu Asn Gin Gly Glu Glu Phe Tyr 305 310 315 320 12 201127961 cac ggc ttc acc tat tcc ggt cac ccg gtg 9cg gcc gcc gtg gcc ctg 1008

His Gly Phe Thr Tyr Ser Gly His Pro Val Ala Ala Ala Val Ala Leu 325 330 335 gag aac ate ege ate ctg ege gaa gag aag ate ate gag aag gtg aag 1056

Glu Asn lie Arg lie Leu Arg Glu Glu Lys lie lie Glu Lys Val Lys 340 345 350 geg gaa aeg gca ccg tat ttg cag aaa ege tgg cag gag ctg gcc gac 1104

Ala Glu Thr Ala Pro Tyr Leu Gin Lys Arg Trp Gin Glu Leu Ala Asp 355 360 365 cac ccg ttg gtg ggc gaa geg ege ggg gtc ggc atg gtc gcc gcc ctg 1152

His Pro Leu Val Gly Glu Ala Arg Gly Val Gly Met Val Ala Ala Leu 370 375 380 gag ctg gtc aag aac aag aag acc ege gag cgt ttc acc gac aag ggc 1200

Glu Leu Val Lys Asn Lys Lys Thr Arg Glu Arg Phe Thr Asp Lys Gly 385 390 395 400 gtc ggg atg ctg tgc egg gaa cat tgt ttc ege aac ggt ttg ate atg 1248

Val Gly Met Leu Cys Arg Glu His Cys Phe Arg Asn Gly Leu lie Met 405 410 415 ege geg gtg ggc gac act atg att ate teg ccg ccg ctg gtg ate gat 1296

Arg Ala Val Gly Asp Thr Met lie lie Ser Pro Pro Leu Val lie Asp 420 425 430 ccg teg cag ate gat gag ttg ate acc ctg geg ege aag tgc etc gat 1344

Pro Ser Gin lie Asp Glu Leu lie Thr Leu Ala Arg Lys Cys Leu Asp 435 440 445 cag acc gcc gcc gcc gtc ctg get tga 1371

Gin Thr Ala Ala Ala Val Leu Ala 450 455 &lt;210&gt; 8 &lt;211&gt; 456

&lt;212&gt; PRT &lt;213&gt; Pseudomonas aeruginosa} &lt;400&gt; 8

Met Asn Ser Gin lie Thr Asn Ala Lys Thr Arg Glu Trp Gin Ala Leu 15 10 15

Ser Arg Asp His His Leu Pro Pro Phe Thr Asp Tyr Lys Gin Leu Asn 20 25 3〇

Glu Lys Gly Ala Arg lie lie Thr Lys Ala Glu Gly Val Tyr lie Trp 35 40 45

Asp Ser Glu Gly Asn Lys lie Leu Asp Ala Met Ala Gly Leu Trp Cys 50 55 60 13 M, 201127961

Val Asn Val Gly Tyr Gly Arg Glu Glu Leu Val Gin Ala Ala Thr Arg 65 70 75 80

Gin Met Arg Glu Leu Pro Phe Tyr Asn Leu Phe Phe Gin Thr Ala His 85 90 95

Pro Pro Val Val Glu Leu Ala Lys Ala lie Ala Asp Val Ala Pro Glu 100 105 110

Gly Met Asn His Val Phe Phe Thr Gly Ser Gly Ser Glu Ala Asn Asp 115 120 125

Thr Val Leu Arg Met Val Arg His Tyr Trp Ala Thr Lys Gly Gin Pro 130 135 140

Gin Lys Lys Val Val lie Gly Arg Trp Asn Gly Tyr His Gly Ser Thr 145 150 155 160

Val Ala Gly Val Ser Leu Gly Gly Met Lys Ala Leu His Glu Gin Gly 165 170 175

Asp Phe Pro lie Pro Gly lie Val His lie Ala Gin Pro Tyr Trp Tyr 180 185 190

Gly Glu Gly Gly Asp Met Ser Pro Asp Glu Phe Gly Val Trp Ala Ala 195 200 205

Glu Gin Leu Glu Lys Lys lie Leu Glu Val Gly Glu Glu Asn Val Ala 210 215 220

Ala Phe lie Ala Glu Pro lie Gin Gly Ala Gly Gly Val lie Val Pro 225 230 235 240

Pro Asp Thr Tyr Trp Pro Lys lie Arg Glu lie Leu Ala Lys Tyr Asp 245 250 255 lie Leu Phe lie Ala Asp Glu Val lie Cys Gly Phe Gly Arg Thr Gly 260 265 270

Glu Trp Phe Gly Ser Gin Tyr Tyr Gly Asn Ala Pro Asp Leu Met Pro 275 280 285 lie Ala Lys Gly Leu Thr Ser Gly Tyr lie Pro Met Gly Gly Val Val 290 295 300 14 201127961

Val Arg Asp Glu lie Val Glu Val Leu Asn Gin Gly Gly Gin Phe Tyr 305 310 315 320

His Gly Phe Thr Tyr Ser Gly His Pro Val Ala Ala Ala Val Ala Leu 325 330 335

Glu Asn lie Arg lie Leu Arg Glu Glu Lys lie lie Glu Lys Val Lys 340 345 350

Ala Glu Thr Ala Pro Tyr Leu Gin Lys Arg Trp Gin Glu Leu Ala Asp 355 360 365

His Pro Leu Val Gly Glu Ala Arg Gly Val Gly Met Val Ala Ala Leu 370 375 380

Glu Leu Val Lys Asn Lys Lys Thr Arg Glu Arg Phe Thr Asp Lys Gly 385 390 395 400

Val Gly Met Leu Cys Arg Glu His Cys Phe Arg Asn Gly Leu lie Met 405 410 415

Arg Ala Val Gly Asp Thr Met 420

Pro Ser Gin lie Asp Glu Leu 435 lie lie Ser Pro Pro Leu Val lie Asp 425 430 lie Thr Leu Ala Arg Lys Cys Leu Asp 440 445

Gin Thr Ala Ala Ala Val Leu Ala 450 455 &lt;210&gt; 9 &lt;211&gt; 70 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Introduction &lt;400&gt; 9 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgaaca Gccaaatcac 60 caacgccaag 70 &lt;210&gt; 10 &lt;211&gt; 49

&lt;212&gt; DNA 15 49201127961 &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Introduction &lt;400&gt; 10 ggggaccact ttgtacaaga aagctgggtt caagccagga cggcggcgg &lt;210&gt; 11 &lt;211&gt; 1365 &lt;212&gt; DNA &lt;213&gt; Pseudomonas syringae &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) . . (1365) &lt;400&gt; 11 atg agt gcc aac aac ccg caa acc etc gaa tgg cag gcc ctg Age age Met Ser Ala Asn Asn Pro Gin Thr Leu Glu Trp Gin Ala Leu Ser Ser 1 5 10 15 gag cat cac ctg gca ccg ttc age gac tac aaa caa ctg aaa gag aaa Glu His His Leu Ala Pro Phe Ser Asp Tyr Lys Gin Leu Lys Glu Lys 20 25 30 ggc ccg ege ate ate acc cgt gcc gag ggc gtt tat ctg tgg gac age Gly Pro Arg lie lie Thr Arg Ala Glu Gly Val Tyr Leu Trp Asp Ser 35 40 45 gag ggc aac aag ate etc gat ggc Atg tee ggc ctg tgg tgc gtg gcc Glu Gly Asn Lys lie Leu Asp Gly Met Ser Gly Leu Trp Cys Val Ala 50 55 60 ate ggt tat ggc ege gaa gaa ctg gcc gac gca gcc age aaa cag atg lie Gly Tyr Gly Arg Glu Glu Leu Ala Asp Ala Ala Ser Lys Gi n Met 65 70 75 80 ege gag ctg ccg tac tac aac ctg ttc ttc cag acc gcc cac ccg ccg Arg Glu Leu Pro Tyr Tyr Asn Leu Phe Phe Gin Thr Ala His Pro Pro 85 90 95 gtg ctg gaa ctg gcc aag gcc ate tee Gac ate get ccc gag ggc atg Val Leu Glu Leu Ala Lys Ala lie Ser Asp lie Ala Pro Glu Gly Met 100 105 110 aac cat gtg ttc ttc acc ggt tea ggc tet gaa ggc aat gac aeg atg Asn His Val Phe Phe Thr Gly Ser Gly Ser Glu Gly Asn Asp Thr Met 115 120 125 ctg ege atg gtt cgt cat tac tgg geg ctg aaa ggc cag ccg aac aag Leu Arg Met Val Arg His Tyr Trp Ala Leu Lys Gly Gin Pro Asn Lys 130 135 140 aaa acc ate ate Age ege gtc aat ggc tac cac ggc tee acc gtc gcc Lys Thr lie lie Ser Arg Val Asn Gly Tyr His Gly Ser Thr Val Ala 145 150 155 160 48 96 144 192 240 288 336 384 432 16 480 528201127961 ggt gcc age ctg ggt ggc Atg acc tac atg cac gaa cag ggc gac ctg Gly Ala Ser Leu Gly Gly Met Thr Tyr Met His Glu Gin Gly Asp Leu 165 170 175 ccg ate ccg ggg gtg gtg cac att cca cag cct tac tgg ttc ggc gaa Pro lie Pro Gly Val Val His lie Pro Gin Pro Tyr Trp Phe Gly Glu 180 185 190 ggc ggc gac atg aeg ccg gac gag ttc ggc ate tgg geg gcc gag caa Gly Gly Asp Met Thr Pro Asp Glu Phe Gly lie Trp Ala Ala Glu Gin 195 200 205 ctg gaa Aag aaa att etc gag ctg ggc gtc gag aac gtc ggt geg ttc Leu Glu Lys Lys lie Leu Glu Leu Gly Val Glu Asn Val Gly Ala Phe 210 215 220 att gcc gag cca ate cag ggc geg ggc ggt gtg att gtc ccg cct gat lie Ala Glu Pro lie Gin Gly Ala Gly Gly Val lie Val Pro Pro Asp 225 230 235 240 tcc tac tgg ccg aag ate aag ga ate ett tee ege tac gac ate ctg Ser Tyr Trp Pro Lys lie Lys Glu lie Leu Ser Arg Tyr Asp lie Leu 245 250 255 ttc gcc gcc gat gag gtg att tgt ggc ttc ggg cgt acc agt gag tgg Phe Ala Ala Asp Glu Val lie Cys Gly Phe Gly Arg Thr Ser Glu Trp 260 265 270 ttc ggt age gat ttc tat ggc etc agg ccg gac Atg atg acc ate gcc Phe Gly Ser Asp Phe Tyr Gly Leu Arg Pro Asp Met Met Thr lie Ala 275 280 285 aaa ggc ctg acc tec ggt tac gta ccg atg ggc ggc ctg ate gtg ege Lys Gly Leu Thr Ser Gly Tyr Val Pro Met Gly Gly Leu lie Val Arg 290 295 300 gat gaa ate gtt geg gtg etc aat gag ggt ggc gat ttc aat cac ggc Asp Glu lie Val Ala Val Leu Asn Glu Gly Gly Asp Phe Asn His Gly 305 310 315 320 ttt acc tac tec ggg Cac ccg gtg geg gcc geg gtt geg ctg gag aac Phe Thr Tyr Ser Gly His Pro Val Ala Ala Ala Val Ala Leu Glu Asn 325 330 335 ate cgt ate ctg ege gaa gaa aag ate gtc gaa egg gtc agg teg gaa lie Arg lie Leu Arg Glu Glu Lys lie Val Glu Arg Val Arg Ser Glu 340 345 350 aeg gca ccg tat ttg caa aag cgt ttg cgt gag ttg age gat cat ccg Thr Ala Pro Tyr Leu Gin Lys Arg Leu Arg Glu Leu Ser Asp His Pro 355 360 365 Ctg gtg ggc gaa gtc egg ggt gtc ggg ctg etc ggg gcc att gag ctg Leu Val Gly Glu Val Arg Gly Val Gly Leu Leu Gly Ala lie Glu Leu 370 375 380 gtg aag gac aag acc acc ege gag ege tat acc gac aag ggc geg Gga Val Lys Asp Lys Thr Thr Arg Glu Arg Tyr Thr Asp Lys Gly Ala Gly 576 624 672 720 768 816 864 912 960 1008 1056 1104 1152 17

S 1200 1248 201127961 385 390 395 400 atg ate tgt ega acc ttc tgc ttc gac aat ggc ctg ate atg egg get Met lie Cys Arg Thr Phe Cys Phe Asp Asn Gly Leu lie Met Arg Ala 405 410 415 gtg ggc gat acc atg ate att Geg ccg cca ctg gtg ate agt ttt geg Val Gly Asp Thr Met lie lie Ala Pro Pro Leu Val lie Ser Phe Ala 420 425 430 caa ate gat gag ctg gta gag aag geg ege aeg tgt ctg gat ctg aeg Gin lie Asp Glu Leu Val Glu Lys Ala Arg Thr Cys Leu Asp Leu Thr 435 440 445 ctg geg gtg ttg cag ggc tga Leu Ala Val Leu Gin Gly 450 1296 1344 1365 &lt;210&gt; 12 &lt;211&gt; 454

&lt;212&gt; PRT &lt;213&gt; Pseudomonas syringae &lt;400&gt; 12

Met Ser Ala Asn Asn Pro Gin Thr Leu Glu Trp Gin Ala Leu Ser Ser 15 10 15

Glu His His Leu Ala Pro Phe Ser Asp Tyr Lys Gin Leu Lys Glu Lys 20 25 30

Gly Pro Arg lie 35 lie Thr Arg Ala Glu Gly Val Tyr Leu Trp Asp Ser 40 45

Glu Gly Asn Lys lie Leu Asp Gly Met Ser Gly Leu Trp Cys Val Ala 50 55 60 lie Gly Tyr Gly Arg Glu Glu Leu Ala Asp Ala Ala Ser Lys Gin Met 65 70 75 80

Arg Glu Leu Pro Tyr Tyr Asn Leu Phe Phe Gin Thr Ala His Pro Pro 85 90 95

Val Leu Glu Leu Ala Lys Ala lie Ser Asp lie Ala Pro Glu Gly Met 100 105 110

Asn His Val Phe Phe Thr Gly Ser Gly Ser Glu Gly Asn Asp Thr Met 115 120 125

Leu Arg Met Val Arg His Tyr Trp Ala Leu Lys Gly Gin Pro Asn Lys 18 201127961 130 135 140

Lys Thr lie l Arg Val Asn Gly Tyr His Gly Ser Thr Val Ala 145 150 155 160

Gly Ala Ser Leu Gly Gly Met Thr Tyr Met His Glu Gin Gly Asp Leu 165 170 175

Pro lie Pro Gly Val Val His lie 180

Pro Gin Pro Tyr Trp Phe Gly Glu 185 190

Gly Gly Asp Met Thr Pro Asp Glu 195 200

Phe Gly lie Trp Ala Ala Glu Gin 205

Leu Glu Lys Lys lie Leu Glu Leu Gly Val Glu Asn Val Gly Ala Phe 210 215 220 lie Ala Glu Pro lie Gin Gly Ala Gly Gly Val He Val Pro Pro Asp 225 230 235 240

Ser Tyr Trp Pro Lys lie Lys Glu lie Leu Ser Arg Tyr Asp lie Leu 245 250 255

Phe Ala Ala Asp Glu Val lie Cys Gly Phe Gly Arg Thr Ser Glu Trp 260 265 270

Phe Gly Ser Asp Phe Tyr Gly Leu Arg Pro Asp Met Met Thr lie Ala 275 280 285

Lys Gly Leu Thr Ser Gly Tyr Val 290 295

Pro Met Gly Gly Leu lie Val Arg 300

Asp Glu lie Val Ala Val Leu Asn 305 310

Glu Gly Gly Asp Phe Asn His Gly 315 320

Phe Thr Tyr Ser Gly His Pro Val 325

Ala Ala Ala Val Ala Leu Glu Asn 330 335 lie Arg lie Leu Arg Glu Glu Lys 340 lie Val Glu Arg Val Arg Ser Glu 345 350

Thr Ala Pro Tyr Leu Gin Lys Arg 355 360

Leu Arg Glu Leu Ser Asp His Pro 365 19 201127961

Leu Val Gly Glu Val Arg Gly Val Gly Leu Leu Gly Ala lie Glu Leu 370 375 380

Val Lys Asp Lys Thr Thr Arg Glu Arg Tyr Thr Asp Lys Gly Ala Gly 385 390 395 400

Met lie Cys Arg Thr Phe Cys Phe Asp Asn Gly Leu lie Met Arg Ala 405 410 415

Val Gly Asp Thr Met lie lie Ala Pro Pro Leu Val lie Ser Phe Ala 420 425 430

Gin lie Asp Glu Leu Val Glu Lys Ala Arg Thr Cys Leu Asp Leu Thr 435 440 445

Leu Ala Val Leu Gin Gly 450 &lt;210&gt; 13 &lt;211&gt; 1365 &lt;212&gt; DNA &lt;213 &gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Pseudomonas syringae codon best of aminotransferase gene &lt; 400 &gt; 13 atgtctgcta acaatccaca aactctggaa tggcaggcac tgagctccga acatcacctg gctccgttct ccgactacaa acaactgaaa gagaaaggcc cgcgtatcat tacccgcgct gaaggtgtgt acctgtggga ttctgaaggc aacaaaattc tggacggtat gagcggcctg tggtgcgtag caatcggtta tggccgtgaa gaactggctg acgcggcgag caaacagatg cgtgaactgc cgtattataa cctgttcttc caaaccgcac acccgccggt tctggaactg gctaaagcta tcagcgatat cgcaccggag ggcatgaatc acgtcttctt cactggttcc ggtagcgaag gcaacgacac gatgctgcgc atggtacgtc actattgggc gctgaagggc cagccgaaca agaaaacgat tatcagccgt gtaaacggtt atcacggcag caccgttgcg ggtgcgagcc tgggcggtat gacctacatg cacgaacagg gtgacctgcc gatcccgggt gtagtgcaca ttccgcagcc gtattggttc ggtgaaggcg gtgacatgac gccggacgaa ttcggcatct gggcggcaga gcagctggaa aagaaaatcc tggaactggg cgtggaaaac gtcggcgcgt tcatcgcgga accgattcag ggcgcgggcg gcgt aattgt tccgccggac agctactggc caaaaatcaa agagatcctg tctcgttacg acatcctgtt cgccgcagac 60 120 180 240 300 360 420 480 540 600 660 720 20 780 840 201127961 gaagtgatct gcggttttgg ccgcacctct gaatggttcg gctccgactt ctacggtctg cgtccggaca tgatgaccat cgccaaaggc ctgacctccg gttatgttcc tatgggtggc ctgatcgtgc gcgacgaaat tgttgcggtt ctgaacgaag gcggcgattt caaccacggc ttcacctatt ccggtcaccc agttgctgct gctgtagcac tggaaaacat ccgcatcctg cgtgaagaaa agatcgtaga acgcgtacgt tccgaaaccg caccttacct gcagaagcgc ctgcgcgaac tgagcgacca ccctctggta ggtgaagttc gcggcgtggg cctgctgggc gcgatcgagc tggtgaaaga caaaactacc cgtgaacgtt acaccgacaa aggcgcaggc atgatctgcc gtaccttttg cttcgataac ggtctgatca tgcgcgcagt cggtgatacc atgatcattg ctccgcctct ggttatttct tttgcccaga ttgatgagct ggtcgaaaaa gcgcgcactt gtctggatct gactctggct gttctgcagg gttaa &lt; 210 &gt; 14 &lt; 211 &gt; 849

&lt;212&gt; DNA &lt;213&gt; Bacillus subtilis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) . . . (849) &lt;400&gt; 14 atg aag gtt tta gtc aat ggc egg Ctg att ggg ege agt gaa gca tea

Met Lys Val Leu Val Asn Gly Arg Leu lie Gly Arg Ser Glu Ala Ser 15 10 15 ate gat ttg gaa gat ege ggt tat cag ttt ggt gac ggc ate tat gaa lie Asp Leu Glu Asp Arg Gly Tyr Gin Phe Gly Asp Gly lie Tyr Glu 20 25 30 gtg ate agg gtg tac aaa gga gta ttg ttc ggc tta cgt gag cat gca

Val lie Arg Val Tyr Lys Gly Val Leu Phe Gly Leu Arg Glu His Ala 35 40 45 gag cgt ttt ttc aga agt get get gaa ate gga att tea ctg cca ttc

Glu Arg Phe Phe Arg Ser Ala Ala Glu lie Gly lie Ser Leu Pro Phe 50 55 60 agt ata gaa gat etc gag tgg gac ctg caa aag ett gta cag gaa aat

Ser lie Glu Asp Leu Glu Trp Asp Leu Gin Lys Leu Val Gin Glu Asn 65 70 75 80 geg gtc agt gag gga geg gta tac att cag aca aca aga ggt gtg gee

Ala Val Ser Glu Gly Ala Val Tyr He Gin Thr Thr Arg Gly Val Ala 85 90 95 ccg ega aaa cac cag tat gaa gee ggc etc gag ccg cag act act gee

Pro Arg Lys His Gin Tyr Glu Ala Gly Leu Glu Pro Gin Thr Thr Ala 100 105 110 900 960 1020 1080 1140 1200 1260 1320 1365 48 96 144 192 240 288 21 336 384201127961 tat acg ttt acg gtg aaa aaa ccg gag caa gag cag gca Tac gga gtg Tyr Thr Phe Thr Val Lys Lys Pro Glu Gin Glu Gin Ala Tyr Gly Val 115 120 125 9cg gcc att aca gat gag gat ctt cgc tgg tta aga tgt gat ate aaa Ala Ala lie Thr Asp Glu Asp Leu Arg Trp Leu Arg Cys Asp lie Lys 130 135 140 agt ctg aat tta ctg tat aat gtc atg acg aag caa agg gcc tat gaa Ser Leu Asn Leu Leu Tyr Asn Val Met Thr Lys Gin Arg Ala Tyr Glu 145 150 155 160 gcc gga gca ttt gaa gcc att Tt ctt agg gac ggc gtt gtt acg gag Ala Gly Ala Phe Glu Ala lie Leu Leu Arg Asp Gly Val Val Thr Glu 165 170 175 ggt aca tcc tet aac gtt tat gcc gtt ate aac ggc aca gtg ega aca Gly Thr Ser Ser Asn Val Tyr Ala Val lie Asn Gly Thr Val Arg Thr 180 185 190 cat ccg get aat egg etc att etc aat gga att aca egg atg aat att His Pro Ala Asn Arg Leu lie Leu Asn Gly lie Thr Arg Met Asn lie 195 200 205 tta gga ctg att gag aag aat ggg ate aaa ctg gat gag act cct gtc Leu Gly Leu lie Glu Lys Asn Gly lie Lys Leu Asp Glu Thr Pro Val 210 215 220 agt gaa gaa gag ttg aaa cag geg gaa gag ate ttt att teg Tea acg Ser Glu Glu Glu Leu Lys Gin Ala Glu Glu lie Phe lie Ser Ser Thr 225 230 235 240 acg gca gaa att att ccg gtc gtg acg etc gat gga caa teg ate gga Thr Ala Glu lie lie Pro Val Val Thr Leu Asp Gly Gin Ser lie Gly 245 250 255 agc ggg aaa ccc gga ccg gtg acc aaa cag ctt cag get get ttt caa Ser Gly Lys Pro Gly Pro Val Thr Lys Gin Leu Gin Ala Ala Phe Gin 260 265 270 gaa age att caa cag get get age Att tea taa Glu Ser L Gin Gin Ala Ala Ser lie Ser 275 280 432 480 528 576 624 672 720 768 816 849 &lt;210&gt; 15 &lt;211&gt; 282 &lt;212&gt; PRT &lt;213&gt; Bacillus subtilis (Bacillus Subtilis) &lt;400&gt; 15 Met Lys Val Leu Val Asn Gly Arg Leu lie Gly Arg Ser Glu Ala Ser 15 10 15 lie Asp Leu Glu Asp Arg Gly Tyr Gin Phe Gly Asp Gly lie Tyr Glu 20 25 30 22 201127961

Val lie Arg Val Tyr Lys Gly Val Leu Phe Gly Leu Arg Glu His Ala 35 40 45

Glu Arg Phe Phe Arg Ser Ala Ala Glu lie Gly lie Ser Leu Pro Phe 50 55 60

Ser He Glu Asp Leu Glu Trp Asp Leu Gin Lys Leu Val Gin Glu Asn 65 70 75 80

Ala Val Ser Glu Gly Ala Val Tyr lie Gin Thr Thr Arg Gly Val Ala 85 90 95

Pro Arg Lys His Gin Tyr Glu Ala Gly Leu Glu Pro Gin Thr Thr Ala 100 105 110

Tyr Thr Phe Thr Val Lys Lys Pro Glu Gin Glu Gin Ala Tyr Gly Val 115 120 125

Ala Ala lie Thr Asp Glu Asp Leu Arg Trp Leu Arg Cys Asp lie Lys 130 135 140

Ser Leu Asn Leu Leu Tyr Asn Val Met Thr Lys Gin Arg Ala Tyr Glu 145 150 155 160

Ala Gly Ala Phe Glu Ala lie Leu Leu Arg Asp Gly Val Val Thr Glu 165 170 175

Gly Thr Ser Ser Asn Val Tyr Ala Val lie Asn Gly Thr Val Arg Thr 180 185 190

His Pro Ala Asn Arg Leu lie Leu Asn Gly lie Thr Arg Met Asn lie 195 200 205

Leu Gly Leu lie Glu Lys Asn Gly lie Lys Leu Asp Glu Thr Pro Val 210 215 220

Ser Glu Glu Glu Leu Lys Gin Ala Glu Glu lie Phe lie Ser Ser Thr 225 230 235 240

Thr Ala Glu lie lie Pro Val Val Thr Leu Asp Gly Gin Ser lie Gly 245 250 255

Ser Gly Lys Pro Gly Pro Val Thr Lys Gin Leu Gin Ala Ala Phe Gin 23 201127961 260 265 270

Glu Ser lie Gin Gin Ala Ala Ser lie Ser 275 280 &lt;210&gt; 16 &lt;211&gt; 1347 &lt;212&gt; DNA &lt;213&gt; Bacillus subtilis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1} . . (1347) &lt;400&gt; 16 atg act cat gat ttg ata gaa aaa agt aaa aag cac etc tgg ctg cca Met Thr His Asp Leu lie Glu Lys Ser Lys Lys His Leu Trp Leu Pro 15 10 15 ttt Acc caa atg aaa gat tat gat gaa aac ccc tta ate ate gaa age Phe Thr Gin Met Lys Asp Tyr Asp Glu Asn Pro Leu lie lie Glu Ser 20 25 30 ggg act gga ate aaa gtc aaa gac ata aac ggc aag gaa tac tat gac Gly Thr Gly He Lys Val Lys Asp He Asn Gly Lys Glu Tyr Tyr Asp 35 40 45 ggt ttt tea teg gtt tgg ett aat gtc cac gga cac ege aaa aaa gaa Gly Phe Ser Ser Val Trp Leu Asn Val His Gly His Arg Lys Lys Glu 50 55 60 eta gat gac gcc ata aaa aaa cag etc gga aaa att geg cac tee aeg Leu Asp Asp Ala lie Lys Lys Gin Leu Gly Lys lie Ala His Ser Thr 65 70 75 80 tta ttg ggc atg acc aat gtt cca gca acc Cage ett gcc gaa aca tta Leu Leu Gly Met Thr Asn Val Pro Ala Thr Gin Leu Ala Glu Thr Leu 85 90 95 ate gac ate age cca aaa aag etc aeg egg gtc ttt tat tea gac age lie Asp lie Ser Pro Lys Lys Leu Thr Arg Val Phe Tyr Ser Asp Ser 100 105 110 Ggc gca gag geg atg gaa ata gcc eta aaa atg geg ttt cag tat tgg Gly Ala Glu Ala Met Glu lie Ala Leu Lys Met Ala Phe Gin Tyr Trp 115 120 125 aag aac ate ggg aag ccc gag aaa caa aaa ttc ate gca atg aaa Aac Lys Asn He Gly Lys Pro Glu Lys Gin Lys Phe lie Ala Met Lys Asn 130 135 140 ggg tat cac ggt gat aeg att ggc gcc gtc agt gtc ggt tea att gag Gly Tyr His Gly Asp Thr lie Gly Ala Val Ser Val Gly Ser Lie Glu 145 150 155 160 ett ttt cac cac gta tac ggc ccg ttg atg ttc gag agt tac aag gcc 48 96 144 192 240 288 336 384 432 480 528 24 201127961

Leu Phe His His Val Tyr Gly Pro Leu Met Phe Glu Ser Tyr Lys Ala 165 170 175 ccg att cct tat gtg tat cgt tct gaa age ggt gat cct gat gag tgc 576

Pro lie Pro Tyr Val Tyr Arg Ser Glu Ser Gly Asp Pro Asp Glu Cys 180 185 190 cgt gat cag tgc etc ega gag ett gca cag ctg ett gag gaa cat cat 624

Arg Asp Gin Cys Leu Arg Glu Leu Ala Gin Leu Leu Glu Glu His His 195 200 205 gag gaa att gcc geg ett tee att gaa tea atg gta caa ggc geg tee 672

Glu Glu lie Ala Ala Leu Ser lie Glu Ser Met Val Gin Gly Ala Ser 210 215 220 ggt atg ate gtg atg ccg gaa gga tat ttg gca ggc gtg ege gag eta 720

Gly Met lie Val Met Pro Glu Gly Tyr Leu Ala Gly Val Arg Glu Leu 225 230 235 240 tgt aca aca tac gat gtc tta atg ate gtt gat gaa gtc get aca ggc 768

Cys Thr Thr Tyr Asp Val Leu Met lie Val Asp Glu Val Ala Thr Gly 245 250 255 ttt ggc cgt aca gga aaa atg ttt geg tgc gag cac gag aat gtc cag 816

Phe Gly Arg Thr Gly Lys Met Phe Ala Cys Glu His Glu Asn Val Gin 260 265 270 cct gat ctg atg get gcc ggt aaa ggc att aca gga ggc tat ttg cca 864

Pro Asp Leu Met Ala Ala Gly Lys Gly lie Thr Gly Gly Tyr Leu Pro 275 280 285 att gee gtt aeg ttt gcc act gaa gac ate tat aag gca ttc tat gat 912 lie Ala Val Thr Phe Ala Thr Glu Asp lie Tyr Lys Ala Phe Tyr Asp 290 295 300 gat tat gaa aac eta aaa acc ttt ttc cat ggc cat tee tat aca ggc 960

Asp Tyr Glu Asn Leu Lys Thr Phe Phe His Gly His Ser Tyr Thr Gly 305 310 315 320 aat cag ett ggc tgt geg gtt geg ett gaa aat ctg gca tta ttt gaa 1008

Asn Gin Leu Gly Cys Ala Val Ala Leu Glu Asn Leu Ala Leu Phe Glu 325 330 335 tct gaa aac att gtg gaa caa gta geg gaa aaa agt aaa aag etc cat 1056

Ser Glu Asn lie Val Glu Gin Val Ala Glu Lys Ser Lys Lys Leu His 340 345 350 ttt ett ett caa gat ctg cac get ett cct cat gtt ggg gat att egg 1104

Phe Leu Leu Gin Asp Leu His Ala Leu Pro His Val Gly Asp lie Arg 355 360 365 cag ett ggc ttt atg tgc ggt gca gag ett gta ega tea aag gaa act 1152

Gin Leu Gly Phe Met Cys Gly Ala Glu Leu Val Arg Ser Lys Glu Thr 370 375 380 aaa gaa cct tac ccg get gat egg egg att gga tac aaa gtt tee tta 1200

Lys Glu Pro Tyr Pro Ala Asp Arg Arg lie Gly Tyr Lys Val Ser Leu 385 390 395 400 25 1248 201127961 aaa atg aga gag tta gga atg ctg aca aga ccg ctt ggg gac gtg att Lys Met Arg Glu Leu Gly Met Leu Thr Arg Pro Leu Gly Asp Val lie 405 410 415 gca ttt ctt cct cct ctt gcc age aca get gaa gag etc teg gaa atg Ala Phe Leu Pro Pro Leu Ala Ser Thr Ala Glu Glu Leu Ser Glu Met 420 425 430 gtt gcc att atg aaa caa geg Ate cac gag gtt aeg age ctt gaa gat Val Ala lie Met Lys Gin Ala lie His Glu Val Thr Ser Leu Glu Asp 435 440 445 tga 1296 1344 1347 &lt;210&gt; 17 &lt;211&gt; 448

&lt;212&gt; PRT &lt;213&gt; Bacillus subtilis &lt;400&gt; 17

Met Thr His Asp Leu lie Glu Lys Ser Lys Lys His Leu Trp Leu Pro 15 10 15

Phe Thr Gin Met Lys Asp Tyr Asp Glu Asn Pro Leu lie lie Glu Ser 20 25 30

Gly Thr Gly lie Lys Val Lys Asp lie Asn Gly Lys Glu Tyr Tyr Asp 35 40 45

Gly Phe Ser Ser Val Trp Leu Asn Val His Gly His Arg Lys Lys Glu 50 55 60

Leu Asp Asp Ala lie Lys Lys Gin Leu Gly Lys lie Ala His Ser Thr 65 70 75 80

Leu Leu Gly Met Thr Asn Val Pro Ala Thr Gin Leu Ala Glu Thr Leu 85 90 95 lie Asp lie Ser Pro Lys Lys Leu Thr Arg Val Phe Tyr Ser Asp Ser 100 105 110

Gly Ala Glu Ala Met Glu lie Ala Leu Lys Met Ala Phe Gin Tyr Trp 115 120 125

Lys Asn lie Gly Lys Pro Glu Lys Gin Lys Phe lie Ala Met Lys Asn 130 135 140

Gly Tyr His Gly Asp Thr lie Gly Ala Val Ser Val Gly Ser lie Glu 26 201127961 145 150 155 160

Leu Phe His His Val Tyr Gly Pro Leu Met Phe Glu Ser Tyr Lys Ala 165 170 175

Pro lie Pro Tyr Val Tyr Arg Ser Glu Ser Gly Asp Pro Asp Glu Cys 180 185 190

Arg Asp Gin Cys Leu Arg Glu Leu Ala Gin Leu Leu Glu Glu His His 195 200 205

Glu Glu lie Ala Ala Leu Ser lie Glu Ser Met Val Gin Gly Ala Ser 210 2X5 220

Gly Met lie Val Met Pro Glu Gly Tyr Leu Ala Gly Val Arg Glu Leu 225 230 235 240

Cys Thr Thr Tyr Asp Val Leu Met lie Val Asp Glu Val Ala Thr Gly 245 250 255

Phe Gly Arg Thr Gly Lys Met Phe Ala Cys Glu His Glu Asn Val Gin 260 265 270

Pro Asp Leu Met Ala Ala Gly Lys Gly lie Thr Gly Gly Tyr Leu Pro 275 280 285 lie Ala Val Thr Phe Ala Thr Glu Asp lie Tyr Lys Ala Phe Tyr Asp 290 295 300

Asp Tyr Glu Asn Leu Lys Thr Phe Phe His Gly His Ser Tyr Thr Gly 305 310 315 320

Asn Gin Leu Gly Cys Ala Val Ala Leu Glu Asn Leu Ala Leu Phe Glu 325 330 335

Ser Glu Asn lie Val Glu Gin Val Ala Glu Lys Ser Lys Lys Leu His 340 345 350

Phe Leu Leu Gin Asp Leu His Ala Leu Pro His Val Gly Asp lie Arg 355 360 365

Gin Leu Gly Phe Met Cys Gly Ala Glu Leu Val Arg Ser Lys Glu Thr 370 375 380 27 201127961

Lys Glu Pro Tyr Pro Ala Asp Arg Arg lie Gly Tyr Lys Val Ser Leu 385 390 395 400

Lys Met Arg Glu Leu Gly Met Leu Thr Arg Pro Leu Gly Asp Val lie 405 410 415

Ala Phe Leu Pro Pro Leu Ala Ser Thr Ala Glu Glu Leu Ser Glu Met 420 425 430

Val Ala lie Met Lys Gin Ala lie His Glu Val Thr Ser Leu Glu Asp 435 440 445 &lt;210&gt; 18 &lt;211&gt; 1467 &lt;212&gt; DNA &lt;213&gt; Rhodobacter sphaeroides &lt;220&gt;;221&gt; CDS &lt;222&gt; (1) . . (1467) &lt;400&gt; 18 atg ccc ggt tgc ggg ggc ttg ccc ggg aat gaa ccg aaa tgc gga cga Met Pro Gly Cys Gly Gly Leu Pro Gly Asn Glu Pro Lys Cys Gly Arg 1 5 10 15 gag ggg agg teg geg atg aeg egg aat gac geg aeg aat get gcc gga Glu Gly Arg Ser Ala Met Thr Arg Asn Asp Ala Thr Asn Ala Ala Gly 20 25 30 geg gtg ggc geg geg atg egg gat Cac ate etc ttg cct gca cag gaa Ala Val Gly Ala Ala Met Arg Asp His lie Leu Leu Pro Ala Gin Glu 35 40 45 atg geg aag etc ggc aag tcc geg cag ccg gtg ctg act cat gcc gag Met Ala Lys Leu Gly Lys Ser Ala Gin Pro Val Leu Thr His Ala Glu 50 55 60 ggc ate tat gtc cat acc gag gac ggc ege ege ctg ate gac ggg ccg Gly lie Tyr Val His Thr Glu Asp Gly Arg Arg Leu lie Asp Gly Pro 65 70 75 80 geg ggc Atg tgg tgc geg cag gtg ggc tac ggc ege ege gag ate gt c Ala Gly Met Trp Cys Ala Gin Val Gly Tyr Gly Arg Arg Glu lie Val 85 90 95 gat gcc atg geg cat cag geg atg gtg ctg ccc tat gcc teg ccc tgg Asp Ala Met Ala His Gin Ala Met Val Leu Pro Tyr Ala Ser Pro Trp 100 105 110 tat atg gcc aeg age ccc geg geg egg ctg geg gag aag ate gcc aeg Tyr Met Ala Thr Ser Pro Ala Ala Arg Leu Ala Glu Lys He Ala Thr 115 120 125 48 96 144 192 240 288 336 28 384 432201127961 Ctg acg ccg ggc gat etc aac egg ate ttt ttc acc aeg ggc ggg teg Leu Thr Pro Gly Asp Leu Asn Arg lie Phe Phe Thr Thr Gly Gly Ser 130 135 140 acc geg gtg gac age geg ctg ege ttc teg gaa ttc tac aac aac Ggg A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A Asp Gly Tyr 165 170 175 cac ggc teg acg geg etc acc gee gee tgc acc ggc ege acc ggc aac His Gly Ser Thr Ala Leu Thr Ala Ala Cys Thr Gly Arg Thr Gly Asn 180 185 190 tgg ccg aac ttc gac ate geg cag gacEgg ate teg ttc etc teg age Trp Pro Asn Phe Asp lie Ala Gin Asp Arg lie Ser Phe Leu Ser Ser 195 200 205 ccc aat ccg ege cac gee ggc aac ege age cag gag geg ttc etc gac Pro Asn Pro Arg His Ala Gly Asn Arg Ser Gin Glu Ala Phe Leu Asp 210 215 220 gat ctg gtg cag gaa ttc gag gac egg ate gag age etc ggc ccc gac Asp Leu Val Gin Glu Phe Glu Asp Arg lie Glu Ser Leu Gly Pro Asp 225 230 235 240 acg ate geg Gee ttc ctg gee gag ccg ate etc gee teg ggc ggc gtc Thr lie Ala Ala Phe Leu Ala Glu Pro lie Leu Ala Ser Gly Gly Val 245 250 255 att att ccg ccc gca ggc tat cat geg ege ttc aag geg ate tgc gag lie lie Pro Pro Ala Gly Tyr His Ala Arg Phe Lys Ala lie Cys Glu 260 265 270 aag cac gac ate etc tat ate teg gac gag gtg gtg acg ggc ttc ggc Lys His Asp lie Leu Tyr lie Ser Asp Glu Val Val Thr Gly Phe Gly 275 280 285 cgt tgc ggc gag tgg ttc gee teg gag aag gtg ttc ggg gtg gtg ccg Arg Cys Gly Glu Trp Phe Ala Ser Glu Lys Val Phe Gly Val Val Pro 290 295 300 gac ate ate acc ttc gee aag ggc gtg acc teg ggc tatGtg ccg etc Asp lie lie Thr Phe Ala Lys Gly Val Thr Ser Gly Tyr Val Pro Leu 305 310 315 320 ggc ggc ett geg ate tee gag geg gtg ctg geg egg ate teg ggc gag Gly Gly Leu Ala lie Ser Glu Ala Val Leu Ala Arg lie Ser Gly Glu 325 330 335 aat gee aag gga age tgg ttc acc aac ggc tat acc tac age aat cag Asn Ala Lys Gly Ser Trp Phe Thr Asn Gly Tyr Thr Tyr Ser Asn Gin 340 345 350 ccg gtg gee tgc gee geg geg Ett gee aac ate gag ctg atg gag ege Pro Val Ala Cys Ala Ala Ala Leu Ala Asn lie Glu Leu Met Glu Arg 355 360 365 480 528 576 624 720 768 816 864 912 960 1008 1056 29 s 1104 1152201127961 gag ggc atc 9tc 9at ca9 9C9 cgc 9a9 atg gcg gac tat ttc gcc gcg Glu Gly lie Val Asp Gin Ala Arg Glu Met Ala Asp Tyr Phe Ala Ala 370 375 380 gcg ctg get teg ctg ege gat ctg ccg ggc gtg gcg gaa acc egg teg Ala Leu Ala Ser Leu Arg Asp Leu Pro Gly Val Ala Glu Thr Arg Ser 385 390 395 400 gtg ggc etc gtg ggt tgc gtg caa tgc ctg etc gac ccg acc egg gcg Val Gly Leu Val Gly Cys Val Gin Cys Leu Leu Asp Pro Thr Arg Ala 40 5 410 415 gac ggc aeg gcc gag gac aag gcc ttc acc ctg aag ate gac gag ege Asp Gly Thr Ala Glu Asp Lys Ala Phe Thr Leu Lys lie Asp Glu Arg 420 425 430 tgc ttc gag etc ggg ctg ate gtg ege ccg ctg ggc Gat etc tgc gtg Cys Phe Glu Leu Gly Leu lie Val Arg Pro Leu Gly Asp Leu Cys Val 435 440 445 ate teg ccg ccg etc ate ate teg ege gcg cag ate gac gag atg gtc lie Ser Pro Pro Leu lie lie Ser Arg Ala Gin Lie Asp Glu Met Val 450 455 460 gcg ate atg egg cag gcc ate acc gaa gtg age gcc gcc cac ggt ctg Ala lie Met Arg Gin Ala lie Thr Glu Val Ser Ala Ala His Gly Leu 465 470 475 480 acc gcg aaa gaa ccg gee Gee gtc tga Thr Ala Lys Glu Pro Ala Ala Val 485 1200 1248 1296 1344 1392 1440 1467 &lt;210&gt; 19 &lt;211&gt; 488 &lt;212&gt; PRT &lt;213&gt; Rhodobacter sphaeroides &lt;400&gt;

Met Pro Gly Cys Gly Gly Leu Pro Gly Asn Glu Pro Lys Cys Gly Arg 15 10 15

Glu Gly Arg Ser Ala Met Thr Arg Asn Asp Ala Thr Asn Ala Ala Gly 20 25 30

Ala Val Gly Ala Ala Met Arg Asp His lie Leu Leu Pro Ala Gin Glu 35 40 45

Met Ala Lys Leu Gly Lys Ser Ala Gin Pro Val Leu Thr His Ala Glu 50 55 60

Gly lie Tyr Val His Thr Glu Asp Gly Arg Arg Leu lie Asp Gly Pro 65 70 75 80 30 201127961

Ala Gly Met Trp Cys Ala Gin Val Gly Tyr Gly Arg Arg Glu lie Val 85 90 95

Asp Ala Met Ala His Gin Ala Met Val Leu Pro Tyr Ala Ser Pro Trp 100 105 HO

Tyr Met Ala Thr Ser Pro Ala Ala Arg Leu Ala Glu Lys lie Ala Thr 115 120 125

Leu Thr Pro Gly Asp Leu Asn Arg lie Phe Phe Thr Thr Gly Gly Ser 130 135 140

Thr Ala Val Asp Ser Ala Leu Arg Phe Ser Glu Phe Tyr Asn Asn Val 145 150 155 160

Leu Gly Arg Pro Gin Lys Lys Arg lie lie Val Arg Tyr Asp Gly Tyr 165 170 175

His Gly Ser Thr Ala Leu Thr Ala Ala Cys Thr Gly Arg Thr Gly Asn 180 185 190

Trp Pro Asn Phe Asp lie Ala Gin Asp Arg lie Ser Phe Leu Ser Ser 195 200 205

Pro Asn Pro Arg His Ala Gly Asn Arg Ser Gin Glu Ala Phe Leu Asp 210 215 220

Asp Leu Val Gin Glu Phe Glu Asp Arg lie Glu Ser Leu Gly Pro Asp 225 230 235 240

Thr lie Ala Ala Phe Leu Ala Glu Pro lie Leu Ala Ser Gly Gly Val 245 250 255 lie lie Pro Pro Ala Gly Tyr His Ala Arg Phe Lys Ala lie Cys Glu 260 265 270

Lys His Asp lie Leu Tyr lie Ser Asp Glu Val Val Thr Gly Phe Gly 275 280 285

Arg Cys Gly Glu Trp Phe Ala Ser Glu Lys Val Phe Gly Val Val Pro 290 295 300

Asp He lie Thr Phe Ala Lys Gly Val Thr Ser Gly Tyr Val Pro Leu 31 £ 201127961 305 310 315 320

Gly Gly Leu Ala lie Ser Glu Ala Val Leu Ala Arg lie Ser Gly Glu 32S 330 335

Asn Ala Lys Gly Ser Trp Phe Thr Asn Gly Tyr Thr Tyr Ser Asn Gin 340 345 350

Pro Val Ala Cys Ala Ala Ala Leu Ala Asn lie Glu Leu Met Glu Arg 355 360 365

Glu Gly lie Val Asp Gin Ala Arg Glu Met Ala Asp Tyr Phe Ala Ala 370 375 380

Ala Leu Ala Ser Leu Arg Asp Leu Pro Gly Val Ala Glu Thr Arg Ser 385 390 395 400

Val Gly Leu Val Gly Cys Val Gin Cys Leu Leu Asp Pro Thr Arg Ala 405 410 415

Asp Gly Thr Ala Glu Asp Lys Ala Phe Thr Leu Lys lie Asp Glu Arg 420 425 430

Cys Phe Glu Leu Gly Leu lie Val Arg Pro Leu Gly Asp Leu Cys Val 435 440 445 lie Ser Pro Pro Leu lie lie Ser Arg Ala Gin lie Asp Glu Met Val 450 455 460

Ala lie Met Arg Gin Ala He Thr Glu Val Ser Ala Ala His Gly Leu 465 470 475 480

Thr Ala Lys Glu Pro Ala Ala Val 485 &lt;210&gt; 20 &lt;211&gt; 837

&lt;212&gt; DNA &lt;213&gt; Legionella pneumophila &lt;220&gt;

&lt;221&gt; CDS &lt;222&gt; (1)..(837) &lt;4〇〇&gt; 20 atg agt ate gca ttt gtt aac ggc aag tat tgt tgt caa tet gaa gca 32 201127961

Met Ser lie Ala Phe Val Asn Gly Lys Tyr Cys Cys Gin Ser Glu Ala 15 10 15 aaa att tea ata ttt gat ega ggg ttt ett ttt ggt gac teg gtt tat 96

Lys lie Ser lie Phe Asp Arg Gly Phe Leu Phe Gly Asp Ser Val Tyr 20 25 30 gaa gtg ctg cct gtt tac cat ggg cag cct tac ttt gta gac caa cat 144

Glu Val Leu Pro Val Tyr His Gly Gin Pro Tyr Phe Val Asp Gin His 35 40 45 ett gac ega tta ttc tea aat atg aaa aaa att aag atg att ata cca 192

Leu Asp Arg Leu Phe Ser Asn Met Lys Lys lie Lys Met lie lie Pro 50 55 60 aat tat gat tgg cat ggt tta att cat aga eta ata tea gaa aat aat 240

Asn Tyr Asp Trp His Gly Leu lie His Arg Leu lie Ser Glu Asn Asn 65 70 75 80 ggc ggt aat tta caa gta tat ate caa gtc aca ega ggg aat caa ggg 288

Gly Gly Asn Leu Gin Val Tyr lie Gin Val Thr Arg Gly Asn Gin Gly 85 90 95 gtg ege aag cat gat ate cct act tcc ate aca cct tet gtt ate gca 336

Val Arg Lys His Asp lie Pro Thr Ser lie Thr Pro Ser Val lie Ala 100 105 110 ttc act atg cat aat cca ttt ccc acc etc gaa gat aag gaa cag gga 384

Phe Thr Met His Asn Pro Phe Pro Thr Leu Glu Asp Lys Glu Gin Gly 11B 120 125 atg tea gca aaa ctg gtt gaa gat ttt egg tgg atg aga tgt gat ata 432

Met Ser Ala Lys Leu Val Glu Asp Phe Arg Trp Met Arg Cys Asp lie 130 135 140 aaa act act tet tta att gcc aat ata tta ctg aat gat gag get gta 480

Lys Thr Thr Ser Leu lie Ala Asn lie Leu Leu Asn Asp Glu Ala Val 145 150 155 160 tet gca gga ttc cac act gca att ett gcc egg aac ggt eta att aca 528

Ser Ala Gly Phe His Thr Ala lie Leu Ala Arg Asn Gly Leu lie Thr 165 170 175 gag gga agt agt acc aac gta ttt att gtc gca cag gat ggt gtt att 576

Glu Gly Ser Ser Thr Asn Val Phe lie Val Ala Gin Asp Gly Val lie 180 185 190 aag aca cca ccc atg aat aat ttc tgt tta cca gga att act egg caa 624

Lys Thr Pro Pro Met Asn Asn Phe Cys Leu Pro Gly lie Thr Arg Gin 195 200 205 gtt gtt att gaa ata att aaa aaa tta gat tta aag ttc aga gaa ata 672

Val Val lie Glu lie lie Lys Lys Leu Asp Leu Lys Phe Arg Glu lie 210 215 220 gaa att age att tea gag ett ttt tet get cag gaa gtt tgg ata aca 720

Glu lie Ser lie Ser Glu Leu Phe Ser Ala Gin Glu Val Trp lie Thr 225 230 235 240 33 &amp; 768 201127961 agt acg aca aaa gaa gta ttc cct att aca aag att aat gac tct ttg

Ser Thr Thr Lys Glu Val Phe Pro lie Thr Lys lie Asn Asp Ser Leu 245 250 255 att aat ggc gga aaa gtt ggc gaa tat tgg egg ata att aat gat tcc lie Asn Gly Gly Lys Val Gly Glu Tyr Trp Arg lie lie Asn Asp Ser 260 265 270 tac caa caa eta gta aac taa Tyr Gin Gin Leu Val Asn 275 &lt;210&gt; 21 &lt;211&gt; 278

&lt;212&gt; PRT &lt;213&gt; Legionella pneumophila &lt;400&gt; 21 Met Ser lie Ala Phe Val Asn Gly Lys Tyr Cys Cys Gin Ser Glu Ala 1 5 10 15 Lys lie Ser lie Phe Asp Arg Gly Phe Leu Phe Gly Asp Ser Val Tyr 20 25 30

Glu Val Leu Pro Val Tyr His Gly Gin Pro Tyr Phe Val Asp Gin His 35 40 45

Leu Asp Arg Leu Phe Ser Asn Met Lys Lys lie Lys Met lie lie Pro 50 55 60

Asn Tyr Asp Trp His Gly Leu lie His Arg Leu lie Ser Glu Asn Asn 65 70 75 80

Gly Gly Asn Leu Gin Val Tyr lie Gin Val Thr Arg Gly Asn Gin Gly 85 90 95

Val Arg Lys His Asp lie Pro Thr Ser lie Thr Pro Ser Val lie Ala 100 105 110

Phe Thr Met His As Pro Pro Phe Pro Thr Leu Glu Asp Lys Glu Gin Gly 115 120 125

Met Ser Ala Lys Leu Val Glu Asp Phe Arg Trp Met Arg Cys Asp lie 130 135 140

Lys Thr Thr Ser Leu lie Ala Asn He Leu Leu Asn Asp Glu Ala Val 145 150 155 160 816 837 34 201127961

Ser Ala Gly Phe His Thr Ala lie Leu Ala Arg Asn Gly Leu lie Thr 165 170 175

Glu Gly Ser Ser Thr Asn Val Phe He Val Ala Gin Asp Gly Val lie 180 185 19〇

Lys Thr Pro Pro Met Asn Asn Phe Cys Leu Pro Gly lie Thr Arg Gin 195 200 205

Val Val lie Glu lie lie Lys Lys Leu Asp Leu Lys Phe Arg Glu lie 210 215 220

Glu lie Ser lie Ser Glu Leu Phe Ser Ala Gin Glu Val Trp lie Thr 225 230 235 240

Ser Thr Thr Lys Glu Val Phe Pro lie Thr Lys lie Asn Asp Ser Leu 245 250 255 lie Asn Gly Gly Lys Val Gly Glu Tyr Trp Arg lie lie Asn Asp Ser 260 265 270

Tyr Gin Gin Leu Val Asn 275 &lt;210&gt; 22 &lt;211&gt; 861 &lt;212&gt; DNA &lt;213&gt; European genus Nitrosomonas europaea &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) . . . (861) &lt;400&gt; 22 atg att tac etc aat ggc aaa ttt ctg ccg atg gaa cag get acc gtt Met lie Tyr Leu Asn Gly Lys Phe Leu Pro Met Glu Gin Ala Thr Val 1 5 10 15 Cca gtg ctg gat aga ggc ttc ate ttc ggt gat ggt gtc tat gaa gtc Pro Val Leu Asp Arg Gly Phe lie Phe Gly Asp Gly Val Tyr Glu Val 20 25 30 ata ccg gtt tat tea cgt eiaa ccg ttc egg ctg ggc gaa cat ett Tcc lie Pro Val Tyr Ser Arg Lys Pro Phe Arg Leu Gly Glu His Leu Ser 35 40 45 egg ctg cag cac agt ctg gat ggc ata cgt etc cag aat ccg cac act Arg Leu Gin His Ser Leu Asp Gly lie Arg Leu Gin Asn Pro His Thr 5〇55 60 48 96 144 35 192 240201127961 gaa gaa caa tgg get ggt ctg ate gaa ege ate ate gag ctg aat gaa Glu Glu Gin Trp Ala Gly Leu lie Glu Arg lie lie Glu Leu Asn Glu 65 70 75 80 ggt gat Gat cag tac ett tac ctg cac att aca ege ggg gtg gca aaa Gly A Sp Asp Gin Tyr Leu Tyr Leu His lie Thr Arg Gly Val Ala Lys 85 90 95 cgt gac cat gee ttt cct ege gaa gta aeg ccc act gtc ttc ate atg Arg Asp His Ala Phe Pro Arg Glu Val Thr Pro Thr Val Phe lie Met 100 105 110 age aac ccg ett ccg get cca cct gca aaa ttg etc gtt tee gga gtt Ser Asn Pro Leu Pro Ala Pro Pro Ala Lys Leu Leu Val Ser Gly Val 115 120 125 tea geg att acc gee agg gat aat ege tgg ggg ege Tgt gat ate aaa A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A Gin Leu Ala Val Asp 145 150 155 160 gca caa gee atg gaa aeg ate ctg tta ege gat ggt ctg ttg acc gaa Ala Gin Ala Met Glu Thr lie Leu Leu Arg Asp Gly Leu Leu Thr Glu 165 170 175 ggg gee gee age aat att Ttc ate gta aaa gac gac ctg ctg ctg acc Gly Ala Ala Ser Asn lie Phe lie Val Lys Asp Asp Leu Leu Leu Thr 180 185 190 ccc ccc aaa gat cac cgt ata ttg cct ggc att act tat gat gta gta Pro Pro Lys Asp His Arg Lie Leu Pro Gly lie Thr Tyr Asp Val Val 195 200 205 ctg gaa ctg get gaa aca cat ggt gtt cca cat geg aca aga gaa ata Leu Glu Leu Ala Glu Thr His Gly Val Pro His Ala Thr Arg Glu lie 210 215 220 tea gag Tett gag tta cgt act gca egg gaa ate atg ctg act tet tee Ser Glu Leu Glu Leu Arg Thr Ala Arg Glu lie Met Leu Thr Ser Ser 225 230 235 240 acc aaa gaa att etc ccg ate aca cag ctg gat gga caa ccg ate ggt Thr Lys Glu lie Leu Pro lie Thr Gin Leu Asp Gly Gin Pro lie Gly 24S 250 255 aat ggc acc cca ggg cca gta ttt cag caa ctg gat egg etc tat cag Asn Gly Thr Pro Gly Pro Val Phe Gin Gin Leu Asp Arg Leu Tyr Gin 260 265 270 gca tat aag ctg gaa gtc atg ege ggg cat get cca ege cag taa Ala Tyr Lys Leu Glu Val Met Arg Gly His Ala Pro Arg Gin 275 280 285 288 336 432 480 528 576 624 672 720 768 816 &lt;210&gt; 23 36 861 201127961

&lt;211&gt; 286 &lt;212&gt; PRT &lt;213&gt; European Nitrosomonas europaea &lt;400&gt; 23

Met lie Tyr Leu Asn Gly Lys Phe Leu Pro Met Glu Gin Ala Thr Val 1 5 10 15

Pro Val Leu Asp Arg Gly Phe lie Phe Gly Asp Gly Val Tyr Glu Val 20 25 30 lie Pro Val Tyr Ser Arg Lys Pro Phe Arg Leu Gly Glu His Leu Ser 35 40 45

Arg Leu Gin His Ser Leu Asp Gly lie Arg Leu Gin Asn Pro His Thr 50 55 60

Glu Glu Gin Trp Ala Gly Leu lie Glu Arg He He Glu Leu Asn Glu 65 70 75 80

Gly Asp Asp Gin Tyr Leu Tyr Leu His He Thr Arg Gly Val Ala Lys 85 90 95

Arg Asp His Ala Phe Pro Arg Glu Val Thr Pro Thr Val Phe lie Met 100 105 110

Ser Asn Pro Leu Pro Ala Pro Pro Ala Lys Leu Leu Val Ser Gly Val 115 120 125

Ser Ala lie Thr Ala Arg Asp Asn Arg Trp Gly Arg Cys Asp lie Lys 130 135 140

Ala lie Ser Leu Leu Pro Asn lie Leu Leu Arg Gin Leu Ala Val Asp 145 150 155 160

Ala Gin Ala Met Glu Thr lie Leu Leu Arg Asp Gly Leu Leu Thr Glu 165 170 175

Gly Ala Ala Ser Asn lie Phe lie Val Lys Asp Asp Leu Leu Leu Thr 180 185 190

Pro Pro Lys Asp His Arg lie Leu Pro Gly lie Thr Tyr Asp Val Val 195 200 205

Leu Glu Leu Ala Glu Thr His Gly Val Pro His Ala Thr Arg Glu lie 37 201127961 210 215 220

Ser Glu Leu Glu Leu Arg Thr Ala Arg Glu lie Met Leu Thr Ser Ser 225 230 235 240

Thr Lys Glu lie Leu Pro lie Thr Gin Leu Asp Gly Gin Pro lie Gly 245 250 255

Asn Gly Thr Pro Gly Pro Val Phe Gin Gin Leu Asp Arg Leu Tyr Gin 260 265 270

Ala Tyr Lys Leu Glu Val Met Arg Gly His Ala Pro Arg Gin 275 280 285 &lt;210&gt; 24 &lt;211&gt; 1293 &lt;212&gt; DNA &lt;213&gt; Neisseria gonorrhoeae &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1)..(1293) &lt;400&gt; 24 atg agg ata aat atg aac cgt aac gaa att tta ttc gac cgc gcc aag Met Arg lie Asn Met Asn Arg Asn Glu lie Leu Phe Asp Arg Ala Lys 15 10 15 gcc ate ate ccc ggc ggc gtg aat teg ccc gtg cgc gca ttc ggc age Ala lie lie Pro Gly Gly Val Asn Ser Pro Val Arg Ala Phe Gly Ser 20 25 30 gtc ggc ggc gtg ccg cgc ttc ate aaa aaa gcc Gaa ggc geg tat gtt Val Gly Gly Val Pro Arg Phe lie Lys Lys Ala Glu Gly Ala Tyr Val 35 40 45 tgg gac gaa aac ggc aeg cgc tac acc gat tat gtc ggc tet tgg ggg Trp Asp Glu Asn Gly Thr Arg Tyr Thr Asp Tyr Val Gly Ser Trp Gly 50 55 60 cct geg att gtc gga cac geg cat ccc gaa gtc gtc gaa gcc gtg cgc Pro Ala He Val Gly His Ala His Pro Glu Val Val Glu Ala Val Arg 65 70 75 80 gaa get geg ttg ggc Ggt ttg teg ttc ggc geg ccc acc gaa ggc gaa Glu Ala Ala L Eu Gly Gly Leu Ser Phe Gly Ala Pro Thr Glu Gly Glu 85 90 95 ate gcc att gcc gaa caa att gcc gaa att atg ccg tet gtc gaa egg lie Ala He Ala Glu Gin He Ala Glu He Met Pro Ser Val Glu Arg 100 105 110 48 96 144 192 240 288 336 ctg cgc etc gtc age tcc ggc aeg gaa geg aeg atg act gcc ate cgt 384 38 201127961

Gu gu gu gu gu gu gu gu

Leu Ala Arg Gly Phe Thr Gly Arg Asp Lys lie lie Lys Phe Glu Gly 130 135 140 tgc tac cac ggc cat tcc gac age ctg ttg gtg aaa gca ggc age ggt 480

Cys Tyr His Gly His Ser Asp Ser Leu Leu Val Lys Ala Gly Ser Gly 145 150 155 160 ctg ett acc ttc ggc aat cct tet tcc gcc ggt gtg cct gcc gac ttt 528

Leu Leu Thr Phe Gly Asn Pro Ser Ser Ala Gly Val Pro Ala Asp Phe 165 170 175 acc aaa cat act ttg gta etc gaa tac aac aac ate gcc caa etc gaa 576

Thr Lys His Thr Leu Val Leu Glu Tyr Asn Asn lie Ala Gin Leu Glu 180 185 190 gaa gcc ttt gcc caa age ggc gac gaa ate gcc tgc gtg att gtc gaa 624

Glu Ala Phe Ala Gin Ser Gly Asp Glu lie Ala Cys Val lie Val Glu 195 200 205 ccc ttc gtc ggc aat atg aac etc gtc cgc ccg acc gaa gcc ttt gtc 672

Pro Phe Val Gly Asn Met Asn Leu Val Arg Pro Thr Glu Ala Phe Val 210 215 220 aaa gcc ttg ege gga ttg acc gaa aaa cac ggc geg gtg ttg att tac 720

Lys Ala Leu Arg Gly Leu Thr Glu Lys His Gly Ala Val Leu lie Tyr 225 230 235 240 gac gaa gtg atg acc ggt ttc cgc gtc geg etc ggc ggc geg cag teg 768

Asp Glu Val Met Thr Gly Phe Arg Val Ala Leu Gly Gly Ala Gin Ser 245 250 255 ctg cac ggc ate aeg ccc gac ctg acc aeg atg ggc aaa gtc ate ggc 816

Leu His Gly lie Thr Pro Asp Leu Thr Thr Met Gly Lys Val lie Gly 260 265 270 ggc ggt atg ccg ett gcc geg ttc ggc gga cgc aaa gac ate atg gaa 864

Gly Gly Met Pro Leu Ala Ala Phe Gly Gly Arg Lys Asp lie Met Glu 275 280 285 tgt att tcc ccg ttg ggc ggc gtg tat cag gca ggt aca tta tea ggc 912

Cys lie Ser Pro Leu Gly Gly Val Tyr Gin Ala Gly Thr Leu Ser Gly 290 295 300 aac ccg att gcc gtc gcc gcc ggc ttg aaa aeg ctg gaa ate ate cag 960

Asn Pro lie Ala Val Ala Ala Gly Leu Lys Thr Leu Glu lie lie Gin 305 310 315 320 cgc gaa ggc ttc tat gaa aac ctg acc gcc ttg aca caa cgc ett gcc 1008

Arg Glu Gly Phe Tyr Glu Asn Leu Thr Ala Leu Thr Gin Arg Leu Ala 325 330 335 aac ggt att gcc gcc gcc aaa geg cac ggt ate gag ttt gcc gcc gac 1056

Asn Gly lie Ala Ala Ala Lys Ala His Gly lie Glu Phe Ala Ala Asp 340 345 350 39 1104201127961 age gtg ggc ggt atg ttc ggt ctg tat ttc gee gca cac gtg ccg ega Ser Val Gly Gly Met Phe Gly Leu Tyr Phe Ala Ala His Val Pro Arg 355 360 365 aac tat gee gat atg geg ege tee aat ate gac get ttc aaa ege ttc Asn Tyr Ala Asp Met Ala Arg Ser Asn lie Asp Ala Phe Lys Arg Phe 370 375 380 ttc cac ggc atg etc gac ege ggc att Gee ttc ggc ccg tee get tat Phe His Gly Met Leu Asp Arg Gly lie Ala Phe Gly Pro Ser Ala Tyr 385 390 395 400 gaa geg ggt ttc gtt tee gee geg cat aeg ccc gag ctg att gac gaa Glu Ala Gly Phe Val Ser Ala Ala His Thr Pro Glu Leu lie Asp Glu 405 410 415 aeg gtt geg gtt geg gtt gaa gtg ttc aag geg atg get gca tga Thr Val Ala Val Ala Val Glu Val Phe Lys Ala Met Ala Ala 420 425 430 1152 1200 1248 1293 &lt;210&gt; 25 &lt;211&gt; 430 &lt;212&gt; PRT &lt;213&gt; Neisseria gonorrhoeae &lt;400&gt; 25 Met Arg lie Asn Met Asn Arg Asn Glu lie Leu Phe Asp Arg Ala Lys 1 5 10 15 Ala lie lie Pro Gly Gly Val Asn Ser Pro Val Arg Ala Phe Gly Ser 20 25 30 Val Gly Gly Val Pro Arg Phe lie Lys Lys Ala Glu Gly Ala Tyr Val 35 40 45

Trp Asp Glu Asn Gly Thr Arg Tyr Thr Asp Tyr Val Gly Ser Trp Gly 50 55 60

Pro Ala lie Val Gly His Ala His Pro Glu Val Val Glu Ala Val Arg 65 70 75 80

Glu Ala Ala Leu Gly Gly Leu Ser Phe Gly Ala Pro Thr Glu Gly Glu 85 90 95 lie Ala lie Ala Glu Gin lie Ala Glu lie Met Pro Ser Val Glu Arg 100 105 110

Leu Arg Leu Val Ser Ser Gly Thr Glu Ala Thr Met Thr Ala lie Arg 115 120 125 40 201127961

Leu Ala Arg Gly Phe Thr Gly Arg Asp Lys lie lie Lys Phe Glu Gly 130 135 140

Cys Tyr His Gly His Ser Asp Ser Leu Leu Val Lys Ala Gly Ser Gly 145 150 155 160

Leu Leu Thr Phe Gly Asn Pro Ser Ser Ala Gly Val Pro Ala Asp Phe 165 170 175

Thr Lys His Thr Leu Val Leu Glu Tyr Asn Asn lie Ala Gin Leu Glu 180 185 190

Glu Ala Phe Ala Gin Ser Gly Asp Glu lie Ala Cys Val lie Val Glu 195 200 205

Pro Phe Val Gly Asn Met Asn Leu Val Arg Pro Thr Glu Ala Phe Val 210 215 220

Lys Ala Leu Arg Gly Leu Thr Glu Lys His Gly Ala Val Leu lie Tyr 225 230 235 240

Asp Glu Val Met Thr Gly Phe Arg Val Ala Leu Gly Gly Ala Gin Ser 245 250 255

Leu His Gly lie Thr Pro Asp Leu Thr Thr Met Gly Lys Val lie Gly 260 265 270

Gly Gly Met Pro Leu Ala Ala Phe Gly Gly Arg Lys Asp lie Met Glu 275 280 285

Cys lie Ser Pro Leu Gly Gly Val Tyr Gin Ala Gly Thr Leu Ser Gly 290 295 300

Asn Pro lie Ala Val Ala Ala Gly Leu Lys Thr Leu Glu lie lie Gin 305 310 315 320

Arg Glu Gly Phe Tyr Glu Asn Leu Thr Ala Leu Thr Gin Arg Leu Ala 325 330 335

Asn Gly lie Ala Ala Ala Lys Ala His Gly lie Glu Phe Ala Ala Asp 340 345 350

Ser Val Gly Gly Met Phe Gly Leu Tyr Phe Ala Ala His Val Pro Arg 355 360 365 41 201127961

Asn Tyr Ala Asp Met Ala Arg Ser Asn lie Asp Ala Phe Lys Arg Phe 370 375 380

Phe His Gly Met Leu Asp Arg Gly lie Ala Phe Gly Pro Ser Ala Tyr 385 390 395 400

Glu Ala Gly Phe Val Ser Ala Ala His Thr Pro Glu Leu lie Asp Glu 405 410 415

Thr Val Ala Val Ala Val Glu Val Phe Lys Ala Met Ala Ala 420 425 430 &lt;210&gt; 26 &lt;211&gt; 924 &lt;212&gt; DNA &lt;213&gt; Pseudomonas aeruginosa} &lt;220&gt;&lt;;221&gt; CDS &lt;222&gt; (1) . . (924) &lt;400&gt; 26 atg teg atg gcc gat cgt gat ggc gtg ate tgg tat gac ggt gaa ctg Met Ser Met Ala Asp Arg Asp Gly Val lie Trp Tyr Asp Gly Glu Leu 15 10 15 gtg cag tgg ege gac geg acc aeg cac gtg ctg acc cat acc ctg cac Val Gin Trp Arg Asp Ala Thr Thr His Val Leu Thr His Thr Leu His 20 25 30 tat gga atg ggc gtg ttc gag ggc gtg Ege gcc tac gac acc ccg cag Tyr Gly Met Gly Val Phe Glu Gly Val Arg Ala Tyr Asp Thr Pro Gin 3S 40 45 ggc aeg geg ate ttc ege ctg cag geg cat acc gac egg ctg ttc gac Gly Thr Ala lie Phe Arg Leu Gin Ala His Thr Asp Arg Leu Phe Asp 50 55 60 tcc geg cac ate atg aac atg cag ate ccg tac age ege gac gag ate Ser Ala His lie Met Asn Met Gin lie Pro Tyr Ser Arg Asp Glu lie 65 70 75 80 aac gag geg Acc ege gcc gcc gtg ege gag aac aac ctg gaa age gcc Asn Glu Ala Thr Arg Ala Ala Val Arg Glu Asn Asn Leu Glu Ser Ala 85 90 95 tat ate ege ccg atg gtg ttc tac gga age gaa ggc atg ggc ctg ege Tyr lie Arg Pro Met Val Phe Tyr Gly Ser Glu Gly Met Gly Leu Arg 100 105 110 gcc age ggc ctg aag gtc cat gtg ate ate gcc gcc tgg age tgg ggc Ala Ser Gly Leu Lys Val His Val lie lie Ala Ala Trp Ser Trp Gly 48 96 144 192 240 288 336 42 384 201127961 115 120 125 gcc tac atg Ggc gag gaa gcc ctg cag caa ggc ate aag gtg ege acc 432

Ala Tyr Met Gly Glu Glu Ala Leu Gin Gin Gly ly Lys Val Arg Thr 130 135 140 agt tee ttc acc ege cac cac gtc aac ate teg atg acc ege gcc aag 480

Ser Ser Phe Thr Arg His His Val Asn lie Ser Met Thr Arg Ala Lys 145 150 155 160 tee aac ggc gcc tac ate aac teg atg ctg gcc etc cag gaa geg ate 528

Ser Asn Gly Ala Tyr He Asn Ser Met Leu Ala Leu Gin Glu Ala lie 165 170 175 tee ggc ggc gcc gac gag gcc atg at g etc ccg gaa ggc tac gtg 576

Ser Gly Gly Ala Asp Glu Ala Met Met Leu Asp Pro Glu Gly Tyr Val 180 185 190 gcc gaa ggc tee ggc gag aac ate ttc ate ate aag gat ggc gtg ate 624

Ala Glu Gly Ser Gly Glu Asn lie Phe lie lie Lys Asp Gly Val lie 195 200 205 tac acc ccg gaa gtc acc gcc tgc ctg aac ggc ate act cgt aac act 672

Tyr Thr Pro Glu Val Thr Ala Cys Leu Asn Gly lie Thr Arg Asn Thr 210 215 220 ate ctg acc ctg gcc gcc gaa cac ggt ttt aaa ctg gtc gag aag ege 720 lie Leu Thr Leu Ala Ala Glu His Gly Phe Lys Leu Val Glu Lys Arg 225 230 235 240 ate acc ege gac gag gtg tac ate gcc gac gag gcc ttc ttc act ggc 768 lie Thr Arg Asp Glu Val Tyr lie Ala Asp Glu Ala Phe Phe Thr Gly 245 250 255 act gcc geg gaa gtc aeg ccg ate Ege gaa gtg gac ggt ege aag ate 816

Thr Ala Ala Glu Val Thr Pro lie Arg Glu Val Asp Gly Arg Lys lie 260 265 270 ggc gcc ggc ege cgt ggc ccg gtc acc gaa aag ctg cag aaa gcc tat 864

Gly Ala Gly Arg Arg Gly Pro Val Thr Glu Lys Leu Gin Lys Ala Tyr 275 280 285 ttc gac ctg gtc age ggc aag acc gag gcc cac gcc gag tgg cgt acc 912

Phe Asp Leu Val Ser Gly Lys Thr Glu Ala His Ala Glu Trp Arg Thr 290 295 300 ctg gtc aag taa 924

Leu Val Lys 305 &lt;210&gt; 27 &lt;211&gt; 307

&lt;212&gt; PRT &lt;213&gt; Pseudomonas aeruginosa &lt;400&gt; 27

Met Ser Met Ala Asp Arg Asp Gly Val lie Trp Tyr Asp Gly Glu Leu

S 43 201127961 1 5 10 15

Val Gin Trp Arg Asp Ala Thr Thr His Val Leu Thr His Thr Leu His 20 25 30

Tyr Gly Met Gly Val Phe Glu Gly Val Arg Ala Tyr Asp Thr Pro Gin 35 40 45

Gly Thr Ala lie Phe Arg Leu Gin Ala His Thr Asp Arg Leu Phe Asp 50 55 60

Ser Ala His lie Met Asn Met Gin lie Pro Tyr Ser Arg Asp Glu lie 65 70 75 80

Asn Glu Ala Thr Arg Ala Ala Val Arg Glu Asn Asn Leu Glu Ser Ala 85 90 95

Tyr lie Arg Pro Met Val Phe Tyr Gly Ser Glu Gly Met Gly Leu Arg 100 105 110

Ala Ser Gly Leu Lys Val His Val lie lie Ala Ala Trp Ser Trp Gly 115 120 125

Ala Tyr Met Gly Glu Glu Ala Leu Gin Gin Gly lie Lys Val Arg Thr 130 135 140

Ser Ser Phe Thr Arg His His Val Asn lie Ser Met Thr Arg Ala Lys 145 150 155 160

Ser Asn Gly Ala Tyr lie Asn Ser Met Leu Ala Leu Gin Glu Ala lie 165 170 175

Ser Gly Gly Ala Asp Glu Ala Met Met Leu Asp Pro Glu Gly Tyr Val 180 185 190

Ala Glu Gly Ser Gly Glu Asn lie Phe lie lie Lys Asp Gly Val lie 195 200 205

Tyr Thr Pro Glu Val Thr Ala Cys Leu Asn Gly lie Thr Arg Asn Thr 210 215 220 lie Leu Thr Leu Ala Ala Glu His Gly Phe Lys Leu Val Glu Lys Arg 225 230 235 240 44 201127961

He Thr Arg Asp Glu Val Tyr He Ala Asp Glu Ala Phe Phe Thr Gly 245 250 255

Thr Ala Ala Glu Val Thr Pro He Arg Glu Val Asp Gly Arq Lys He 260 265 270 Gly Ala Gly Arg Arg Gly Pro Val Thr Glu Lys Leu Gin Lys Ala Tyr 275 280 285

Phe Asp Leu Val Ser Gly Lys Thr Glu Ala His Ala Glu Trp Arg Thr 290 295 300

Leu Val Lys 305 &lt;210&gt; 28 &lt;211&gt; 1407 &lt;212&gt; DNA &lt;213&gt;: Rhodopseudomonas palustris &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) (1407) &lt;400&gt; 28 atg aag ctg ata ccg tgc cgc gcc ttt cac ccc ccg gcc gcg cag tgc Met Lys Leu He Pro Cys Arg Ala Phe His Pro Pro Ala Ala Gin Cys 1 5 10 15 atg agg age gcc Atg tta gac aag ate aag ccc aeg tcc gcc gtc aac Met Arg Ser Ala Met Leu Asp Lys lie Lys Pro Thr Ser Ala Val Asn 20 25 30 gcg ccg aac gat etc aac gcg ttc tgg atg ccg ttc acc gcg aac egg Ala Pro Asn Asp Leu Asn Ala Phe Trp Met Pro Phe Thr Ala Asn Arg 35 40 45 gcc ttc aag cgc gcg ccg aag atg gtc gtg ggt gcc gaa ggc atg cac Ala Phe Lys Arg Ala Pro Lys Met Val Val Gly Ala Glu Gly Met His 50 55 60 tac ate acc gcc gat ggt cgc ag ate ate gac gcc gcc teg ggc atg Tyr lie Thr Ala Asp Gly Arg Lys lie lie Asp Ala Ala Ser Gly Met 65 70 75 80 tgg tgc acc aat gcg ggc cat ggc cgc aag gaa ate gcc Gag gcg ate Trp Cys Thr Asn Ala Gly His Gly Arg Lys Glu lie Ala Glu Ala lie 85 90 95 aag gcg cag gcc gat gaa etc gac ttc teg ccg ccg ttc cag ttc ggc Lys Ala Gin Ala Asp Glu Leu Asp Phe Ser Pro Pro Phe Gin Phe Gly 100 105 110 48 96 144 192 240 288 45 336 201127961 Cag ccg aag gcg ttc gaa etc gee age egg ate gee gat ctg geg ccg 334

Gin Pro Lys Ala Phe Glu Leu Ala Ser Arg lie Ala Asp Leu Ala Pro 115 120 125 gaa ggc etc gat cac gtg ttc ttc tgc aat teg ggc teg gaa gee ggc 432

Glu Gly Leu Asp His Val Phe Phe Cys Asn Ser Gly Ser Glu Ala Gly 130 135 140 gac acc gcg ctg aag ate gcg gtc gee tat cag cag ate aag ggc cag 48〇

Asp Thr Ala Leu Lys lie Ala Val Ala Tyr Gin Gin lie Lys Gly Gin 145 150 155 160 ggc tea ege acc ege ctg ate ggc ege gag ege ggc tat cac ggc gtc 52a

Gly Ser Arg Thr Arg Leu lie Gly Arg Glu Arg Gly Tyr His Gly Val 165 170 175 ggc ttc ggc ggc acc gcg gtc ggc ggc ate ggc aac aac ege aag atg 575

Gly Phe Gly Gly Thr Ala Val Gly Gly lie Gly Asn Asn Arg Lys Met 180 185 190 ttc ggt ccg ctg etc aac ggc gtc gat cat ctg cct gcg act tat gat 624

Phe Gly Pro Leu Leu Asn Gly Val Asp His Leu Pro Ala Thr Tyr Asp 195 200 205 ege gac aag cag get ttc acc ate ggc gag ccg gaa tac ggc gcg cac 672

Arg Asp Lys Gin Ala Phe Thr lie Gly Glu Pro Glu Tyr Gly Ala His 210 215 220 ttc gee gaa gcg ett gaa ggc etc gtc aat ctg.cac ggc gee aac acc 720

Phe Ala Glu Ala Leu Glu Gly Leu Val Asn Leu His Gly Ala Asn Thr 225 230 235 240 ate gcg gcg gtg ate gtc gag ccg atg gee ggc tee acc ggc gtg ctg 768 lie Ala Ala Val lie Val Glu Pro Met Ala Gly Ser Thr Gly Val Leu 245 250 255 ccg gcg ccg aag ggc tat etc aag aag ctg ege gag ate acc aag aag 816

Pro Ala Pro Lys Gly Tyr Leu Lys Lys Leu Arg Glu lie Thr Lys Lys 260 265 270 cac ggc ate ctg ctg ate ttc gac gag gtc ate acc ggc tac ggc cgt 864

His Gly lie Leu Leu lie Phe Asp Glu Val lie Thr Gly Tyr Gly Arg 275 280 285 etc ggc tat gee ttc gcg tee gaa cgt tac ggc gtc acc ccg gac atg 912

Leu Gly Tyr Ala Phe Ala Ser Glu Arg Tyr Gly Val Thr Pro Asp Met 290 295 300 ate acc ttc gee aag ggc gtc acc aat ggt gcg gtg ccg atg ggc ggc 960 lie Thr Phe Ala Lys Gly Val Thr Asn Gly Ala Val Pro Met Gly Gly 305 310 315 320 gtg ate acc teg gcg gag ate cac gat gcg ttc atg acc ggc ccc gag 1008

Val He Thr Ser Ala Glu lie His Asp Ala Phe Met Thr Gly Pro Glu 325 330 335 cac gcg gtc gag ctg gcg cac ggc tac acc tat teg gcg cat ccg etc 1056

His Ala Val Glu Leu Ala His Gly Tyr Thr Tyr Ser Ala His Pro Leu 340 345 350 46 1104201127961 gcc tgc gcg gcc ggc ate gee acc etc gac ate tac ege gac gag aag Ala Cys Ala Ala Gly lie Ala Thr Leu Asp lie Tyr Arg Asp Glu Lys 355 360 365 ctg ttc gag ege gcc aag gcg ctg gag ccg aag ttt gcc gag gcg gtg Leu Phe Glu Arg Ala Lys Ala Leu Glu Pro Lys Phe Ala Glu Ala Val 370 375 380 atg teg ctg aag teg gcc ccg aac gtg Gtc gac ate ege acc gtc ggc Met Ser Leu Lys Ser Ala Pro Asn Val Val Asp lie Arg Thr Val Gly 385 390 395 400 ctg aeg gcg ggt ate gac etc get teg ate gcc gat gcg gtc ggc aag Leu Thr Ala Gly lie Asp Leu Ala Ser lie Ala Asp Ala Val Gly Lys 405 410 415 cgt ggc ttc gaa gcg atg aat gcc ggc ttc cac gac cac gag ctg atg Arg Gly Phe Glu Ala Met Asn Ala Gly Phe His Asp His Glu Leu Met 420 425 430 ctg egg ate Gcc ggc gac acc ctg gcg ctg acc ccg ccg ctg ate etc Leu Arg lie Ala Gly Asp Thr Leu Ala Leu Thr Pro Pro Leu lie Leu 435 440 445 age gag gac cac ate ggt gag ate gtc gac aag gtc ggc aag gtg ate Ser Glu Asp His lie Gly Glu lie Val Asp Lys Val Gly Lys Val lie 450 455 460 ege gcg gtc gcc tga Arg Ala Val Ala 465 1152 1200 1248 1296 1344 1392 1407 &lt;210&gt; 29 &lt;211&gt; 468 &lt;212&gt; PRT &lt;213&gt; Rhodopseudomonas palustris &lt;400&gt; 29

Met Lys Leu lie Pro Cys Arg Ala Phe His Pro Pro Ala Ala Gin Cys 1 5 10 15

Met Arg Ser Ala Met Leu Asp Lys lie Lys 20 25

Pro Thr Ser Ala Val Asn 30

Ala Pro Asn Asp Leu Asn Ala Phe Trp Met Pro Phe Thr Ala Asn Arg 35 40 45 Ala Phe Lys Arg Ala Pro Lys Met Val Val Gly Ala Glu Gly Met His 50 55 60

Tyr lie Thr Ala Asp Gly Arg Lys lie lie Asp Ala Ala Ser Gly Met 65 70 75 80 47 £ 201127961

Trp Cys Thr Asn Ala Gly His Gly Arg Lys Glu lie Ala Glu Ala lie 85 90 95

Lys Ala Gin Ala Asp Glu Leu Asp Phe Sex Pro Pro Phe Gin Phe Gly 100 105 110

Gin Pro Lys Ala Phe Glu Leu Ala Ser Arg lie Ala Asp Leu Ala Pro 115 120 125

Glu Gly Leu Asp His Val Phe Phe Cys Asn Ser Gly Ser Glu Ala Gly 130 135 140

Asp Thr Ala Leu Lys lie Ala Val Ala Tyr Gin Gin lie Lys Gly Gin 145 150 155 160

Gly Ser Arg Thr Arg Leu lie Gly Arg Glu Arg Gly Tyr His Gly Val 165 170 175

Gly Phe Gly Gly Thr Ala Val Gly Gly lie Gly Asn Asn Arg Lys Met 180 185 190

Phe Gly Pro Leu Leu Asn Gly Val Asp His Leu Pro Ala Thr Tyr Asp 195 200 205

Arg Asp Lys Gin Ala Phe Thr lie Gly Glu Pro Glu Tyr Gly Ala His 210 215 220

Phe Ala Glu Ala Leu Glu Gly Leu Val Asn Leu His Gly Ala Asn Thr 225 230 235 240 lie Ala Ala Val lie Val Glu Pro Met Ala Gly Ser Thr Gly Val Leu 245 250 255

Pro Ala Pro Lys Gly Tyr Leu Lys Lys Leu Arg Glu lie Thr Lys Lys 260 265 270

His Gly lie Leu Leu lie Phe Asp Glu Val lie Thr Gly Tyr Gly Arg 275 280 285

Leu Gly Tyr Ala Phe Ala Ser Glu Arg Tyr Gly Val Thr Pro Asp Met 290 295 300 lie Thr Phe Ala Lys Gly Val Thr Asn Gly Ala Val Pro Met Gly Gly 48 201127961 305 310 315 320

Val lie Thr Ser Ala Glu lie His Asp Ala Phe Met Thr Gly Pro Glu 325 330 335

His Ala Val Glu Leu Ala His Gly Tyr Thr Tyr Ser Ala His Pro Leu 340 345 350

Ala Cys Ala Ala Gly lie Ala Thr Leu Asp lie Tyr Arg Asp Glu Lys 355 360 365

Leu Phe Glu Arg Ala Lys Ala Leu Glu Pro Lys Phe Ala Glu Ala Val 370 375 380

Met Ser Leu Lys Ser Ala Pro Asn Val Val Asp lie Arg Thr Val Gly 385 390 395 400

Leu Thr Ala Gly lie Asp Leu Ala Ser lie Ala Asp Ala Val Gly Lys 405 410 415

Arg Gly Phe Glu Ala Met Asn Ala Gly Phe His Asp His Glu Leu Met 420 425 430

Leu Arg lie Ala Gly Asp Thr Leu Ala Leu Thr Pro Pro Leu lie Leu 435 440 445

Ser Glu Asp His lie Gly Glu lie Val Asp Lys Val Gly Lys Val lie 450 455 460

Arg Ala Val Ala 465 &lt;21〇&gt;3〇&lt;211&gt; 1263 &lt;212&gt; DNA &lt;213>Escherichia coli &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1). ·(1263) &lt;400&gt; 3〇at9 cca cat tea ctg ttc age acc gat acc gat etc acc gcc gaa aat

Met Pro His Ser Leu Phe Ser Thr Asp Thr Asp Leu Thr Ala Glu Asn 1 5 10 15 ctg ctg cgt ttg ccc get gaa ttt ggc tgc ccg gtg tgg gtc tac gat 48 96 49 201127961

Leu Leu Arg Leu Pro Ala 20 gcg caa att att cgt egg Ala Gin lie lie Arg Arg 35 gtg ege ttt gca cag aaa Val Arg Phe Ala Gin Lys 50 atg cgt gag cag ggc gtg Met Arg Glu Gin Gly Val 65 70 gag cgt gcg Ttg gcg gcg Glu Arg Ala Leu Ala Ala 85 att gtt ttt aeg gca gat lie VaX Phe Thr Ala Asp 100 agt gaa ttg caa att ccg Ser Glu Leu Gin lie Pro 115 caa ctg ggc cag gtt teg Gin Leu Gly Gin Val Ser 130 ccg Ggg ttt ggt cac gga Pro Gly Phe Gly His Gly 145 150 aac age aag cac ggt ate Asn Ser Lys His Gly lie 165 gtg ata caa cgt cat cat Val lie Gin Arg His His 180 ggt tet ggc gtt gat tat Gly Ser Gly Val Asp Tyr 195 gtg cgt cag gtc ate gaa Val Arg Gin Val lie Glu 210 ggc ggt ggg ett tet gtt Gly Gly Gly Leu Ser Val 225 230 acc gaa cat tat tat ggt Thr Glu His Tyr Tyr Gly 245

Glu Phe Gly Cys Pro Val 25 cag att gca gcg ctg aaa Gin lie Ala Ala Leu Lys 40 gcc tgt tcc aat att cat Ala Cys Ser Asn lie His 55 60 aaa gtg gat tcc gtc teg Lys Val Asp Ser Val Ser 75 ggt tac aat Ccg caa aeg Gly Tyr Asn Pro Gin Thr 90 gtt ate gat cag gcg aeg Val lie Asp Gin Ala Thr 105 gtg aat gcg ggt tet gtt Val Asn Ala Gly Ser Val 120 cca ggg cat egg gta tgg Pro Gly His Arg Val Trp 135 140 Cat age caa aaa acc aat His Ser Gin Lys Thr Asn 155 tgg tac acc gat ctg ccc Trp Tyr Thr Asp Leu Pro 170 ctg cag ctg gtc ggc att Leu Gin Leu Val Gly lie 185 gcc cat ctg gaa cag gtg Ala His Leu Glu Gin Val 200 ttc ggt cag gat tta cag Phe Gly Gin Asp Leu Gin 215 220 cct tat caa cag ggt gaa Pro Tyr Gin Gin Gly Glu 235 ctg tgg aat gcc gcg cgt Leu Trp Asn Ala Ala Arg 250

Trp Val Tyr Asp 30 cag ttt gat gtg 144

Gin Phe Asp Val 45 att ttg ege tta 192 lie Leu Arg Leu tta ggc gaa ata 240

Leu Gly Glu lie 80 cac ccc gat gat 288

His Pro Asp Asp 95 ett gaa ege gtc 336

Leu Glu Arg Val 110 gat atg etc gac 384

Asp Met Leu Asp 125 ctg ege gtt aat 432

Leu Arg Val Asn acc ggt ggc gaa 480

Thr Gly Gly Glu 160 gcc gca ctg gac 528

Ala Ala Leu Asp 175 cac atg cac att 576

His Met His lie 190 tgt ggt get atg 624

Cys Gly Ala Met 205 get att tet gcg 672

Ala lie Ser Ala gag gcg gtt gat 720

Glu Ala Val Asp 240 gag caa ate gcc 768

Glu Gin lie Ala 255 50 201127961 cgc cat ttg ggc cac cct gtg aaa ctg gaa att gaa ccg ggt cgc ttc 816

Arg His Leu Gly His Pro Val Lys Leu Glu lie Glu Pro Gly Arg Phe 260 265 270 ctg gta gcg cag tct ggc gta tta att act cag gtg egg age gtc aaa 864

Leu Val Ala Gin Ser Gly Val Leu lie Thr Gin Val Arg Ser Val Lys 275 280 285 caa atg ggg age cgc cac ttt gtg ctg gtt gat gcc ggg ttc aac gat 912

Gin Met Gly Ser Arg His Phe Val Leu Val Asp Ala Gly Phe Asn Asp 290 295 300 ctg atg cgc ccg gca atg tac ggt agt tac cac cat ate agt gcc ctg 960

Leu Met Arg Pro Ala Met Tyr Gly Ser Tyr His His lie Ser Ala Leu 305 310 315 320 gca get gat ggt cgt tet ctg gaa cac geg cca aeg gtg gaa acc gtc 1008

Ala Ala Asp Gly Arg Ser Leu Glu His Ala Pro Thr Val Glu Thr Val 325 330 335 gtc gcc gga ccg tta tgt gaa teg ggc gat gtc ttt acc cag cag gaa 1056

Val Ala Gly Pro Leu Cys Glu Ser Gly Asp Val Phe Thr Gin Gin Glu 340 345 350 999 99a gtt gaa acc cgc gcc ttg ccg gaa gtg aag gca ggt gat 1104

Gly Gly Asn Val Glu Thr Arg Ala Leu Pro Glu Val Lys Ala Gly Asp 355 360 365 tat ctg gta ctg cat gat aca ggg gca tat ggc gca tea atg tea tcc 1152

Tyr Leu Val Leu His Asp Thr Gly Ala Tyr Gly Ala Ser Met Ser Ser 370 375 380 aac tac aat age cgt ccg ctg tta cca gaa gtt ctg ttt gat aat ggt 1200

Asn Tyr Asn Ser Arg Pro Leu Leu Pro Glu Val Leu Phe Asp Asn Gly 385 390 395 400 cag geg egg ttg att cgc cgt cgc cag acc ate gaa gaa tta ctg geg 1248

Gin Ala Arg Leu He Arg Arg Arg Gin Thr lie Glu Glu Leu Leu Ala 405 410 415 ctg gaa ttg ett taa 1263

Leu Glu Leu Leu 420 &lt;210&gt; 31 &lt;211&gt; 420

&lt;212&gt; PRT &lt;213&gt; Escherichia coli &lt;400&gt;

Met Pro His Ser Leu Phe Ser Thr Asp Thr Asp Leu Thr Ala Glu Asn 1 5 i〇 15

Leu Leu Arg Leu Pro Ala Glu Phe Gly Cys Pro Val Trp Val Tyr Asp 20 25 30 e; 51 201127961

Ala Gin lie lie Arg Arg Gin lie Ala Ala Leu Lys Gin Phe Asp Val 35 40 45

Val Arg Phe Ala Gin Lys Ala Cys Ser Asn lie His lie Leu Arg Leu 50 55 60

Met Arg Glu Gin Gly Val Lys Val Asp Ser Val Ser Leu Gly Glu lie 65 70 75 80

Glu Arg Ala Leu Ala Ala Gly Tyr Asn Pro Gin Thr His Pro Asp Asp 85 90 95 lie Val Phe Thr Ala Asp Val lie Asp Gin Ala Thr Leu Glu Arg Val 100 105 110

Ser Glu Leu Gin lie Pro Val Asn Ala Gly Ser Val Asp Met Leu Asp 115 120 125

Gin Leu Gly Gin Val Ser Pro Gly His Arg Val Trp Leu Arg Val Asn 130 135 140

Pro Gly Phe Gly His Gly His Ser Gin Lys Thr Asn Thr Gly Gly Glu 145 150 155 160

Asn Ser Lys His Gly lie Trp Tyr Thr Asp Leu Pro Ala Ala Leu Asp 165 170 175

Val lie Gin Arg His His Leu Gin Leu Val Gly lie His Met His lie 180 185 190

Gly Ser Gly Val Asp Tyr Ala His Leu Glu Gin Val Cys Gly Ala Met 195 200 205

Val Arg Gin Val lie Glu Phe Gly Gin Asp Leu Gin Ala lie Ser Ala 210 215 220

Gly Gly Gly Leu Ser Val Pro Tyr Gin Gin Gly Glu Glu Ala Val Asp 225 230 235 240

Thr Glu His Tyr Tyr Gly Leu Trp Asn Ala Ala Arg Glu Gin lie Ala 245 250 255

Arg His Leu Gly His Pro Val Lys Leu Glu lie Glu Pro Gly Arg Phe 260 265 270 52 201127961

Leu Val Ala Gin Ser Gly Val Leu lie Thr Gin Val Arg Ser Val Lys 275 280 285

Gin Met Gly Ser Arg His Phe Val Leu Val Asp Ala Gly Phe Asn Asp 290 295 300

Leu Met Arg Pro Ala Met Tyr Gly Ser Tyr His His lie Ser Ala Leu 305 310 315 320

Ala Ala Asp Gly Arg Ser Leu Glu His Ala Pro Thr Val Glu Thr Val 325 330 335

Val Ala Gly Pro Leu Cys Glu Ser Gly Asp Val Phe Thr Gin Gin Glu 340 345 350

Gly Gly Asn Val Glu Thr Arg Ala Leu Pro Glu Val Lys Ala Gly Asp 355 360 365

Tyr Leu Val Leu His Asp Thr Gly Ala Tyr Gly Ala Ser Met Ser Ser 370 375 380

Asn Tyr Asn Ser Arg Pro Leu Leu Pro Glu Val Leu Phe Asp Asn Gly 385 390 395 400

Gin Ala Arg Leu lie Arg Arg Arg Gin Thr lie Glu Glu Leu Leu Ala 405 410 415

Leu Glu Leu Leu 420 &lt;210&gt; 32 &lt;211&gt; 1265 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Escherichia, coli diaminopimelate enzyme LysA codon-optimized gene &lt; 400 &gt; 32 60 120 180 240 atatgccaca ctctctgttt tctactgata ctgatctgac tgcggaaaac ctgctgcgtc tgccggctga attcggttgt ccggtatggg tgtacgacgc tcagattatt cgtcgccaga tcgcagcact gaagcagttc gatgtagtgc gttttgcaca gaaggcgtgc tccaacatcc atatcctgcg cctgatgcgt gagcagggcg ttaaagttga ctccgtctct ctgggtgaga 53 £ 300 201127961 ttgagcgcgc cctggcagcc ggctataacc cacagaccca tcctgacgac attgtattta ctgccgacgt gatcgaccag gctactctgg aacgcgtttc tgaactgcag atcccggtta atgctggttc tgtggacatg ctggaccagc tgggccaggt atccccaggt catcgtgtgt 99Ctgcgtgt caacccaggt ttcggccacg gccactctca gaaaactaac actggtggtg agaactccaa gcatggcatt tggtataccg atctgccggc tgcactggac gtaatccagc gtcaccacct gcagctggtg ggcatccaca tgcacattgg ctccggcgta gactacgccc acctggagca agtctgcggt gctatggtac gtcaggtaat cgagttcggc caagatctgc aggcaatcag cgct ggtggc ggcctgtctg taccttatca gcagggcgag gaggcggttg acactgagca ctactacggt ctgtggaacg ccgctcgtga gcaaattgca cgtcacctgg gccacccggt gaaactggag atcgagccgg gccgcttcct ggtagcacag tccggcgtac tgattaccca ggtacgctct gttaaacaga tgggctcccg tcactttgtg ctggtagacg caggcttcaa cgacctgatg cgtccggcta tgtatggttc ctatcatcac atctctgcgc tggccgccga cggccgctct ctggaacacg cgccgacggt tgaaacggtg gtggctggtc cgctgtgcga gtccggcgac gttttcactc agcaggaggg cggcaatgta gagacgcgtg cgctgccgga agtgaaagcc ggtgattatc tggtgctgca tgataccggc gcctatggtg cgagcatgag cagcaactac aactctcgcc Cgctgctgcc ggaggtcctg ttcgataacg gccaagcccg cctgatccgt cgtcgtcaga ccatcgagga actgctggca ctggagctgc tgtaa &lt;210&gt; 33 &lt;211&gt; 1692

&lt;212&gt; DNA &lt;213&gt; Saccharomyces cerevisiae 360 420 480 540 600 660 720 780 840 900 960 1020 1080 1140 1200 1260 1265 &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) (1692) &lt;400&gt; 33 atg tct gaa att act ttg ggt aaa tat ttg ttc gaa aga tta aag caa Met Ser Glu lie Thr Leu Gly Lys Tyr Leu Phe Glu Arg Leu Lys Gin 15 10 15 gtc aac gtt aac Val Asn Val Asn 20 ttg ttg gac aag Leu Leu Asp Lys 35 acc gtt ttc ggt Thr Val Phe Gly ate tac gaa gtt He Tyr Glu Val 40 ttg cca ggt gac Leu Pro Gly Asp 25 gaa ggt atg aga Glu Gly Met Arg ttc aac ttg Tcc Phe Asn Leu Ser 30 tgg get ggt aac Trp Ala Gly Asn 45 48 96 54 144 192201127961 gcc aac gaa ttg aac get get tac gee get gat ggt tac get cgt ate Ala Asn Glu Leu Asn Ala Ala Ala Ala Ala Ala Asp Gly Tyr Ala Arg lie 50 55 60 aag ggt atg tet tgt ate ate acc acc ttc ggt gtc ggt gaa ttg tet Lys Gly Met Ser Cys lie lie Thr Thr Phe Gly Val Gly Glu Leu Ser 65 70 75 80 get ttg aac ggt att gcc ggt tet tac Get gaa cac gtc ggt gtt ttg Ala Leu Asn Gly lie Ala Gly Ser Tyr Ala Glu His Val Gly Val Leu 85 90 95 cac gtt gtt ggt gtc cca tee ate tet get caa get aag caa ttg ttg His Val Val Gly Val Pro Ser lie Ser Ala Gin Ala Lys Gin Leu Leu 100 105 110 ttg cac cac acc ttg ggt aac ggt gac ttc act gtt ttc cac aga atg Leu His His Thr Leu Gly Asn Gly Asp Phe Thr Val Phe His Arg Met 115 120 125 tet gcc aac att tet gaa acc act get atg ate act gac Att get acc Ser Ala Asn lie Ser Glu Thr Thr Ala Met lie Thr Asp lie Ala Thr 130 135 140 gee cca get gaa att gac aga tgt ate aga acc act tac gtc acc caa Ala Pro Ala Glu lie Asp Arg Cys lie Arg Thr Thr Tyr Val Thr Gin 145 150 155 160 aga cca gtc tac tta ggt ttg cca get aac ttg gtc gac ttg aac gtc Arg Pro Val Tyr Leu Gly Leu Pro Ala Asn Leu Val Asp Leu Asn Val 165 170 175 cca get aag ttg ttg caa act Cca att gac atg tet ttg aag cca aac Pro Ala Lys Leu Leu Gin Thr Pro lie Asp Met Ser Leu Lys Pro Asn 180 185 190 gat get gaa tee gaa aag gaa gtc att gac acc ate ttg get ttg gtc Asp Ala Glu Ser Glu Lys Gl u Val lie Asp Thr lie Leu Ala Leu Val 195 200 205 aag gat get aag aac cca gtt ate ttg get gat get tgt tgt tee aga Lys Asp Ala Lys Asn Pro Val lie Leu Ala Asp Ala Cys Cys Ser Arg 210 215 220 cac gac Gtc ag get gaa act aag aag ttg att gac ttg act caa ttc His Asp Val Lys Ala Glu Thr Lys Lys Leu lie Asp Leu Thr Gin Phe 225 230 235 240 cca get ttc gtc acc cca atg ggt aag ggt tee att gac gaa caa cac Pro Ala Phe Val Thr Pro Met Gly Lys Gly Ser lie Asp Glu Gin His 245 250 255 cca aga tac ggt ggt gtt tac gtc ggt acc ttg tee aag cca gaa gtt Pro Arg Tyr Gly Gly Val Tyr Val Gly Thr Leu Ser Lys Pro Glu Val 260 265 270 aag gaa gcc gtt gaa tet get gac ttg att ttg tet gtc ggt get ttg Lys Glu Ala Val Glu Ser Ala Asp Leu lie Leu Ser Val Gly Ala Leu 240 288 336 384 432 480 528 576 624 672 720 768 816 55 ε 864 201127961 275 ttg tct gat ttc aac acc Leu Ser Asp Phe Asn Thr 290 aac att gtc gaa ttc cac Asn lie Val Glu Phe His 305 310 ttc cca ggt gtc caa atg Phe Pro Gly Val Gin Met 325 att get gac gcc get aa g Ala Asp Ala Ala Lys 340 act cca get aac get get Thr Pro Ala Asn Ala Ala 355 tgg atg tgg aac caa ttg Trp Met Trp Asn Gin Leu 370 att get gaa acc ggt acc lie Ala Glu Thr Gly Thr 385 390 cca Aac aac acc tac ggt Pro Asn Asn Thr Tyr Gly 405 ttc acc act ggt get acc Phe Thr Thr Gly Ala Thr 420 gat cca aag aag aga gtt Asp Pro Lys Lys Arg Val 435 ttg act gtt caa gaa ate Leu Thr Val Gin Glu lie 450 tac ttg ttc gtc ttg aac Tyr Leu Phe Val Leu Asn 465 470 cac ggt cca aag get caa His Gly Pro Lys Ala Gin 485 tee ttg ttg cca act ttc Ser Leu Leu Pro Thr Phe 500 get acc acc ggt gaa tgg 280 ggt tct Ttc tct tac tct Gly Ser Phe Ser Tyr Ser 295 300 tcc gac cac atg ag ate Ser Asp His Met Lys lie 315 aaa ttc gtt ttg caa aag Lys Phe Val Leu Gin Lys 330 ggt tac aag cca gtt get Gly Tyr Lys Pro Val Ala 345 gtc cca get tct acc cca Val Pro Ala Ser Thr Pro 360 ggt aac ttc ttg caa gaa Gly Asn Phe Leu Gin Glu 375 380 tcc get ttc ggt ate aac Ser Ala Phe Gly lie Asn 395 ate tct caa gtc tta tgg lie Ser Gin Val Leu Trp 410 tt:9 99fc 9ct 9ct ttc get Leu Gly Ala Ala Phe Ala 425 ate tta ttc att ggt gac lie Leu Phe lie Gly Asp 440 tcc acc atg ate aga tgg Ser Thr Met lie Arg Trp 455 460 aac gat ggt tac acc Att Asn Asp Gly Tyr Thr lie 475 tac aac gaa att caa ggt Tyr Asn Glu lie Gin Gly 490 ggt get aag gac tat gaa Gly Ala Lys Asp Tyr Glu 505 gac aag ttg acc caa gac 285 tac aag acc aag 912

Tyr Lys Thr Lys aga aac gcc act 960

Arg Asn Ala Thr 320 ttg ttg acc act 1008

Leu Leu Thr Thr 335 gtc cca get aga 1056

Val Pro Ala Arg 350 ttg aag caa gaa 1104

Leu Lys Gin Glu 365 ggt gat gtt gtc 1152

Gly Asp Val Val caa acc act ttc 1200

Gin Thr Thr Phe 400 ggt tcc att ggt 1248

Gly Ser lie Gly 415 get gaa gaa att 1296

Ala Glu Glu He 430 ggt tct ttg caa 1344

Gly Ser Leu Gin 445 ggc ttg aag cca 1392

Gly Leu Lys Pro gaa aag ttg att 1440

Glu Lys Leu lie 480 tgg 9ac cac eta 1488

Trp Asp His Leu 495 acc cac aga gtc 1536

Thr His Arg Val 510 aag tct ttc aac 1584 56 201127961

Ala Thr Thr Gly Glu Trp Asp Lys Leu Thr Gin Asp Lys Ser Phe Asn 515 520 525 gac aac tct aag ate aga atg att gaa ate atg ttg cca gtc ttc gat Asp Asn Ser Lys lie Arg Met lie Glu lie Met Leu Pro Val Phe Asp 530 535 540 get cca caa aac ttg gtt gaa caa get aag ttg act get get acc aac Ala Pro Gin Asn Leu Val Glu Gin Ala Lys Leu Thr Ala Ala Thr Asn 545 550 555 560 get aag caa taa Ala Lys Gin 1632 1680 1692 &lt;210&gt; 34 &lt;211&gt; 563

&lt;212&gt; PRT &lt;213&gt; Saccharomyces cerevisiae &lt;400&gt; 34

Met Ser Glu lie Thr Leu Gly Lys Tyr Leu Phe Glu Arg Leu Lys Gin 15 10 15

Val Asn Val Asn Thr Val Phe Gly Leu Pro Gly Asp Phe Asn Leu Ser 20 25 30

Leu Leu Asp Lys lie Tyr Glu Val Glu Gly Met Arg Trp Ala Gly Asn 35 40 _ 45

Ala Asn Glu Leu Asn Ala Ala Tyr Ala Ala Asp Gly Tyr Ala Arg lie 50 55 60

Lys Gly Met Ser Cys lie lie Thr Thr Phe Gly Val Gly Glu Leu Ser 65 70 75 80

Ala Leu Asn Gly lie Ala Gly Ser Tyr Ala Glu His Val Gly Val Leu 85 90 95

His Val Val Gly Val Pro Ser lie Ser Ala Gin Ala Lys Gin Leu Leu 100 105 110

Leu His His Thr Leu Gly Asn Gly Asp Phe Thr Val Phe His Arg Met 115 120 125

Ser Ala Asn lie Ser Glu Thr Thr Ala Met lie Thr Asp lie Ala Thr 130 135 140 57 201127961

Ala Pro Ala Glu lie Asp Arg Cys lie Arg Thr Thr Tyr Val Thr Gin 145 150 155 160

Arg Pro Val Tyr Leu Gly Leu Pro Ala Asn Leu Val Asp Leu Asn Val 165 170 175

Pro Ala Lys Leu Leu Gin Thr Pro lie Asp Met Ser Leu Lys Pro Asn 180 185 190

Asp Ala Glu Ser Glu Lys Glu Val lie Asp Thr He Leu Ala Leu Val 195 200 205

Lys Asp Ala Lys Asn Pro Val lie Leu Ala Asp Ala Cys Cys Ser Arg 210 215 220

His Asp Val Lys Ala Glu Thr Lys Lys Leu lie Asp Leu Thr Gin Phe 225 230 235 240

Pro Ala Phe Val Thr Pro Met Gly Lys Gly Ser lie Asp Glu Gin His 245 250 255

Pro Arg Tyr Gly Gly Val Tyr Val Gly Thr Leu Ser Lys Pro Glu Val 260 265 270

Lys Glu Ala Val Glu Ser Ala Asp Leu lie Leu Ser Val Gly Ala Leu 275 280 285

Leu Ser Asp Phe Asn Thr Gly Ser Phe Ser Tyr Ser Tyr Lys Thr Lys 290 295 300

Asn lie Val Glu Phe His Ser Asp His Met Lys lie Arg Asn Ala Thr 305 310 315 320

Phe Pro Gly Val Gin Met 325

Lys Phe Val Leu Gin Lys Leu Leu Thr Thr 330 335 lie Ala Asp Ala Ala Lys Gly Tyr Lys Pro Val Ala Val Pro Ala Arg 340 345 350

Thr Pro Ala Asn Ala Ala Val Pro Ala Ser Thr Pro Leu Lys Gin Glu 355 360 365

Trp Met Trp Asn Gin Leu Gly Asn Phe Leu Gin Glu Gly Asp Val Val 370 375 380 58 201127961 lie Ala Glu Thr Gly Thr Ser Ala Phe Gly lie Asn Gin Thr Thr Phe 385 390 395 400

Pro Asn Asn Thr Tyr Gly lie Ser Gin Val Leu Trp Gly Ser lie Gly 405 410 415

Phe Thr Thr Gly Ala Thr Leu Gly Ala Ala Phe Ala Ala Glu Glu lie 420 425 430

Asp Pro Lys Lys Arg Val lie Leu Phe lie Gly Asp Gly Ser Leu Gin 435 440 445

Leu Thr Val Gin Glu lie Ser Thr Met lie Arg Trp Gly Leu Lys Pro 450 455 460

Tyr Leu Phe Val Leu Asn Asn Asp Gly Tyr Thr lie Glu Lys Leu lie 465 470 475 480

His Gly Pro Lys Ala Gin Tyr Asn Glu lie Gin Gly Trp Asp His Leu 485 490 495

Ser Leu Leu Pro Thr Phe Gly Ala Lys Asp Tyr Glu Thr His Arg Val 500 505 510

Ala Thr Thr Gly Glu Trp Asp Lys Leu Thr Gin Asp Lys Ser Phe Asn 515 520 525

Asp Asn Ser Lys lie Arg Met lie Glu lie Met Leu Pro Val Phe Asp 530 535 540

Ala Pro Gin Asn Leu Val Glu Gin Ala Lys Leu Thr Ala Ala Thr Asn 545 550 555 560

Ala Lys Gin &lt;210&gt; 35 &lt;211&gt; 1692 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;22j&gt; Saccharomyces cerevisiae propionate dehydrogenase Pdc codon optimized gene &lt; 400 &gt; 35 atgtccgaga tcactctggg caaatacctg tttgaacgtc tgaaacaggt gaacgttaat 60 59 201127961 accgtattcg gcctgccggg tgatttcaac ctgtccctgc tggacaaaat ctatgaagtt 120 gaaggtatgc gttgggctgg caacgctaac gagctgaacg cagcgtacgc ggcagatggt 180 tacgctcgta tcaaaggtat gtcttgtatc atcaccacct tcggtgttgg tgagctgagc 240 gccctgaacg gcatcgccgg ctcctatgca gagcacgtgg gcgtgctgca cgttgtgggt 300 gtaccgtcca tcagcgccca ggcaaaacag ctgctgctgc accacaccct gggtaacggc 360 gactttaccg ttttccatcg tatgtctgcg aacatcagcg aaactactgc aatgattact 420 gacatcgcta cggcaccggc agaaatcgac cgttgcattc gtaccacgta cgttactcag 480 cgcccggttt atctgggcct gccagccaac ctggtggatc tgaacgtccc ggctaaactg 540 ctgcagactc cgatcgatat gtctctgaaa cctaacgacg cagaatctga gaaagaagtt 600 atcgatacta ttctggctct ggtgaaagat gcaaagaacc cagttatcct ggctgacg ca 660 tgttgctctc gtcatgatgt aaaggcagaa accaaaaagc tgatcgacct gacgcagttc 720 ccggcgttcg ttaccccgat gggcaagggt tccatcgatg agcagcaccc gcgttatggt 780 ggtgtatacg ttggcacgct gtccaaaccg gaggtaaaag aagcggttga aagcgcagat 840 ctgatcctgt ctgttggtgc actgctgagc gacttcaaca ccggttcttt ctcctatagc 900 tacaagacca aaaacattgt ggagtttcac tccgatcaca tgaaaatccg caacgcgacc 960 tttcctggtg tgcagatgaa attcgtactg cagaaactgc tgaccaccat cgccgacgct 1020 gcgaaaggtt ataaaccggt agctgtgccg gcacgtaccc cggcgaacgc cgcggttcct 1080 gcatccactc cactgaagca ggaatggatg tggaatcagc tgggtaattt cctgcaagaa 1140 ggcgacgttg taatcgcaga aaccggcact agcgcgtttg gcattaacca gacgaccttc 1200 ccaaacaaca cctacggtat cagccaagtc ctgtggggct ctatcggctt caccaccggt 1260 gcaaccctgg gtgcggcttt cgctgctgag gagatcgacc cgaagaaacg tgttatcctg 1320 ttcatcggtg acggctccct gcagctgacc gtccaggaga tttctaccat gatccgctgg 1380 ggcctgaaac cgtacctgtt tgtgctgaac aacgacggct acactattga gaaactgatc 1440 cacggtccga aagcacagta taatgagatc cagggttggg atcatctgtc tctgct gccg 1500 acctttggcg ctaaagacta cgagacccac cgcgtggcta ccaccggcga gtgggataaa 1560 ctgacgcagg ataaatcctt caatgacaat agcaagattc gtatgatcga aatcatgctg 1620 ccggtctttg atgctccgca gaacctggta gagcaagcaa aactgaccgc ggcaactaac 1680 gctaaacagt aa 1692 &lt; 210 &gt; 36 60 201127961 &lt; 211 &gt; 1707 &lt; 212 &gt; DNA &lt; 213 &gt; motion Aeromonas Zymomonas mobilis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1)..(1707) &lt;400&gt; 36 atg agt tat act gtc ggt acc tat tta gcg gag egg ett gtc cag att Met Ser Tyr Thr Val Gly Thr Tyr Leu Ala Glu Arg Leu Val Gin lie 15 10 15 ggt etc aag cat cac ttc gca gtc gcg ggc gac tac aac etc gtc ett Gly Leu Lys His His Phe Ala Val Ala Gly Asp Tyr Asn Leu Val Leu 20 25 30 ett gac aac ctg ett ttg aac aaa aac atg gag cag gtt tat tgc tgt Leu Asp Asn Leu Leu Leu Asn Lys Asn Met Glu Gin Val Tyr Cys Cys 35 40 45 aac gaa ctg aac tgc ggt ttc agt gca gaa ggt tat get cgt Gee aaa Asn Glu Leu Asn Cys Gly Phe Ser Ala Glu Gly Tyr Ala Arg Ala Lys 50 55 60 ggc gca gca gca gee Gtc gtt acc tac age gtc ggt gcg ett tee gca Gly Ala Ala Ala Ala Val Val Thr Tyr Ser Val Gly Ala Leu Ser Ala 65 70 75 80 ttt gat get ate ggt ggc gee tat gca gaa aac ett ccg gtt ate ctg Phe Asp Ala Lie Gly Gly Ala Tyr Ala Glu Asn Leu Pro Val lie Leu 85 90 95 ate tee ggt get ccg aac aac aat gat cac get get ggt cac gtg ttg lie Ser Gly Ala Pro Asn Asn Asn His Ala Ala Gly His Val Leu 100 105 110 cat cac get ett ggc aaa acc gac tat cac tat cag ttg gaa atg gee His His Ala Leu Gly Lys Thr Asp Tyr His Tyr Gin Leu Glu Met Ala 115 120 125 aag aac ate aeg gee gee get gaa gcg att tac acc ccg gaa Gaa get Lys Asn lie Thr Ala Ala Ala Glu Ala lie Tyr Thr Pro Glu Glu Ala 130 135 140 ccg get aaa ate gat cac gtg att aaa act get ett cgt gag aag aag Pro Ala Lys lie Asp His Val lie Lys Thr Ala Leu Arg Glu Lys Lys 145 150 155 160 ccg gtt tat etc gaa ate get tgc aac att get tee atg ccc tgc gee Pro Val Tyr Leu Glu He Ala Cys Asn lie Ala Ser Met Pro Cys Ala 165 170 175 get cct gga ccg gca age gca ttg Ttc aat Gac gaa gee age gac gaa Ala Pro Gly Pro Ala Ser Ala Leu Phe Asn Asp Glu Ala Ser Asp Glu 180 185 190 get tet ttg aat gca gcg gtt gaa gaa acc ctg aaa ttc ate gee aac 48 96 144 192 240 288 336 384 432 480 528 576 624 61

E 201127961

Ala Ser Leu Asn Ala Ala Val Glu Glu 195 200 cgc gac aaa gtt gcc gtc etc gtc ggc

Arg Asp Lys Val Ala Val Leu Val Gly 210 215 get gaa gaa get get gtc aaa ttt get

Ala Glu Glu Ala Ala Val Lys Phe Ala 225 230 get acc atg get get gca aaa age ttc

Ala Thr Met Ala Ala Ala Lys Ser Phe 245 tac ate ggc acc tea tgg ggt gaa gtc

Tyr lie Gly Thr Ser Trp Gly Glu Val 260 265 aeg atg aaa gaa gcc gat geg gtt ate

Thr Met Lys Glu Ala Asp Ala Val lie 275 280 gac tac tee acc act ggt tgg aeg gat

Asp Tyr Ser Thr Thr Gly Trp Thr Asp 290 295 gtt etc get gaa ccg cgt tet gtc gtc

Val Leu Ala Glu Pro Arg Ser Val Val 305 310 age gtc cat ctg aaa gac tat ctg acc

Ser Val His Leu Lys Asp Tyr Leu Thr 325 aag aaa acc ggt gca ttg gac ttc ttc

Lys Lys Thr Gly Ala Leu Asp Phe Phe 340 345 ctg aag aaa gcc get ccg get gat ccg

Leu Lys Lys Ala Ala Pro Ala Asp Pro 355 360 gaa ate gcc cgt cag gtc gaa get ett

Glu lie Ala Arg Gin Val Glu Ala Leu 370 375 att get gaa acc ggt gac tet tgg ttc lie Ala Glu Thr Gly Asp Ser Trp Phe 385 390 ccg aac ggt get cgc gtt gaa tat gaa

Pro Asn Gly Ala Arg Val Glu Tyr Glu 405 tgg tee gtt cct gcc gcc ttc ggt tat

Trp Ser Val Pro Ala Ala Phe Gly Tyr 420 425

Thr Leu Lys Phe lie Ala Asn 205 age aag ctg cgc gca get ggt 672

Ser Lys Leu Arg Ala Ala Gly 220 gat get etc ggt ggc gca gtt 720

Asp Ala Leu Gly Gly Ala Val 235 240 ttc cca gaa gaa aac ccg cat 768

Phe Pro Glu Glu Asn Pro His 250 255 age tat ccg ggc gtt gaa aag 816

Ser Tyr Pro Gly Val Glu Lys 270 get ctg get cct gtc ttc aac 864

Ala Leu Ala Pro Val Phe Asn 285 att cct gat cct aag aaa ctg 912 lie Pro Asp Pro Lys Lys Leu 300 gtt aac ggc att cgc ttc ccc 960

Val Asn Gly lie Arg Phe Pro 315 320 cgt ttg get cag aaa gtt tee 1008 Arg Leu Ala Gin Lys Val Ser 330 335 aaa tee etc aat gca ggt gaa 1056 Lys Ser Leu Asn Ala Gly Glu 350 agt get ccg ttg gtc aac gca 1104 Ser Ala Pro Leu Val Asn Ala 365 ctg acc ccg aac aeg aeg gtt 1152 Leu Thr Pro Asn Thr Thr Val 380 aat get cag cgc atg ag etc 1200 Asn Ala Gin Arg Met Lys Leu 395 400 atg cag tgg ggt cac att ggt 1248 Met Gin Trp Gly His lie Gly 410 415 gee gtc ggt get ccg gaa cgt 1296 Ala Val Gly Ala Pro Glu Arg 430 62 1344 201127961 1344 cgc aac ate etc atg gtt ggt gat ggt tee ttc cag ctg aeg get cag Arg Asn lie Leu Met Val Gly Asp Gly Ser Phe Gin Leu Thr Ala Gin 435 440 445 gaa gtc get cag atg gtt cgc ctg aaa ctg ccg gtt ate ate ttc ttg Glu Val Ala Gin Met Val Arg Leu Lys Leu Pro Val lie lie Phe Leu 450 455 460 ate aat Aac tat ggt tac acc gee gaa gtt atg ate cat gat ggt ccg lie Asn Asn Tyr Gly Tyr Thr Ala GXu Val Met lie His Asp Gly Pro 465 470 475 480 tac aac aac ate aag aac tgg gat tat gee ggt ctg at g gaa gtg ttc Tyr Asn Asn lie Lys Asn Trp Asp Tyr Ala Gly Leu Met Glu Val Phe 485 490 495 aac ggt aac ggt ggt tat gac age ggt get ggt aaa ggc ctg aag get Asn Gly Asn Gly Gly Tyr Asp Ser Gly Ala Gly Lys Gly Leu Lys Ala 500 505 510 aaa acc ggt ggc gaa ctg gca gaa get ate aag gtt get ctg gca aac Lys Thr Gly Gly Glu Leu Ala Glu Ala He Lys Val Ala Leu Ala Asn 515 520 525 acc gac ggc cca acc ctg ate Gaa tgc ttc ate ggt cgt gaa gac tgc Thr Asp Gly Pro Thr Leu He Glu Cys Phe lie Gly Arg Glu Asp Cys 530 535 540 act gaa gaa ttg gtc aaa tgg ggt aag cgc gtt get gee gee aac age Thr Glu Glu Leu Val Lys Rp Lys Leu Leu 565 1392 1440 1488 1536 1584 1632 1680 1707 &lt;210&gt; 37 &lt

&lt;212&gt; PRT &lt;213&gt; Zymomonas mobilis &lt;400&gt; 37

Met Ser Tyr Thr Val Gly Thr Tyr Leu Ala Glu Arg Leu Val Gin He 1 5 10 15

Gly Leu Lys His His Phe Ala Val Ala Gly Asp Tyr Asn Leu Val Leu 20 25 30

Leu Asp Asn Leu Leu Leu Asn Lys Asn Met Glu Gin Val Tyr Cys Cys 35 40 45

Asn Glu Leu Asn Cys Gly Phe Ser Ala Glu Gly Tyr Ala Arg Ala Lys 5〇 55 60 g 63 201127961

Gly Ala Ala Ala Ala Val Val Thr Tyr Ser Val Gly Ala Leu Ser Ala 65 70 75 80

Phe Asp Ala lie Gly Gly Ala Tyr Ala Glu Asn Leu Pro Val lie Leu 85 90 95 lie Ser Gly Ala Pro Asn Asn Asn His Ala Ala Gly His Val Leu 100 105 110

His His Ala Leu Gly Lys Thr Asp Tyr His Tyr Gin Leu Glu Met Ala 115 120 125

Lys Asn lie Thr Ala Ala Ala Glu Ala lie Tyr Thr Pro Glu Glu Ala 130 135 140

Pro Ala Lys lie Asp His Val lie Lys Thr Ala Leu Arg Glu Lys Lys 145 150 155 160

Pro Val Tyr Leu*Glu lie Ala Cys Asn lie Ala Ser Met Pro Cys Ala 165 170 175

Ala Pro Gly Pro Ala Ser Ala Leu Phe Asn Asp Glu Ala Ser Asp Glu 180 185 190

Phe lie Ala Asn 205

Ala Ser Leu Asn Ala Ala Val Glu Glu Thr Leu Lys 195 200

Arg Asp Lys Val Ala Val Leu Val Gly Ser Lys Leu Arg Ala Ala Gly 210 215 220

Ala Glu Glu Ala Ala Val Lys Phe Ala Asp Ala Leu Gly Gly Ala Val 225 230 235 240

Ala Thr Met Ala Ala Ala Lys Ser Phe Phe Pro Glu Glu Asn Pro His 245 250 255

Tyr lie Gly Thr Ser Trp Gly Glu Val Ser Tyr Pro Gly Val Glu Lys 260 265 270

Thr Met Lys Glu Ala Asp Ala Val lie Ala Leu Ala Pro Val Phe Asn 275 280 285

Asp Tyr Ser Thr Thr Gly Trp Thr Asp lie Pro Asp Pro Lys Lys Leu 290 295 300 64 201127961

Val Leu Ala Glu Pro Arg Ser Val Val Val Asn Gly lie Arg Phe Pro 305 310 315 320

Ser Val His Leu Lys Asp Tyr Leu Thr Arg Leu Ala Gin Lys Val Ser 325 330 335

Lys Lys Thr Gly Ala Leu Asp Phe Phe Lys Ser Leu Asn Ala Gly Glu 340 345 350

Leu Lys Lys Ala Ala Pro Ala Asp Pro Ser Ala Pro Leu Val Asn Ala 355 360 365

Glu lie Ala Arg Gin Val Glu Ala Leu Leu Thr Pro Asn Thr Thr Val 370 375 380 lie Ala Glu Thr Gly Asp Ser Trp Phe Asn Ala Gin Arg Met Lys Leu 385 390 395 400

Pro Asn Gly Ala Arg Val Glu Tyr Glu Met Gin Trp Gly His lie Gly 405 410 415

Trp Ser Val Pro Ala Ala Phe Gly Tyr Ala Val Gly Ala Pro Glu Arg 420 425 430

Arg Asn lie Leu Met Val Gly Asp Gly Ser Phe Gin Leu Thr Ala Gin 435 440 445

Glu Val Ala Gin Met Val Arg Leu Lys Leu Pro Val lie lie Phe Leu 450 455 460 lie Asn Asn Tyr Gly Tyr Thr Ala Glu Val Met lie His Asp Gly Pro 465 470 475 480

Tyr Asn Asn lie Lys Asn Trp Asp Tyr Ala Gly Leu Met Glu Val Phe 485 490 495

Asn Gly Asn Gly Gly Tyr Asp Ser Gly Ala Gly Lys Gly Leu Lys Ala 500 S05 510

Lys Thr Gly Gly Glu Leu Ala Glu Ala lie Lys Val Ala Leu Ala Asn 515 520 525

Thr Asp Gly Pro Thr Leu lie Glu Cys Phe lie Gly Arg Glu Asp Cys 65 201127961 530 535 540

Thr Glu Glu Leu Val Lys Trp Gly Lys Arg Val Ala Ala Ala Asn Ser 545 550 555 560

Arg Lys Pro Val Asn Lys Leu Leu 565 &lt;210&gt; 38 &lt;211&gt; 1707 &lt;212&gt; DNA &lt;213&gt; Human 1 sequence &lt;220&gt;&lt;223&gt; Zymomonas mobilis after acid to enzyme PdcI472A codon-optimized gene &lt; 400 &gt; 38 atgtcttata ctgttggtac ttatctggct gagcgtctgg tgcaaatcgg cctgaaacac 60 cactttgcag ttgctggcga ctacaacctg gttctgctgg ataacctgct gctgaacaaa 120 aacatggagc aagtttattg ctgtaacgag ctgaactgcg gcttctctgc ggagggttat 180 gcgcgtgcga aaggtgccgc tgcagcagtc gtaacctact ctgtgggcgc tctgtccgcg 240 ttcgacgcaa tcggtggcgc ttacgctgaa aacctgccgg tgatcctgat tagcggtgcg 300 ccgaataata acgaccatgc tgctggccac gttctgcacc acgccctggg taaaactgat 360 taccattacc agctggagat ggctaaaaac atcactgcag cagcagaagc gatctacacc 420 ccggaagagg ctccggcaaa aatcgaccac gtgattaaaa ccgctctgcg tgagaaaaag 480 ccggtatacc tggaaatcgc gtgcaacatc gcgtctatgc cgtgcgccgc accgggtccg 540 gcttctgccc tgttcaacga tgaggcgagc gatgaggcat ctctgaacgc agcagtagaa 600 gaaaccctga aatttatcgc aaaccgtgac aaagtagcag tcctggtagg ttctaaactg 660 cgtgcggctg gtgcggaaga ggctgcggta aagttcgcgg atgctctggg cggtgcagtg 720 gcgaccatgg cagcggctaa atccttcttc ccagaggaga acccgcatta cattggtacc 780 tcctggggcg aagtttccta ccctggtgtg gagaaaacca tgaaagaagc cgatgctgtg 840 attgccctgg cgcctgtatt caacgattat tccaccaccg gttggaccga tatcccggac 900 ccgaagaaac tggtcctggc tgaaccgcgc tccgtagtag tgaatggcat tcgtttcccg 960 tccgtacacc tgaaggatta cctgacgcgt ctggcacaga aagtatccaa gaaaactggc 1020 gcgctggact tctttaaatc cctgaacgct ggtgagctga aaaaggcggc tccggccgat 1080 ccgtccgcac cgctggtgaa cgcagagatt gcacgtcagg ttgaggcact gctgacgccg 1140 aacaccaccg taatcgcgga aacgggcgac tcttggttca acgcacagcg catgaaactg 1200 66 201127961 ccgaacggtg cccgcgttga atatgaaatg cagtggggtc acatcggctg gtctgtccca gcagcgtttg gttacgcggt tggtgcaccg gagcgtcgca acatcctgat ggtgggtgac ggctccttcc agctgactgc tcaggaggtg gcgcagatgg tgcgcctgaa gctgccggtt atcattttcc tgatcaacaa ctacggctac accgccgagg taatgatcca cgatggtccg tacaacaaca tcaaaaactg ggactacgcc ggtctgatgg aggtttttaa cggtaacggc ggttacgaca gcggtgctgg taagggtctg aaagccaaaa ccggtggcga actggcagag gcgattaaag ttgcgctggc aaacaccgat ggcccgaccc tgatcgagtg cttcatcggc cgtgaggact gcaccgagga gctggtcaaa tggggcaaac gtgtggcggc tgctaactct cgcaagccgg taaacaaact gctgtaa 1260 1320 1380 1440 1500 1560 1620 1680 17 07 &lt; 210 &gt; 39 &lt; 211 &gt; 1644 &lt; 212 &gt; DNA &lt; 213 &gt; Lactococcus lactis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) . . (1644) &lt;400&gt; 39 atg tat aca gta gga gat tac ctg tta gac cga tta cac gag ttg Gga Met Tyr Thr Val Gly Asp Tyr Leu Leu Asp Arg Leu His Glu Leu Gly 15 10 15 att gaa gaa att ttt gga gtt cct ggt gac tat aac tta caa ttt tta lie Glu Glu lie Phe Gly Val Pro Gly Asp Tyr Asn Leu Gin Phe Leu 20 25 30 gat caa att att tea ege gaa gat atg aaa tgg att gga aat get aat Asp Gin lie lie Ser Arg Glu Asp Met Lys Trp lie Gly Asn Ala Asn 35 40 45 gaa tta aat get tet tat atg get gat ggt Tat get cgt act aaa aaa Glu Leu Asn Ala Ser Tyr Met Ala Asp Gly Tyr Ala Arg Thr Lys Lys 50 55 60 get Gcc gca ttt etc acc aca ttt gga gtc ggc gaa ttg agt geg ate Ala Ala Ala Phe Leu Thr Thr Phe Gly Val Gly Glu Leu Ser Ala lie 65 70 75 80 aat gga ctg gca gga agt tat gcc gaa aat tta cca gta gta gaa Att Asn Gly Leu Ala Gly Ser Tyr Ala Glu Asn Leu Pro Val Val Glu lie 85 90 95 gtt ggt tea cca act tea aaa gta caa aat gac gga aaa ttt gtc cat Val Gly Ser Pro Thr Ser Lys Val Gin Asn Asp Gly Lys Phe Val His 100 105 110 48 96 144 192 240 288 67

S 336 201127961 cat aca eta gca gat ggt gat ttt aaa cac ttt atg aag atg cat gaa 384

His Thr Leu Ala Asp Gly Asp Phe Lys His Phe Met Lys Met His Glu 115 120 125 cct gtt aca gca geg egg act tta ctg aca gca gaa aat gee aca tat 432

Pro Val Thr Ala Ala Arg Thr Leu Leu Thr Ala Glu Asn Ala Thr Tyr 130 135 140 gaa att gac ega gta ett tet caa tta eta aaa gaa aga aaa cca gtc 480

Glu lie Asp Arg Val Leu Ser Gin Leu Leu Lys Glu Arg Lys Pro Val 145 150 155 160 tat att aac tta cca gtc gat gtt get gca gca aaa gca gag aag cct 528

Tyr work Le Asn Leu Pro Val Asp Val Ala Ala Ala Lys Ala Glu Lys Pro 165 170 175 gca tta tet tta gaa aaa gaa age tet aca aca aat aca act gaa caa 576

Ala Leu Ser Leu Glu Lys Glu Ser Ser Thr Thr Asn Thr Thr Glu Gin 180 185 190 gtg att ttg agt ag att gaa gaa agt ttg aaa aat gee caa aaa cca 624

Val lie Leu Ser Lys lie Glu Glu Ser Leu Lys Asn Ala Gin Lys Pro 195 200 205 gta gtg att gca gga cac gaa gta att agt ttt ggt tta gaa aaa aeg 672

Val Val lie Ala Gly His Glu Val lie Ser Phe Gly Leu Glu Lys Thr 210 215 220 gta act cag ttt gtt tea gaa aca aaa eta ccg att aeg aca eta aat 720

Val Thr Gin Phe Val Ser Glu Thr Lys Leu Pro lie Thr Thr Leu Asn 225 230 235 240 ttt ggt aaa agt get gtt gat gaa tet ttg ccc tea ttt tta gga ata 768

Phe Gly Lys Ser Ala Val Asp Glu Ser Leu Pro Ser Phe Leu Gly lie 245 250 255 tat aac ggg aaa ett tea gaa ate agt ett aaa aat ttt gtg gag tee 816

Tyr Asn Gly Lys Leu Ser Glu lie Ser Leu Lys Asn Phe Val Glu Ser 260 265 270 gca gac ttt ate eta atg ett gga gtg aag ett aeg gac tee tea aca 864

Ala Asp Phe lie Leu Met Leu Gly Val Lys Leu Thr Asp Ser Ser Thr 275 280 285 ggt gca ttc aca cat cat tta gat gaa aat aaa atg att tea eta aac 912

Gly Ala Phe Thr His His Leu Asp Glu Asn Lys Met lie Ser Leu Asn 290 295 300 ata gat gaa gga ata att ttc aat aaa gtg gta gaa gat ttt gat ttt 960 lie Asp Glu Gly lie lie Phe Asn Lys Val Val Glu Asp Phe Asp Phe 305 310 315 320 aga gca gtg gtt tet tet tta tea gaa tta aaa gga ata gaa tat gaa 1008

Arg Ala Val Val Ser Ser Leu Ser Glu Leu Lys Gly lie Glu Tyr Glu 325 330 335 gga caa tat att gat aag caa tat gaa gaa ttt att cca tea agt get 1056

Gly Gin Tyr lie Asp Lys Gin Tyr Glu Glu Phe lie Pro Ser Ser Ala 340 345 350 68 201127961 ccc tta tea caa gac cgt eta tgg cag gca gtt gaa agt ttg act caa 1104

Pro Leu Ser Gin Asp Arg Leu Trp Gin Ala Val Glu Ser Leu Thr Gin 355 360 365 age aat gaa aca ate gtt get gaa caa gga acc tea ttt ttt gga get 1152

Ser Asn Glu Thr lie Val Ala Glu Gin Gly Thr Ser Phe Phe Gly Ala 370 375 380 tea aca att ttc tta aaa tea aat agt cgt ttt att gga caa cct tta 1200

Ser Thr lie Phe Leu Lys Ser Asn Ser Arg Phe lie Gly Gin Pro Leu 385 390 395 400 tgg ggt tet att gga tat act ttt cca geg get tta gga age caa att 1248

Trp Gly Ser lie Gly Tyr Thr Phe Pro Ala Ala Leu Gly Ser Gin lie 405 410 415 geg gat aaa gag age aga cac ett tta ttt att ggt gat ggt tea ett 1296

Ala Asp Lys Glu Ser Arg His Leu Leu Phe lie Gly Asp Gly Ser Leu 420 425 430 caa ett acc gta caa gaa tta gga eta tea ate aga ga gaa aaa etc aat 1344

Gin Leu Thr Val Gin Glu Leu Gly Leu Ser lie Arg Glu Lys Leu Asn 435 440 445 cca att tgt ttt ate ata aat aat gat ggt tat aca gtt gaa aga gaa 1392

Pro lie Cys Phe lie lie Asn Asn Asp Gly Tyr Thr Val Glu Arg Glu 450 455 460 ate cac gga cct act caa agt tat aac gac att cca atg tgg aat tac 1440 lie His Gly Pro Thr Gin Ser Tyr Asn Asp lie Pro Met Trp Asn Tyr 465 470 475 480 teg aaa tta cca gaa aca ttt gga gca aca gaa gat cgt gta gta tea 1488

Ser Lys Leu Pro Glu Thr Phe Gly Ala Thr Glu Asp Arg Val Val Ser 485 490 495 aaa att gtt aga aca gag aat gaa ttt gtg tet gtc atg aaa gaa gee 1536

Lys lie Val Arg Thr Glu Asn Glu Phe Val Ser Val Met Lys Glu Ala 500 505 510 caa gca gat gtc aat aga atg tat tgg ata gaa eta gtt ttg gaa aaa 1584

Gin Ala Asp Val Asn Arg Met Tyr Trp lie Glu Leu Val Leu Glu Lys 515 520 525 gaa gat geg cca aaa tta ctg aaa aaa atg ggt aaa tta ttt get gag 1632

Glu Asp Ala Pro Lys Leu Leu Lys Lys Met Gly Lys Leu Phe Ala Glu 530 535 540 caa aat aaa tag 1644

Gin Asn Lys 545 &lt;210&gt; 40 &lt;211&gt; 547

&lt;212&gt; PRT &lt;213&gt; Lactococcus lactis 69 201127961 &lt;400&gt; 40

Met Tyr Thr Val Gly Asp Tyr Leu Leu Asp Arg Leu His Glu Leu Gly 15 10 15 lie Glu Glu lie Phe Gly Val Pro Gly Asp Tyr Asn Leu Gin Phe Leu 20 25 30

Asp Gin lie lie Ser Arg Glu Asp Met Lys Trp lie Gly Asn Ala Asn 35 40 45

Glu Leu Asn Ala Ser Tyr Met Ala Asp Gly Tyr Ala Arg Thr Lys Lys 50 55 60

Ala Ala Ala Phe Leu Thr Thr Phe Gly Val Gly Glu Leu Ser Ala lie 65 70 75 80

Asn Gly Leu Ala Gly Ser Tyr Ala Glu Asn Leu Pro Val Val Glu lie 85 90 95

Val Gly Ser Pro Thr Ser Lys Val Gin Asn Asp Gly Lys Phe Val His 100 105 110

His Thr Leu Ala Asp Gly Asp Phe Lys His Phe Met Lys Met His Glu 115 120 125

Pro Val Thr Ala Ala Arg Thr Leu Leu Thr Ala Glu Asn Ala Thr Tyr 130 135 140

Glu lie Asp Arg Val Leu Ser Gin Leu Leu Lys Glu Arg Lys Pro Val 145 150 155 160

Tyr lie Asn Leu Pro Val Asp Val Ala Ala Ala Lys Ala Glu Lys Pro 165 170 175

Ala Leu Ser Leu Glu Lys Glu Ser Ser Thr Thr Asn Thr Thr Glu Gin 180 185 190

Val lie Leu Ser Lys lie Glu Glu Ser Leu Lys Asn Ala Gin Lys Pro 195 200 205

Val Val lie Ala Gly His Glu Val lie Ser Phe Gly Leu Glu Lys Thr 210 215 220

Val Thr Gin Phe Val Ser Glu Thr Lys Leu Pro He Thr Thr Leu Asn 70 201127961 225 230 235 240 Phe Gly Lys Ser Ala Val Asp Glu Ser Leu Pro Ser Phe Leu Gly lie 245 250 255

Tyr Asn Gly Lys Leu Ser Glu lie Ser Leu Lys Asn Phe Val Glu Ser 260 265 270

Ala Asp Phe lie Leu Met Leu Gly Val Lys Leu Thr Asp Ser Ser Thr 275 280 285

Gly Ala Phe Thr His His Leu Asp Glu Asn Lys Met lie Ser Leu Asn 290 295 300 lie Asp Glu Gly lie lie Phe Asn Lys Val Val Glu Asp Phe Asp Phe 305 310 315 320

Arg Ala Val Val Ser Ser Leu Ser Glu Leu Lys Gly lie Glu Tyr Glu 325 330 335

Gly Gin Tyr lie Asp Lys Gin Tyr Glu Glu Phe lie Pro Ser Ser Ala 340 345 350

Pro Leu Ser Gin Asp Arg Leu Trp Gin Ala Val Glu Ser Leu Thr Gin 355 360 365

Ser Asn Glu Thr lie Val Ala Glu Gin Gly Thr Ser Phe Phe Gly Ala 370 375 380

Ser Thr lie Phe Leu Lys Ser Asn Ser Arg Phe lie Gly Gin Pro Leu 385 390 395 400

Trp Gly Ser lie Gly Tyr Thr Phe Pro Ala Ala Leu Gly Ser Gin lie 405 410 415

Ala Asp Lys Glu Ser Arg His Leu Leu Phe lie Gly Asp Gly Ser Leu 420 425 430

Gin Leu Thr Val Gin Glu Leu Gly Leu Ser lie Arg Glu Lys Leu Asn 435 440 445

Pro He Cys Phe He He Asn Asn Asp Gly Tyr Thr Val Glu Arg Glu 450 455 460 71 201127961 lie His Gly Pro Thr Gin Ser Tyr Asn Asp lie Pro Met Trp Asn Tyr 465 470 475 480

Ser Lys Leu Pro Glu Thr Phe Gly Ala Thr Glu Asp Arg Val Val Ser 485 490 495

Lys lie Val Arg Thr Glu Asn Glu Phe Val Ser Val Met Lys Glu Ala 500 505 510

Gin Ala Asp Val Asn Arg Met Tyr Trp lie Glu Leu Val Leu Glu Lys 515 520 525

Glu Asp Ala Pro Lys Leu Leu Lys Lys Met Gly Lys Leu Phe Ala Glu 530 535 540

Gin Asn Lys 545 &lt;210&gt; 41 &lt;211&gt; 1644 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Branch ot-ketoacid decarboxylation of Lactococcus lactis enzyme KdcA codon-optimized gene &lt; 400 &gt; 41 atgtatactg ttggtgatta tctgctggac cgtctgcatg aactgggcat tgaagaaatc 60 ttcggtgtcc caggcgacta caacctgcag ttcctggacc agatcatctc ccgcgaagat 120 atgaaatgga tcggtaacgc aaacgagctg aacgcgtctt atatggctga tggttatgct 180 cgcaccaaaa aggctgcggc ctttctgacc acctttggtg tgggcgagct gagcgcgatc 240 aacggcctgg caggttccta cgctgagaac ctgccggtag tagaaatcgt tggttccccg 300 acctctaagg ttcagaacga cggcaaattc gtacatcaca ccctggcgga cggcgatttt 360 aagcacttta tgaaaatgca cgaaccggtc accgccgctc gcactctgct gaccgcggaa 420 aacgcaacgt acgagatcga tcgtgtactg tcccagctgc tgaaagaacg taaaccggtg 480 tatatcaatc tgccggttga tgtcgctgcg gccaaagcag agaaaccggc actgtccctg 540 gagaaggaga gctccactac taacaccacc gaacaggtta tcctgtccaa aattgaagaa 600 tctctgaaaa acgcacagaa accggtggtt atcgcaggtc acgaggttat ctccttcgg c 660 ctggagaaaa ctgttactca attcgtctct gaaacgaaac tgccgatcac gaccctgaac 720 tttggcaagt ccgcagttga cgaatctctg ccttctttcc tgggcattta caacggcaaa 780 72 840 201127961 ctgtccgaga tctccctgaa gaacttcgta gaatccgctg actttatcct gatgctgggt gtgaaactga ccgactcctc taccggtgcg ttcacgcacc atctggatga aaacaaaatg atcagcctga acatcgacga gggtatcatc ttcaacaagg tagttgaaga tttcgacttc cgtgctgttg tcagcagcct gtccgagctg aaaggcattg agtacgaggg tcaatacatc gataaacagt acgaagagtt tattccgtct tctgcaccgc tgagccagga ccgcctgtgg caggcagttg agtccctgac gcagtccaac gaaactatcg tagcggaaca aggtacctct ttcttcggtg cttctaccat ctttctgaag tccaactctc gctttatcgg tcagccgctg tggggttcta tcggttacac gttcccggct gcgctgggta gccagatcgc tgataaagag tctcgtcatc tgctgttcat cggtgatggt tccctgcagc tgactgtaca ggaactgggt ctgtctatcc gtgaaaaact gaacccgatt tgttttatca tcaataacga tggctacact gttgagcgtg aaattcatgg tccgactcag tcttacaacg atattccgat gtggaactac tctaaactgc cggaaacctt cggtgcaact gaggatcgcg tcgtgagcaa gattgtgcgt actgagaacg agttcgtatc tgttatgaaa gaggcgcagg Cagatgtgaa ccgcatgtac tggatcgaac tggttctgga aaaagaggat gcaccgaaac tgctgaagaa aatgggtaaa ctgtttgcgg agcagaacaa gtaa &lt;210&gt; 42 &lt;211&gt; 1647

&lt;212&gt; DNA &lt;213&gt; Lactococcus lactis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1) , . (1647) &lt;400&gt; 42 atg tat aca gta gga gat Tac eta tta gac ega tta cac gag tta gga

Met Tyr Thr Val Gly Asp Tyr Leu Leu Asp Arg Leu His Glu Leu Gly 15 10 15 att gaa gaa att ttt gga gtc act gga gac tat aac tta caa ttt tta lie Glu Glu lie Phe Gly Val Pro Gly Asp Tyr Asn Leu Gin Phe Leu 20 25 30 gat caa att att tcc cac aag gat atg aaa tgg gtc gga aat get aat

Asp Gin lie lie Ser His Lys Asp Met Lys Trp Val Gly Asn Ala Asn 35 40 45 gaa tta aat get tea tat atg get gat ggc tat get cgt act aaa aaa

Glu Leu Asn Ala Ser Tyr Met Ala Asp Gly Tyr Ala Arg Thr Lys Lys 50 55 60 get gcc gca ttt ett aca acc ttt gga gta ggt gaa ttg agt gca gtt 73 900 960 1020 1080 1140 1200 1260 1320 1380 1440 1500 1560 1620 1644 48 96 144 192 240 Lane 201127961

Ala Ala Ala Phe Leu Thr Thr Phe Gly Val Gly Glu Leu Ser Ala Val 65 70 75 80 aat gga tta gca gga agt tac gcc gaa aat tta cca gta gta gaa ata 288

Asn Gly Leu Ala Gly Ser Tyr Ala Glu Asn Leu Pro Val Val Glu lie 85 90 95 gtg gga tea cct aca tea aaa gtt caa aat gaa gga aaa ttt gtt cat 336

Val Gly Ser Pro Thr Ser Lys Val Gin Asn Glu Gly Lys Phe Val His 100 105 110 cat aeg ctg get gac ggt gat ttt aaa cac ttt atg aaa atg cac gaa 384

His Thr Leu Ala Asp Gly Asp Phe Lys His Phe Met Lys Met His Glu 115 120 125 cct gtt aca gca get ega act tta ctg aca gca gaa aat gca acc gtt 432

Pro Val Thr Ala Ala Arg Thr Leu Leu Thr Ala Glu Asn Ala Thr Val 130 135 140 gaa att gac ega gta ett tet gca eta tta aaa gaa aga aaa cct gtc 480

Glu lie Asp Arg Val Leu Ser Ala Leu Leu Lys Glu Arg Lys Pro Val 145 150 155 160 tat ate aac tta cca gtt gat gtt get get gca aaa gca gag aaa ccc 528

Tyr lie Asn Leu Pro Val Asp Val Ala Ala Ala Lys Ala Glu Lys Pro 165 170 175 tea etc cct ttg aaa aag gaa aac tea act tea aat aca agt gac caa 576

Ser Leu Pro Leu Lys Lys Glu Asn Ser Thr Ser Asn Thr Ser Asp Gin 180 185 190 gaa att ttg aac aaa att caa gaa age ttg aaa aat gcc aaa aaa cca 624

Glu lie Leu Asn Lys lie Gin Glu Ser Leu Lys Asn Ala Lys Lys Pro 195 200 205 ate gtg att aca gga cat gaa ata att agt ttt ggc tta gaa aaa aca 672

He Val He Thr Gly His Glu lie lie Ser Phe Gly Leu Glu Lys Thr 210 215 220 gtc act caa ttt att tea aag aca aaa eta cct att aeg aca tta aac 720

Val Thr Gin Phe lie Ser Lys Thr Lys Leu Pro lie Thr Thr Leu Asn 225 230 235 240 ttt ggt aaa agt tea gtt gat gaa gcc etc cct tea ttt tta gga ate 768

Phe Gly Lys Ser Ser Val Asp Glu Ala Leu Pro Ser Phe Leu Gly lie 245 250 255 tat aat ggt aca etc tea gag cct aat ett aaa gaa ttc gtg gaa tea 816

Tyr Asn Gly Thr Leu Ser Glu Pro Asn Leu Lys Glu Phe Val Glu Ser 260 265 270 gcc gac ttc ate ttg atg ett gga gtt aaa etc aca gac tet tea aca 864

Ala Asp Phe lie Leu Met Leu Gly Val Lys Leu Thr Asp Ser Ser Thr 275 280 285 gga gcc ttc act cat cat tta aat gaa aat aaa atg att tea ctg aat 912

Gly Ala Phe Thr His His Leu Asn Glu Asn Lys Met lie Ser Leu Asn 290 295 300 74 201127961 ata gat gaa gga aaa ata ttt aac gaa aga ate caa aat ttt gat ttt lie Asp Glu Gly Lys lie Phe Asn Glu Arg lie Gin Asn Phe Asp Phe 305 310 315 320 gaa tcc etc ate tcc tet etc tta gac eta age gaa ata gaa tac aaa Glu Ser Leu lie Ser Ser Leu Leu Asp Leu Ser Glu lie Glu Tyr Lys 325 330 335 gga aaa tat ate gat aaa aag caa Gaa gac ttt gtt cca tea aat geg Gly Lys Tyr lie Asp Lys Lys Gin Glu Asp Phe Val Pro Ser Asn Ala 340 345 350 ett tta tea caa gac ege eta tgg caa gca gtt gaa aac eta act caa Leu Leu Ser Gin Asp Arg Leu Trp Gin Ala Val Glu Asn Leu Thr Gin 355 360 365 age aat gaa aca ate gtt get gaa caa ggg aca tea ttc ttt ggc get Ser Asn Glu Thr lie Val Ala Glu Gin Gly Thr Ser Phe Phe Gly Ala 370 375 380 tea tea att Ttc tta aaa tea aag agt cat ttt att ggt caa ccc tta Ser Ser lie Phe Leu Lys Ser Lys Ser His Phe lie Gly Gin Pro Leu 385 390 395 400 99a tea att gga tat aca ttc cca gca gca tta gga age caa att Trp Gly Ser lie Gly Tyr Thr Phe Pro Ala Ala Leu Gly Ser Gin lie 405 410 415 gca gat aaa gaa age aga cac ett tta ttt att ggt gat ggt tea ett Ala Asp Lys Glu Ser Arg His Leu Leu Phe lie Gly Asp Gly Ser Leu 420 425 430 caa ett aca gtg caa gaa tta gga tta gca ate aga gaa aaa att aat Gin Leu Thr Val Gin Glu Leu Gly Leu Ala lie Arg Glu Lys lie Asn 435 440 445 cca att tgc ttt att ate aat aat gat ggt tat aca Gtc gaa aga gaa Pro lie Cys Phe lie lie Asn Asn Asp Gly Tyr Thr Val Glu Arg Glu 450 455 460 att cat gga cca aat caa age tac aat gat att cca atg tgg aat tac lie His Gly Pro Asn Gin Ser Tyr Asn Asp lie Pro Met Trp Asn Tyr 465 470 475 480 tea aaa tta cca gaa teg ttt gga gca aca gaa gat ega gta gtc tea Ser Lys Leu Pro Glu Ser Phe Gly Ala Thr Glu Asp Arg Val Val Ser 485 490 495 aaa ate gtt aga act gaa Aat gaa ttt gtg tet gtc atg aaa gaa get Lys lie Val Arg Thr Glu Asn Glu Phe Val Ser Val Met Lys Glu Ala 500 505 510 caa gca gat cca aat aga atg tac tg att gag tta att ttg gca aaa Gin Ala Asp Pro Asn Arg Met Tyr Trp He Glu Leu He Leu Ala Lys 515 520 525 gaa ggt gca cca aaa gta ctg aaa aaa atg ggc aaa eta ttt get gaa Glu Gly Ala Pro Lys Val Leu Lys Lys Met Gly Lys Leu Phe Ala Glu 530 535 540 9S0 1008 1056 1104 1152 1200 1248 1296 1344 1392 1440 1488 1536 1584 1632 75

S 201127961 1647 caa aat aaa tea taa Gin Asn Lys Ser 545 &lt;210&gt; 43 &lt;211&gt; 548

&lt;212&gt; PRT &lt;213&gt; Lactococcus lactis &lt;400&gt; 43

Met Tyr Thr Val Gly Asp Tyr Leu Leu Asp Arg Leu His Glu Leu Gly 15 10 15 lie Glu Glu lie Phe Gly Val Pro Gly Asp Tyr Asn Leu Gin Phe Leu 20 25 30

Asp Gin lie lie Ser His Lys Asp Met Lys Trp Val Gly Asn Ala Asn 35 40 45

Glu Leu Asn Ala Ser Tyr Met Ala Asp Gly Tyr Ala Arg Thr Lys Lys 50 55 60

Ala Ala Ala Phe Leu Thr Thr Phe Gly Val Gly Glu Leu Ser Ala Val 65 70 75 80

Asn Gly Leu Ala Gly Ser Tyr Ala Glu Asn Leu Pro Val Val Glu lie 85 90 95

Val Gly Ser Pro Thr Ser Lys Val Gin Asn Glu Gly Lys Phe Val His 100 105 110

His Thr Leu Ala Asp Gly Asp Phe Lys His Phe Met Lys Met His Glu 115 120 125

Pro Val Thr Ala Ala Arg Thr Leu Leu Thr Ala Glu Asn Ala Thr Val 130 135 140

Glu lie Asp Arg Val Leu Ser Ala Leu Leu Lys Glu Arg Lys Pro Val 145 150 155 160

Tyr lie Asn Leu Pro Val Asp Val Ala Ala Ala Lys Ala Glu Lys Pro 165 170 175

Ser Leu Pro Leu Lys Lys Glu Asn Ser Thr Ser Asn Thr Ser Asp Gin 180 185 190 76 201127961

Glu lie Leu Asn Lys lie Gin Glu Ser Leu Lys Asn Ala Lys Lys Pro 195 200 205 lie Val 210 lie Thr Gly His Glu lie lie Ser Phe Gly Leu Glu Lys Thr 215 220 Val Thr 225 Gin Phe lie Ser Lys Thr Lys Leu Pro Lie Thr Thr Leu Asn 230 235 240 Phe Gly Lys Ser Ser Val Asp Glu Ala Leu Pro Ser Phe Leu Gly lie 245 250 255 Tyr Asn Gly Thr Leu Ser Glu Pro Asn Leu Lys Glu Phe Val Glu Ser 260 265 270 Ala Asp Phe lie Leu Met Leu Gly Val Lys Leu Thr Asp Ser Ser Thr 275 280 285 Gly Ala 290 Phe Thr His His Leu Asn Glu Asn Lys Met lie Ser Leu Asn 295 300 lie Asp 305 Glu Gly Lys lie Phe Asn Glu Arg lie Gin Asn Phe Asp Phe 310 315 320 Glu Ser Leu lie Ser Ser Leu Leu Asp Leu Ser Glu lie Glu Tyr Lys 325 330 335 Gly Lys Tyr lie Asp Lys Lys Gin Glu Asp Phe Val Pro Ser Asn Ala 340 345 350 Leu Leu Ser Gin Asp Arg Leu Trp Gin Ala Val Glu Asn Leu Thr Gin 3S5 360 365 Ser Asn 3 70 Glu Thr lie Val Ala Glu Gin Gly Thr Ser Phe Phe Gly Ala 375 380 Ser Ser 385 lie Phe Leu Lys Ser Lys Ser His Phe lie Gly Gin Pro Leu 390 395 400 Trp Gly Ser lie Gly Tyr Thr Phe Pro Ala Ala Leu Gly Ser Gin lie 405 410 415 Ala Asp Lys Glu Ser Arg His Leu Leu Phe lie Gly Asp Gly Ser Leu 77 201127961 420 425 430

Gin Leu Thr Val Gin Glu Leu Gly Leu Ala lie Arg Glu Lys lie Asn 435 440 445

Pro lie Cys Phe lie lie Asn Asn Asp Gly Tyr Thr Val Glu Arg Glu 450 455 460 lie His Gly Pro Asn Gin Ser Tyr Asn Asp lie Pro Met Trp Asn Tyr 465 470 475 480

Ser Lys Leu Pro Glu Ser Phe Gly Ala Thr Glu Asp Arg Val Val Ser 485 490 495

Lys lie Val Arg Thr Glu Asn Glu Phe Val Ser Val Met Lys Glu Ala 500 505 510

Gin Ala Asp Pro Asn Arg Met Tyr Trp lie Glu Leu lie Leu Ala Lys 515 520 525

Glu Gly Ala Pro Lys Val Leu Lys Lys Met Gly Lys Leu Phe Ala Glu 530 535 540

Gin Asn Lys Ser 545 &lt;210&gt; 44 &lt;211&gt; 1647 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; ?-ketoisovalerate of Lactococcus lactis ester to carboxylase KivD codon-optimized gene &lt; 400 &gt; 44 atgtatactg ttggtgatta cctgctggat cgtctgcatg aactgggcat cgaggaaatt 60 ttcggcgtac ctggtgacta taacctgcag ttcctggatc agatcatttc ccacaaagat 120 atgaaatggg ttggtaacgc gaacgagctg aatgcaagct acatggctga cggttatgca 180 cgcaccaaga aagctgcggc gttcctgact acttttggcg tcggcgagct gtctgcggta 240 aacggtctgg ccggctccta cgcggaaaac ctgccggtag tagaaatcgt cggttccccg 300 acctctaaag ttcagaacga gggtaaattc gtgcaccata ctctggccga tggtgacttc 360 aaacacttca tgaagatgca cgaaccggtc actgctgctc gtacgctgct gaccgcggaa 420 78 201127961 aatgcgactg tacatcaacc aaaaaggaga tctctgaaga ctggagaaaa ttcggtaaat ctgtctgagc gtgaaactga atctctctga gaatccctga gataagaagc caagctgtgg ttcttcggtg tggggctcta tcccgccacc ctggcgattc gttgagcgtg tctaaactgc accgaaaacg tggatcgagc ctgttcgcag tcgagattga tgccggttga acagcacctc acgctaaaaa ctgtcaccca cctccgttga cgaacctgaa cggattcctc acattgatga tctcctccct aagaggactt aaaacctgac ctagctctat ttggctacac tgctgttcat gtgagaaaat agatccacgg cggaatcctt aattcgtgtc tgattctggc aacagaacaa tcgtgtactg tgtcgcggcc taacaccagc gccgatcgta gttcatcagc cgaagcgctg agagttcgtg caccggtgca gggcaaaatc gctggatctg cgtaccgtct ccagtccaac cttcctgaaa cttcccggca caacccgatt cccgaaccag cggtgcgact tgtcatgaaa taaagagggc atcctaa agcgcactgc gccaaagcgg gaccaggaaa atcaccggcc aaaaccaaac ccgtcctttc gagtctgcgg ttcacccacc ttcaacgagc tccgagattg aacgcgctgc tggtgacggc gaaaccatcg tctaaaagcc gcgctgggtt tctctgcaac tgtttcatca tcctacaacg gaagaccgtg gaagcacagg gcgccaaaag tgaaagaacg aaaaaccatc tcctgaacaa atgagattat tgccgatcac tgggtattta attttatcct acctgaatga gtattcagaa aatataaagg tgagccagga tggcggaaca acttcatcgg cccaaatcgc tgaccgtaca tcaacaacga acattccgat tcgtaagcaa cggacccgaa tactgaaaaa caagcctgta tctgccgctg gatccaggag ctctttcggt caccctgaac caacggcact gatgctgggc gaataaaatg cttcgatttc caaatacatt ccgtctgtgg gggtacctcc t Cagccactg agacaaagaa ggagctgggt tggctacact gtggaactac gatcgtccgt ccgcatgtac gatgggtaaa 480 540 600 660 720 780 840 900 960 1020 1080 1140 1200 1260 1320 1380 1440 1500 1560 1620 1647 &lt;210&gt; 45 &lt;211&gt; 3696

&lt;212&gt; DNA &lt;213&gt; Mycobacterium tuberculosis &lt;220&gt;&lt;221&gt; CDS &lt;222&gt; (1)..(3696) &lt;400&gt; 45 48 96 gtg gcc aac ata agt tea Cca ttc ggg caa aac gaa tgg ctg gtc gaa Val Ala Asn He Ser Ser Pro Phe Gly Gin Asn Glu Trp Leu Val Glu 15 10 15 gag atg tac ege aag ttc ege gac gac ccc tee teg gtc gat ccc age 79 201127961

Glu Met Tyr Arg Lys Phe 20 tgg cac gag ttc ctg gtt Trp His Glu Phe Leu Val 35 get gcc gaa cca acc egg Ala Ala Glu Pro Thr Arg 50 get geg gcc gcc ccg cag Ala Ala Ala Ala Pro Gin 65 70 9cg ggc aac Ggc gtg gtc Ala Gly Asn Gly Val Val 85 ccg cca gcc gaa ggt gac Pro Pro Ala Glu Gly Asp 100 gee gtc gtc aag aac atg Ala Val Val Lys Asn Met 115 age gtc egg geg gtc ccg Ser Val Arg Ala Val Pro 130 ate Aac aac cag ttg aag lie Asn Asn Gin Leu Lys 145 150 cat ttg ctg ggc tac gcc His Leu Leu Gly Tyr Ala 165 atg aac egg cac tac acc Met Asn Arg His Tyr Thr 180 ccg geg cac acc aat etc Pro Ala His Thr Asn Leu 195 ggg aag cgt tee ctg gtg Gly Lys Arg Ser Leu Val 210 ega ttc geg cag ttc gtc Arg Phe Ala Gin Phe Val 225 230 ege gac ggc aag ctg acc Arg Asp Gly Lys Leu Thr 245

Arg Asp Asp Pro Ser Ser 25 gac tac age ccc gaa ccc Asp Tyr Ser Pro Glu Pro 40 gtt acc teg cca etc gtt Val Thr Ser Pro Leu Val 55 60 gca ccc ccc aag ccg gcc Ala Pro Pro Lys Pro Ala 75 gcc gca ctg Gcc gcc aaa Ala Ala Leu Ala Ala Lys 90 gag gta geg gtg ctg ege Glu Val Ala Val Leu Arg 105 tee geg teg ttg gag gtg Ser Ala Ser Leu Glu Val 120 gcc aag eta ctg ate gac Ala Lys Leu Leu lie Asp 135 140 Egg acc ege ggc ggc aag Arg Thr Arg Gly Gly Lys 155 ctg gtg cag geg gtg aag Leu Val Gin Ala Val Lys 170 gaa gtc gac ggc aag ccc Glu Val Asp Gly Lys Pro 185 ggc ctg geg ate gac ctg Gly Leu Ala lie Asp Leu 200 gtg gcc ggc ate aag egg Val Ala Gly lie Lys Arg 215 220 aeg gcc tac gaa gac ate Thr Ala Tyr Glu Asp lie 235 act gaa gac ttt gcc ggc Thr Glu Asp Phe Ala Gly 250

Val Asp Pro Ser 30 acc tee caa cca 144

Thr Ser Gin Pro 45 gcc gag egg gcc 192

Ala Glu Arg Ala gac acc geg gcc 240

Asp Thr Ala Ala 80 act gcc gtt ccc 288

Thr Ala Val Pro 95 ggc gcc gcc geg 336

Gly Ala Ala Ala 110 ccg aeg geg acc 384

Pro Thr Ala Thr 125 aac egg ate gtc 432

Asn Arg lie Val ate teg ttc aeg 480 lie Ser Phe Thr 160 aaa ttc ccg aac 528

Lys Phe Pro Asn 175 acc geg gtc aeg 576

Thr Ala Val Thr 190 caa ggc aag gac 624

Gin Gly Lys Asp 205 tgc gag acc atg 672

Cys Glu Thr Met gta ege egg gcc 720

Val Arg Arg Ala 240 gtg aeg att teg 768

Val Thr lie Ser 255 80 201127961 ctg acc aat ccc gga acc ate ggc acc gtg cat teg gtg ccg egg ctg 816

Leu Thr Asn Pro Gly Thr lie Gly Thr Val His Ser Val Pro Arg Leu 260 265 270 atg ccc ggc cag ggc gee ate ate ggc gtg ggc gee atg gaa tac ccc 864

Met Pro Gly Gin Gly Ala lie lie Gly Val Gly Ala Met Glu Tyr Pro 275 280 285 gee gag ttt caa ggc gee age gag gaa ege ate gee gag ctg ggc ate 912

Ala Glu Phe Gin Gly Ala Ser Glu Glu Arg lie Ala Glu Leu Gly lie 290 295 300 ggc aaa ttg ate act ttg acc tee acc tac gac cac ege ate ate cag 960

Gly Lys Leu lie Thr Leu Thr Ser Thr Tyr Asp His Arg lie lie Gin 305 310 315 320 ggc geg gaa teg ggc gac ttc ctg ege acc ate cac gag ttg ctg etc 1008

Gly Ala Glu Ser Gly Asp Phe Leu Arg Thr lie His Glu Leu Leu Leu 325 330 335 teg gat ggc ttc tgg gac gag gtc ttc ege gaa ctg age ate cca tat 1056

Ser Asp Gly Phe Trp Asp Glu Val Phe Arg Glu Leu Ser lie Pro Tyr 340 345 350 ctg ccg gtg ege tgg age acc gac aac ccc gac teg ate gtc gac aag 1104

Leu Pro Val Arg Trp Ser Thr Asp Asn Pro Asp Ser lie Val Asp Lys 355 360 365 aac get ege gtc atg aac ttg ate geg gee tac ege aac ege ggc cat 1152

Asn Ala Arg Val Met Asn Leu lie Ala Ala Tyr Arg Asn Arg Gly His 370 375 380 ctg atg gee gat acc gac ccg ctg egg ttg gac aaa get egg ttc ege 1200

Leu Met Ala Asp Thr Asp Pro Leu Arg Leu Asp Lys Ala Arg Phe Arg 385 390 395 400 agt cac ccc gac etc gaa gtg ctg acc cac ggc ctg aeg ctg tgg gat 1248

Ser His Pro Asp Leu Glu Val Leu Thr His Gly Leu Thr Leu Trp Asp 405 410 415 etc gat egg gtg ttc aag gtc gac ggc ttt gee ggt geg cag tac aag 1296

Leu Asp Arg Val Phe Lys Val Asp Gly Phe Ala Gly Ala Gin Tyr Lys 420 425 430 aaa ctg ege gac gtg ctg ggc ttg ctg ege gat gee tac tgc ege cac 1344

Lys Leu Arg Asp Val Leu Gly Leu Leu Arg Asp Ala Tyr Cys Arg His 435 440 445 ate ggc gtg gag tac gee cat ate etc gac ccc gaa caa aag gag tgg 1392 lie Gly Val Glu Tyr Ala His lie Leu Asp Pro Glu Gin Lys Glu Trp 450 455 460 etc gaa caa egg gtc gag acc aag cac gtc aaa ccc act gtg gee caa 1440

Leu Glu Gin Arg Val Glu Thr Lys His Val Lys Pro Thr Val Ala Gin 465 470 475 480 cag aaa tac ate etc age aag etc aac gee gee gag gee ttt gaa aeg 1488

Gin Lys Tyr lie Leu Ser Lys Leu Asn Ala Ala Glu Ala Phe Glu Thr 485 490 495 81 201127961 ttc eta cag acc aag tac gtc ggc cag aag egg ttc teg ctg gaa ggc 1536

Phe Leu Gin Thr Lys Tyr Val Gly Gin Lys Arg Phe Ser Leu Glu Gly 500 505 510 gee gaa age gtg ate ccg atg atg gac geg ate gac cag tgc get 1584

Ala Glu Ser Val lie Pro Met Met Asp Ala Ala lie Asp Gin Cys Ala 515 520 525 gag cac ggc etc gac gag gtg gtc ate ggg atg ccg cac egg ggc egg 1632

Glu His Gly Leu Asp Glu Val Val lie Gly Met Pro His Arg Gly Arg 530 535 540 etc aac gtg ctg gee aac ate gtc ggc aag ccg tac teg cag ate ttc 1680

Leu Asn Val Leu Ala Asn lie Val Gly Lys Pro Tyr Ser Gin lie Phe 545 550 555 560 acc gag ttc gag ggc aac ctg aat ccg teg cag- geg cac ggc tee ggt 1728

Thr Glu Phe Glu Gly Asn Leu Asn Pro Ser Gin Ala His Gly Ser Gly 565 570 575 gac gtc aag tac cac ctg ggc gee acc ggg ctg tac ctg cag atg ttc 1776

Asp Val Lys Tyr His Leu Gly Ala Thr Gly Leu Tyr Leu Gin Met Phe 580 585 590 ggc gac aac gac att cag gtg teg ctg acc gee aac ccg teg cat ctg 1824

Gly Asp Asn Asp lie Gin Val Ser Leu Thr Ala Asn Pro Ser His Leu 595 600 605 gag gee gtc gac ccg gtg ctg gag gga ttg gtg egg gee aag cag gat 1872

Glu Ala Val Asp Pro Val Leu Glu Gly Leu Val Arg Ala Lys Gin Asp 610 615 620 ctg etc gac cac gga age ate gac age gac ggc caa egg geg ttc teg 1920

Leu Leu Asp His Gly Ser lie Asp Ser Asp Gly Gin Arg Ala Phe Ser 625 630 635 640 gtg gtg ccg ctg atg ttg cat ggc gat gee geg ttc gee ggt cag ggt 1968

Val Val Pro Leu Met Leu His Gly Asp Ala Ala Phe Ala Gly Gin Gly 645 650 655 gtg gtc gee gag aeg ctg aac ctg geg aat ctg ccg ggc tac ege gtc 2016

Val Val Ala Glu Thr Leu Asn Leu Ala Asn Leu Pro Gly Tyr Arg Val 660 665 670 ggc ggc acc ate cac ate ate gtc aac aac cag ate ggc ttc acc acc 2064

Gly Gly Thr lie His lie lie Val Asn Asn Gin lie Gly Phe Thr Thr 675 680 685 geg ccc gag tat tee agg tee age gag tac tgc acc gac gtc gca aag 2112

Ala Pro Glu Tyr Ser Arg Ser Ser Glu Tyr Cys Thr Asp Val Ala Lys 690 695 700 atg ate ggg gca ccg ate ttt cac gtc aac ggc gac gac ccg gag geg 2160

Met He Gly Ala Pro He Phe His Val Asn Gly Asp Asp Pro Glu Ala 705 710 715 720 tgt gtc tgg gtg geg egg ttg geg gtg gac ttc ega caa egg ttc aag 2208

Cys Val Trp Val Ala Arg Leu Ala Val Asp Phe Arg Gin Arg Phe Lys 82 201127961 725 730 735 aag Lys gac Asp gtc Val gtc Val 740 ate lie gac Asp atg Met ctg Leu tgc Cys 745 tac Tyr ege Arg ege Arg ege Arg ggg Gly 7S0 cac His aac Asn 2256 gag Glu ggt gac Gly Asp 755 gac Asp ccg Pro teg Ser atg Met acc Thr 760 aac Asn GCC Pro tac Tyr gtg Val tac Tyr 765 gac Asp gtc Val gtc Val 2304 gac Asp acc Thr 770 aag Lys Ege Arg ggg gcc ege Gly Ala Arg 775 aaa Lys age Ser tac Tyr acc Thr gaa Glu 780 gcc Ala ctg Leu ate lie gga Gly 2352 cgt Arg 785 ggc gac Gly Asp ate lie teg Ser atg Met 790 aag Lys gag Glu gcc Ala gag Glu gac Asp 795 gcg Ala ctg ege Leu Arg gac Asp tac Tyr 800 2400 cag ggc Gin Gly cag Gin ctg Leu gaa Glu 805 egg Arg gtg Val ttc Phe aac Asn gaa Glu 810 gtg Val ege Arg gag Glu ctg Leu gag Glu 815 aag Lys 2448 cac His ggt gtg Gly Val cag Gin 820 ccg Pro age Ser gag Glu teg Ser gtc Val 825 gag Glu tec Ser gac Asp cag Gin atg Met 830 att He ccc Pro 2496 gcg Ala ggg Gly ctg L Eu 835 gcc Ala act Thr gcg gtg Ala Val gac Asp 840 aag Lys teg Ser ctg Leu ctg Leu gcc Ala 845 egg Arg ate lie ggc Gly 2544 gat Asp gcg Ala 850 ttc Phe etc Leu gcc Ala ttg Leu ccg Pro 855 aac Asn ggc Gly ttc Phe acc Thr gcg Ala 860 cac His ccg Pro ega Arg gtc Val 2592 caa Gin 865 ccg Pro gtg Val ctg Leu gag Glu aag Lys 870 ege egg Arg Arg gag Glu atg Met gcc Ala 875 tat Tyr gaa Glu ggc aag Gly Lys Ate lie 880 2640 gac Asp tgg Trp gcc Ala ttt Phe ggc gag Gly Glu 885 ctg Leu ctg Leu gcg Ala ctg Leu 890 99C Gly teg Ser ctg Leu gtg Val gcc Ala 895 gaa Glu 2688 ggc aag Gly Lys ctg Leu gtg Val 900 ege Arg ttg Leu teg Ser ggg cag gac Gly Gin Asp 905 age Ser ege ege ggc Arg Arg Gly 910 acc Thr ttc Phe 2736 tcc Ser cag Gin egg Arg 915 cat His teg Ser gtt Val etc Leu ate He 920 gac Asp ege Arg cac His Act Thr ggc gag Gly Glu 925 gag Glu ttc Phe 2784 aca Thr cca Pro 930 ctg Leu cag Gin ctg Leu ctg Leu gcg Ala 935 acc Thr aac Asn tec Ser gac Asp ggc Gly 940 age Ser ccg Pro acc Thr ggc Gly 28 32 gga aag Gly Lys 945 ttc Phe ctg Leu gtc Val tac gac Tyr Asp 950 teg Ser cca Pro ctg Leu teg Ser 955 gag Glu tac gcc Tyr Ala gcc Ala gtc Val 960 2880 ggc ttc gag tac ggc tac act gtg ggc aat ccg gac Gcc gtg gtg etc 2928 83 201127961

Gly Phe Glu Tyr Gly Tyr Thr Val Gly Asn Pro Asp Ala Val Val Leu 965 970 975 tgg gag gcg cag ttc ggc gac ttc gtc aac ggc gcg cag teg ate ate 2976

Trp Glu Ala Gin Phe Gly Asp Phe Val Asn Gly Ala Gin Ser lie 980 985 990 gac gag ttc ate age tee ggt gag gee aag tgg ggc caa ttg tee aac 3024

Asp Glu Phe lie Ser Ser Gly Glu Ala Lys Trp Gly Gin Leu Ser Asn 995 1000 1005 gtc gtg ctg ctg tta ccg cac ggg cac gag ggg cag gga ccc gac 3069

Val Val Leu Leu Leu Pro His Gly His Glu Gly Gin Gly Pro Asp 1010 1015 1020 cac act tet gee egg ate gaa ege ttc ttg cag ttg tgg gcg gaa 3114

His Thr Ser Ala Arg lie Glu Arg Phe Leu Gin Leu Trp Ala Glu 1025 1030 1035 ggt teg atg acc ate gcg atg ccg teg act ccg teg aac tac ttc 3159

Gly Ser Met Thr lie Ala Met Pro Ser Thr Pro Ser Asn Tyr Phe 1040 1045 1050 cac ctg eta ege egg cat gee ctg gac ggc ate caa ege ccg ctg 3204

His Leu Leu Arg Arg His Ala Leu Asp Gly He Gin Arg Pro Leu 1055 1060 1065 ate gtg ttc aeg ccc aag teg atg ttg cgt cac aag gee gee gtc 3249 lie Val Phe Thr Pro Lys Ser Met Leu Arg His Lys Ala Ala Val 1070 1075 1080 age gaa ate aag gac ttc acc gag ate aag ttc ege tea gtg ctg 3294

Ser Glu lie Lys Asp Phe Thr Glu lie Lys Phe Arg Ser Val Leu 1085 1090 1095 gag gaa ccc acc tat gag gac ggc ate gga gac ege aac aag gtc 3339

Glu Glu Pro Thr Tyr Glu Asp Gly lie Gly Asp Arg Asn Lys Val 1100 1105 1110 age egg ate ctg ctg acc agt ggc aag ctg tat tac gag ctg gee 3384

Ser Arg lie Leu Leu Thr Ser Gly Lys Leu Tyr Tyr Glu Leu Ala 1115 1120 1125 gee ege aag gee aag gac aac ege aat gac etc gcg ate gtg egg 3429

Ala Arg Lys Ala Lys Asp Asn Arg Asn Asp Leu Ala lie Val Arg 1130 1135 1140 ett gaa cag etc gee ccg ctg ccc agg cgt ega ctg cgt gaa aeg 3474

Leu Glu Gin Leu Ala Pro Leu Pro Arg Arg Arg Leu Arg Glu Thr 1145 1150 1155 ctg gac ege tac gag aac gtc aag gag ttc ttc tgg gtc caa gag 3519

Leu Asp Arg Tyr Glu Asn Val Lys Glu Phe Phe Trp Val Gin Glu 1160 1165 1170 gaa ccg gee aac cag ggt gcg tgg ccg ega ttc ggg etc gaa eta 3564

Glu Pro Ala Asn Gin Gly Ala Trp pr〇Arg Phe Gly Leu Glu Leu 1175 1180 1185 84 201127961 ccc gag ctg ctg cct gac aag Pro Glu Leu Leu Pro Asp Lys 1190 1195 ttg gcc ggg ate aag Leu Ala Gly lie Lys 1200 ega ate Teg Arg lie Ser 3609 ege egg geg atg tea gcc ccg Arg Arg Ala Met Ser Ala Pro 1205 1210 teg tea ggc teg teg Ser Ser Gly Ser Ser 1215 aag gtg cac Lys Val His 3654 3696 gcc gtc gaa cag cag gag ate etc gac gag Geg ttc ggc tga Ala Val Glu Gin Gin Glu lie Leu Asp Glu Ala Phe Gly 1220 1225 1230 &lt;210&gt; 46 &lt;211&gt; 1231

&lt;212&gt; PRT &lt;213&gt; Mycobacterium tuberculosis &lt;400&gt; 46

Val Ala Asn lie Ser Ser Pro Phe Gly Gin Asn Glu Trp Leu Val Glu 15 10 15

Glu Met Tyr Arg Lys Phe Arg Asp Asp Pro Ser Ser Val Asp Pro Ser 20 25 30

Trp His Glu Phe Leu Val Asp Tyr Ser Pro Glu Pro Thr Ser Gin Pro 35 40 45

Ala Ala Glu Pro Thr Arg Val Thr Ser Pro Leu Val Ala Glu Arg Ala 50 55 60

Ala Ala Ala Ala Pro Gin Ala Pro Pro Lys Pro Ala Asp Thr Ala Ala 65 70 75 80

Ala Gly Asn Gly Val Val Ala Ala Leu Ala Ala Lys Thr Ala Val Pro 85 90 95

Pro Pro Ala Glu Gly Asp Glu Val Ala Val Leu Arg Gly Ala Ala Ala 100 105 110

Ala Val Val Lys Asn Met Ser Ala Ser Leu Glu Val Pro Thr Ala Thr 115 120 125

Ser Val Arg Ala Val Pro Ala Lys Leu Leu lie Asp Asn Arg lie Val 130 135 140

He Asn Asn Gin Leu Lys Arg Thr Arg Gly Gly Lys He Ser Phe Thr 145 150 155 160 85 201127961

His Leu Leu Gly Tyr Ala Leu Val Gin Ala Val Lys Lys Phe Pro Asn 165 170 175

Met Asn Arg His Tyr Thr Glu Val Asp Gly Lys Pro Thr Ala Val Thr 180 185 190

Pro Ala His Thr Asn Leu Gly Leu Ala He Asp Leu Gin Gly Lys Asp 195 200 205

Gly Lys Arg Ser Leu Val Val Ala Gly lie Lys Arg Cys Glu Thr Met 210 215 220

Arg Phe Ala Gin Phe Val Thr Ala Tyr Glu Asp lie Val Arg Arg Ala 225 230 235 240

Arg Asp Gly Lys Leu Thr Thr Glu Asp Phe Ala Gly Val Thr lie Ser 245 250 255

Leu Thr Asn Pro Gly Thr lie Gly Thr Val His Ser Val Pro Arg Leu 260 265 270

Met Pro Gly Gin Gly Ala lie lie Gly Val Gly Ala Met Glu Tyr Pro 275 280 285

Ala Glu Phe Gin Gly Ala Ser Glu Glu Arg lie Ala Glu Leu Gly lie 290 295 300

Gly Lys Leu lie Thr Leu Thr Ser Thr Tyr Asp His Arg lie lie Gin 305 310 315 320

Gly Ala Glu Ser Gly Asp Phe Leu Arg Thr lie His Glu Leu Leu Leu 325 330 335

Ser Asp Gly Phe Trp Asp Glu Val Phe Arg Glu Leu Ser lie Pro Tyr 340 345 350

Leu Pro Val Arg Trp Ser Thr Asp Asn Pro Asp Ser lie Val Asp Lys 355 360 365

Asn Ala Arg Val Met Asn Leu lie Ala Ala Tyr Arg Asn Arg Gly His 370 375 380

Leu Met Ala Asp Thr Asp Pro Leu Arg Leu Asp Lys Ala Arg Phe Arg 385 390 395 400 86 201127961

Ser His Pro Asp Leu Glu Val Leu Thr His Gly Leu Thr Leu Trp Asp 405 410 415

Leu Asp Arg Val Phe Lys Val Asp Gly Phe Ala Gly Ala Gin Tyr Lys 420 425 430

Lys Leu Arg Asp Val Leu Gly Leu Leu Arg Asp Ala Tyr Cys Arg His 435 440 445 lie Gly Val Glu Tyr Ala His lie Leu Asp Pro Glu Gin Lys Glu Trp 450 455 460

Leu Glu Gin Arg Val Glu Thr Lys His Val Lys Pro Thr Val Ala Gin 465 470 475 480

Gin Lys Tyr lie Leu Ser Lys Leu Asn Ala Ala Glu Ala Phe Glu Thr 485 490 495

Phe Leu Gin Thr Lys Tyr Val Gly Gin Lys Arg Phe Ser Leu Glu Gly 500 505 510

Ala Glu Ser Val lie Pro Met Met Asp Ala Ala lie Asp Gin Cys Ala 515 520 525

Glu His Gly Leu Asp Glu Val Val lie Gly Met Pro His Arg Gly Arg 530 535 540

Leu Asn Val Leu Ala Asn lie Val Gly Lys Pro Tyr Ser Gin lie Phe 545 550 555 560

Thr Glu Phe Glu Gly Asn Leu Asn Pro Ser Gin Ala His Gly Ser Gly 565 570 575

Asp Val Lys Tyr His Leu Gly Ala Thr Gly Leu Tyr Leu Gin Met Phe 580 585 590

Gly Asp Asn Asp lie Gin Val Ser Leu Thr Ala Asn Pro Ser His Leu 595 600 605

Glu Ala Val Asp Pro Val Leu Glu Gly Leu Val Arg Ala Lys Gin Asp 610 615 620

Leu Leu Asp His Gly Ser lie Asp Ser Asp Gly Gin Arg Ala Phe Ser e 87 201127961 625 630 635 640 Val Val Pro Leu Met Leu His Gly Asp Ala Ala Phe Ala Gly Gin Gly 645 650 655

Val Val Ala Glu Thr Leu Asn Leu Ala Asn Leu Pro Gly Tyr Arg Val 660 665 670

Gly Gly Thr lie His lie lie Val Asn Asn Gin lie Gly Phe Thr Thr 675 680 685

Ala Pro Glu Tyr Ser Arg Ser Ser Glu Tyr Cys Thr Asp Val Ala Lys 690 695 700

Met lie Gly Ala Pro He Phe His Val Asn Gly Asp Asp Pro Glu Ala 705 710 715 720

Cys Val Trp Val Ala Arg Leu Ala Val Asp Phe Arg Gin Arg Phe Lys 725 730 735

Lys Asp Val Val lie Asp Met Leu Cys Tyr Arg Arg Arg Gly His Asn 740 745 750

Glu Gly Asp Asp Pro Ser Met Thr Asn Pro Tyr Val Tyr Asp Val Val 755 760 765

Asp Thr Lys Arg Gly Ala Arg Lys Ser Tyr Thr Glu Ala Leu lie Gly 770 775 780

Arg Gly Asp worker le Ser Met Lys Glu Ala Glu Asp Ala Leu Arg Asp Tyr 785 790 795 800

Gin Gly Gin Leu Glu Arg Val Phe Asn Glu Val Arg Glu Leu Glu Lys 805 810 815

His Gly Val Gin Pro Ser Glu Ser Val Glu Ser Asp Gin Met lie Pro 820 825 830

Ala Gly Leu Ala Thr Ala Val Asp Lys Ser Leu Leu Ala Arg lie Gly 835 840 845

Asp Ala Phe Leu Ala Leu Pro Asn Gly Phe Thr Ala His Pro Arg Val 850 855 860 88 201127961

Gin Pro Val Leu Glu Lys Arg Arg Glu Met Ala Tyr Glu Gly Lys lie 865 870 875 880

Asp Trp Ala Phe Gly Glu Leu Leu Ala Leu Gly Ser Leu Val Ala Glu 885 890 895

Gly Lys Leu Val Arg Leu Ser Gly Gin Asp Ser Arg Arg Gly Thr Phe 900 905 910

Ser Gin Arg His Ser Val Leu lie Asp Arg His Thr Gly Glu Glu Phe 915 920 925

Thr Pro Leu Gin Leu Leu Ala Thr Asn Ser Asp Gly Ser Pro Thr Gly 930 935 940

Gly Lys Phe Leu Val Tyr Asp Ser Pro Leu Ser Glu Tyr Ala Ala Val 945 950 955 960

Gly Phe Glu Tyr Gly Tyr Thr Val Gly Asn Pro Asp Ala Val Val Leu 965 970 975

Trp Glu Ala Gin Phe Gly Asp Phe Val Asn Gly Ala Gin Ser lie lie 980 985 990

Asp Glu Phe lie Ser Ser Gly Glu Ala Lys Trp Gly Gin Leu Ser Asn 995 1000 1005

Val Val Leu Leu Leu Pro His Gly His Glu Gly Gin Gly Gly Pro Asp 1010 1015 1020

His Thr Ser Ala Arg lie Glu Arg Phe Leu Gin Leu Trp Ala Glu 1025 1030 1035

Gly Ser Met Thr lie Ala Met Pro Ser Thr Pro Ser Asn Tyr Phe 1040 1045 1050

His Leu Leu Arg Arg His Ala Leu Asp Gly lie Gin Arg Pro Leu 10SS 1060 1065 lie Val Phe Thr Pro Lys Ser Met Leu Arg His Lys Ala Ala Val 1070 1075 1080

Ser Glu lie Lys Asp Phe Thr Glu lie Lys Phe Arg Ser Val Leu 1085 1090 1095 e 89 201127961

Glu Glu 1100 Pro Thr Tyr Glu Asp 1105 Gly lie Gly Asp Arg 1110 Asn Lys Val Ser Arg 1115 lie Leu Leu Thr Ser 1120 Gly Lys Leu Tyr Tyr 1125 Glu Leu Ala Ala Arg 1130 Lys Ala Lys Asp Asn 1135 Arg Asn Asp Leu Ala 1140 lie Val Arg Leu Glu 1145 Gin Leu Ala Pro Leu 1150 Pro Arg Arg Arg Leu 1155 Arg Glu Thr Leu Asp 1160 Arg Tyr Glu Asn Val 1165 Lys Glu Phe Phe Trp 1170 Val Gin Glu Glu Pro 1175 Ala Asn Gin Gly Ala 1180 Trp Pro Arg Phe Gly 1185 Leu Glu Leu - Pro Glu 1190 Leu Leu Pro Asp Lys 1195 Leu Ala Gly lie Lys 1200 Arg lie Ser Arg Arg 1205 Ala Met Ser Ala Pro 1210 Ser Ser Gly Ser Ser 1215 Lys Val His Ala Val 1220 Glu Gin Gin Glu lie 1225 Leu Asp Glu Ala Phe 1230 Gly &lt;210&gt; 47 &lt;211&gt; 3696 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; CX- of Mycobacterium tuberculosis Methyl glutamate de-sustaining enzyme Kgd mazin optimized gene &lt;400&gt; 47 atggctaata tctcctctcc gtttggtcag aatgaatggc tggtagaaga aatgtaccgt 60 aaattccgcg atgacccgtc ctctgtgg ac ccgtcctggc atgaattcct ggtagactac 120 agcccggagc cgaccagcca accggcagcg gaaccaaccc gcgttacttc tccgctggta 180 gcggaacgtg cagctgctgc cgcgcctcag gcgccgccta aaccggcgga tactgccgca 240 gccggtaacg gtgtggtggc cgcactggct gctaagactg cggttccgcc gccagcagaa 300 ggcgatgaag ttgcagtcct gcgcggtgcg gcggctgcag tggtgaaaaa catgagcgcg 360 90 201127961 tccctggagg taccgaccgc cacgagcgtg cgcgcggtcc ctgctaaact gctgattgat 420 aaccgtattg tgatcaacaa ccagctgaaa cgtacccgtg gtggcaagat ctccttcact 480 catctgctgg gttatgcact ggtacaagcg gttaagaaat tccctaacat gaaccgtcat 540 tacactgagg tcgacggtaa accgacggct gttactccgg cacacacgaa cctgggcctg 600 gcgatcgacc tgcaaggtaa agatggtaag cgctccctgg tagttgcggg tattaaacgt 660 tgcgaaacca tgcgtttcgc acaattcgta accgcctacg aggacattgt ccgccgtgct 720 cgtgatggca aactgaccac cgaagatttt gcgggcgtta ctattagcct gaccaaccca 780 ggcaccatcg gcaccgtgca cagcgtacct cgtctgatgc cgggccaagg tgcgattatc 840 ggtgtgggtg ccatggagta cccggcagaa tttcagggtg cttctgaaga gcgcatcgcc 900 gagctgggta ttggtaaact Ga tcaccctg acttctacct atgaccaccg catcattcag 960 ggcgcagaat ccggtgactt cctgcgcact attcacgaac tgctgctgtc cgacggtttc 1020 tgggatgaag tttttcgtga actgagcatc ccatatctgc cagttcgctg gtccaccgac 1080 aatccggact ctatcgttga caaaaacgct cgcgtaatga acctgatcgc tgcttatcgt 1140 aatcgtggtc acctgatggc tgatacggat ccgctgcgcc tggataaagc tcgtttccgt 1200 tcccacccgg acctggaagt gctgacccat ggtctgactc tgtgggatct ggaccgcgtg 1260 ttcaaagtag atggtttcgc gggtgctcag tacaagaagc tgcgtgacgt gctgggtctg 1320 ctgcgtgatg cgtactgtcg tcacattggt gtggagtacg cccacattct ggatccggaa 1380 cagaaagaat ggctggagca gcgtgtcgag accaaacacg taaaaccgac cgtagcgcag 1440 cagaaatata tcctgtccaa actgaacgcc gccgaggctt cctgcagacc 1500 aagtacgtgg gccagaaacg cttcagcctg gagggtgcgg aaagcgttat tccgatgatg 1560 gatgcagcta tcgatcagtg cgcggaacat ggtctggatg aagtcgttat cggtatgccg 1620 caccgtggtc gcctgaacgt actggcaaac atcgtcggta aaccatattc tcagatcttc 1680 acggaattcg agggcaacct gaacccgtcc caagcccacg gctccggcga cgtaaaatat 1740 catctgggtg ctaccg tcgaaacttt gcct gtatctgcag atgttcggtg ataacgacat ccaggtatct 1800 ctgactgcta acccgagcca cctggaggcg gttgatcctg ttctggaagg tctggttcgc 1860 gccaaacagg atctgctgga ccacggctct atcgacagcg atggccagcg tgcattcagc 1920 gttgtaccgc tgatgctgca tggcgacgcg gcgttcgccg gtcagggtgt cgtagcagaa 1980 actctgaacc tggcgaacct gcctggctat cgcgtgggtg gcaccattca catcatcgtt 2040 aacaaccaaa tcggtttcac cacggcaccg gagtatagcc gttctagcga atattgcacc 2100 gacgtagcca aaatgatcgg tgcgccgatc ttccatgtaa acggtgacga tccagaggcc 2160 91 201127961 tgcgtgtggg tggctcgtct ggccgtagac ttccgccagc gttttaagaa agatgtggtt atcgacatgc tgtgctaccg ccgtcgtggt cacaacgaag gtgatgatcc gtctatgact aacccgtatg tctatgacgt ggtggacacc aagcgtggtg cacgcaaatc ttacacggag gccctgatcg gtcgtggcga catctctatg aaagaagcgg aagacgctct gcgtgattac cagggtcagc tggaacgtgt gttcaatgag gtgcgtgagc tggaaaagca cggcgtacaa ccgtccgaat ccgtagagtc cgatcagatg atccctgctg gtctggcaac tgctgttgat aaaagcctgc tggcgcgtat cggcgacgca ttcctggcgc tgccgaatgg ctttaccgcg cacccgcgcg tacagccggt actggaaaaa cgtcgt gaaa tggcctacga aggtaaaatc gattgggcct tcggtgagct gctggccctg ggctctctgg tggctgaggg caagctggta cgcctgagcg gccaggactc ccgtcgcggc actttttctc agcgtcacag cgtcctgatc gatcgtcaca ccggcgaaga attcacgccg ctgcaactgc tggctactaa ctccgatggt agcccgaccg gtggtaagtt cctggtgtac gattccccgc tgtccgaata tgctgcagtt ggtttcgagt atggttacac cgttggcaac ccggacgcag tggttctgtg ggaagcgcag ttcggcgatt tcgttaacgg tgcccagtcc attatcgatg agtttattag cagcggcgag gccaaatggg gccagctgtc taacgttgtg ctgctgctgc ctcacggcca cgagggtcaa ggcccggacc acacctccgc ccgtatcgaa cgcttcctgc agctgtgggc tgaaggctct atgaccatcg cgatgccgtc taccccaagc aactacttcc acctgctgcg tcgccacgca ctggacggca ttcagcgccc gctgatcgtt ttcaccccaa aatccatgct gcgccacaaa gcagctgttt ctgaaatcaa agattttacg gaaattaaat tccgttctgt gctggaagaa ccaacctacg aagacggtat tggcgaccgc actacgagct aacaaggtaa gccgtatcct gctgacctcc ggcaaactgt cgctacgaaa acgtaaaaga attcttctgg ggcagcacgt aaggcaaaag ataaccgcaa cgacctggcc atcgtccgcc tggaacagct ggcgccactg ccacgccgtc gcctgcgtga aaccctggat gtgcaggaag aaccg gcaaa ccagggtgcg tggccgcgct ttggtctgga actgccggaa ctgctgccgg ataaactggc aggtatcaag cgcatcagcc gtcgcgctat gagcgccccg tcttctggta gctctaaagt acacgctgta gaacagcaag agatcctgga tgaggccttc ggctaa &lt; 210 &gt; 48 &lt; 211 &gt; 74 &lt; 212 &gt; DNA &lt; 213 &gt; artificial sequence 2,220,228,023,402,400 2,460,252,025,802,640 2700 2760 2820 2880 2940 3000 3060 3120 3180 3240 3300 3360 3420 3480 3540 3600 3660 3696 92 201127961 &lt;220&gt;&lt;223&gt; Forward primer for amplifying the aminotransferase x of Bacillus subtilis &lt;400&gt; 9999This 039 ttgtacaaaa aagcaggcta ggaggaatta accatgaagg ttttagtcaa tggccggctg attg &lt;210&gt; 49 &lt;211&gt; 62 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; for amplifying the grass buds Bacmus subtilis) Aminotransferase x reverse primer &lt;400&gt; 49 ggggaccact ttgtacaaga aagctgggtt tatgaaatgc tagcagcctg ttgaatgctt tc &lt;210&gt; 50 &lt;211&gt; 82 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;;223&gt; for amplifying the grass buds (Bacillus subtilis} A forward transfer of aminotransferase y &lt;400&gt; 50 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgactc atgatttgat agaaaaaagt aaaaagcacc tc &lt;210&gt; 51 &lt;211&gt; 57 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;;220&gt;&lt;223&gt; for amplifying the aminotransferase of Bacillus subtilis. y reverse primer &lt;400&gt; 51 9999accact ttgtacaaga aagctgggtt caatcttcaa ggctcgtaac ctcgtgg &lt;210&gt; 52 &lt;211&gt; 64 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Forward primer for amplifying the aminotransferase of Rhodobacter sphaeroides &lt;400&gt; 52 201127961 99ggacaa9t ttgtacaaaa aagcaggcta ggaggaatta accatgcccg gttgcggggg 60 cttg 64 &lt;210&gt; 53 &lt;211&gt; 51 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; used to amplify the reverse primer of the aminotransferase of Rhodobacter sphaeroides &lt;400&gt; 53 ggggaccact ttgtacaaga aagctgggtt cagacggcgg ccggttcttt c 51 &lt;210&gt; 54 &lt;211&gt; 78 &lt;21 2&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Forward primer for amplifying the aminotransferase of Legionella pneumophila &lt;400&gt; 54 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgagta tcgcatttgt 60 Taacggcaag tattgttg 78 &lt;210&gt; 55 &lt;211&gt; 67 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;:223&gt; for amplifying the aminotransferase of Legionella pneumophila Reverse Inversion &lt;400&gt; 55 ggggaccact ttgtacaaga aagctgggtt tagtttacta gttgttggta ggaatcatta 60 attatcc 67 &lt;210&gt; 56 &lt;211&gt; 76 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Forward introduction of the aminotransferase of Nitrosomonas europaea &lt;400&gt; 56 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgattt acctcaatgg 60 caaatttctg ccgatg 76 94 201127961 &lt;210&gt; 57 &lt;211&gt; 50 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; for amplifying the European genus Nitroso The reverse primer of the aminotransferase of monas europaea &lt;400&gt; 57 ggggacc han tt ttgtacaag ugly aagctgggtt tactggcgtg gagcatgccc &lt;210&gt; 58 &lt;211&gt; 79 &lt;212&gt; DNA &lt;213&gt; artificial sequence &lt;220&gt;;223&gt; Forward primer for amplifying the aminotransferase of Neisseria gonorrhoeae &lt;400&gt; 58 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgagga taaatatgaa ¢0 ccgtaacgaa attttattc 79 &lt;210&gt; 59 &lt;211&gt; 56 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse arch 1 for amplifying the aminotransferase of Neisseria gonorrhoeae &lt;400&gt; 59 ggggaccact ttgtacaaga aagctgggtt catgcagcca tcgccttgaa cacttc &lt;210&gt; 60 &lt;211&gt; 66 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Forward primer for amplifying the aminotransferase of Pseudomonas aeruginosa &lt;400&gt; 60 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgtcga tggccgatcg tgatgg 56 60 66 &lt;210&gt; 61 &lt;211&gt; 53 &lt;212&gt; DNA &lt;213&gt; Artificial sequence 95 $ 201127961 &lt;220&gt;&lt;223&gt; Reverse primer for amplifying the aminotransferase of Pseudomonas aeruginosa &lt;400&gt; 61 ggggaccact ttgtacaaga aagctgggtt tacttgacca gggtacgcca etc 53 &lt;210&gt; 62 &lt;211&gt; 67 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; for magnification; Rhodopseudomonas Rhodopseudomonas palustris) A forward transfer of the aminotransferase &lt;400&gt; 62 ggggacaagt ttgtacaaaa aageaggeta ggaggaatta accatgaagc tgataccgtg 60 ccgcgcc 67 &lt;210&gt; 63 &lt;211&gt; 51 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer for amplifying the aminotransferase of Rhodopseudomonas palustris &lt;400&gt; 63 ggggaccact ttgtacaaga aagctgggtt caggcgaccg cgcggatcac c 51 &lt;210&gt; 64 &lt;211&gt; 1353 &lt;212&gt; DNA &lt;213&gt; Bacillus subtilis &lt;400&gt; 64 atggagatga tggggatgga aaacattcag caaaatcagg gattaaagea aaaagatg ag 60 caatttgtgt ggcatgccat gaagggagcg catcaagcgg acagcctgat agcccagaag 120 gccgaagggg cctgggtaac cgacacagac ggacgccgct atttggatgc gatgtccggt 180 ttgtggtgcg tcaacattgg ttaeggeaga aaggagcttg cggaggctgc ctatgagcaa 240 etaaaggage tgccttacta cccgttaacg caaagtcacg cacccgcaat tcaactggcg 300 gaaaagctga atgaatgget tggeggegat tatgttattt ttttttccaa cagcggatcg 360 gaagcaaacg aaactgcttt taaaattgcc cgccagtacc atctgcaaaa cggcgaccac 420 ageegttata aattcatctc aagatategg gcataccacg gcaatacatt gggagcgctc 480 tcagctaccg gacaggcgca geggaaatat aaataegage ctttgagcca agggttcctg 540 catgcagctc cgccagatat ataccggaat cctgatgatg cagacacgct tgaaagcgca 600 96 201127961 aatgaaatcg accgcatcat gacatgggaa ttaagcgaaa cgattgccgg ggtcattatg 660 gagcccatca ttacaggcgg aggcatccta atgccgccgg acggatatat gaagaaggtg 720 gaggacattt gccggcgcca cggagccctt ttgatttgcg atgaagtgat ctgcgggttt 780 ggacggacag gtgagccgtt cgggtttatg cactacggtg tgaagcctga tatcattacg 840 atggcaaagg gaatcacaag cgcgtatctg ccattgtcag cgactgctgt Gaa acgggac 900 attttcgaag cgtatcaggg ggaagctcct tatgaccgtt tccgccacgt gaacacgttc 960 ggcggaagcc cggctgcctg tgctttggcg ttgaaaaacc tgcaaattat ggaggacgaa 1020 cagctgattc agcgatcccg tgatcttgga gcaaagcttt taggtgagct 1080 agagaacacc cggcagtcgg ggatgttaga ggaaaagggc tgctgatcgg aatcgaactc 1140 gtcaaagaca aattgactaa agagccggct gatgccgcca aagtaaacca agtggttgcg 1200 gcgtgcaaag aaaaagggct gatcatcggc aaaaacggcg atacagtcgc cggctacaac 1260 aatgtcatcc acgttgcgcc gccattttgc ctgacagaag aggacctttc ctttatcgtg tcaagctctg 1320 aaaacggtga aagaaagctt tcaaacgata taa 1353 &lt;210&gt; 65 &lt;211&gt; 450

&lt;212&gt; PRT &lt;213&gt; Bacillus subtilis &lt;400&gt; 65

Met Glu Met Met Gly Met Glu Asn lie Gin Gin Asn Gin Gly Leu Lys 15 10 15

Gin Lys Asp Glu Gin Phe Val Trp His Ala Met Lys Gly Ala His Gin 20 25 30

Ala Asp Ser Leu lie Ala Gin Lys Ala Glu Gly Ala Trp Val Thr Asp 35 40 45

Thr Asp Gly Arg Arg Tyr Leu Asp Ala Met Ser Gly Leu Trp Cys Val 50 55 60

Asn lie Gly Tyr Gly Arg Lys Glu Leu Ala Glu Ala Ala Tyr Glu Gin 65 70 75 80

Leu Lys Glu Leu Pro Tyr Tyr Pro Leu Thr Gin Ser His Ala Pro Ala 85 90 95

S 97 201127961

He Gin Leu Ala Glu Lys Leu Asn Glu Trp Leu Gly Gly Asp Tyr Val 100 105 110 lie Phe Phe 115

Ser Asn Ser Gly Ser Glu Ala Asn Glu Thr Ala Phe Lys 120 125 lie Ala Arg Gin Tyr His Leu Gin Asn Gly Asp His Ser Arg Tyr Lys 130 135 140

Phe lie Ser Arg Tyr Arg Ala Tyr His Gly Asn Thr Leu Gly Ala Leu 145 150 155 160

Ser Ala Thr Gly Gin Ala Gin Arg Lys Tyr Lys Tyr Glu Pro Leu Ser 165 170 175

Gin Gly Phe Leu His Ala Ala Pro Pro Asp lie Tyr Arg Asn Pro Asp 180 185 190

Asp Ala Asp Thr Leu Glu Ser Ala Asn Glu lie Asp Arg lie Met Thr 195 200 205

Trp Glu Leu Ser Glu Thr lie Ala Gly Val lie Met Glu Pro lie lie 210 215 220

Thr Gly Gly Gly lie Leu Met Pro Pro Asp Gly Tyr Met Lys Lys Val 225 230 235 240

Glu Asp lie Cys Arg Arg His Gly Ala Leu Leu lie Cys Asp Glu Val 245 250 255 lie Cys Gly Phe Gly Arg Thr Gly Glu Pro Phe Gly Phe Met His Tyr 260 265 270

Gly Val Lys Pro Asp lie lie Thr Met Ala Lys Gly lie Thr Ser Ala 275 280 285

Tyr Leu Pro Leu Ser Ala Thr Ala Val Lys Arg Asp lie Phe Glu Ala 290 295 300

Tyr Gin Gly Glu Ala Pro Tyr Asp Arg Phe Arg His Val Asn Thr Phe 305 310 315 320

Gly Gly Ser Pro Ala Ala Cys Ala Leu Ala Leu Lys Asn Leu Gin lie 325 330 335 98 201127961

Met Glu Asp Glu Gin Leu lie Gin Arg Ser Arg Asp Leu Gly Ala Lys 340 345 350

Leu Leu Gly Glu Leu Gin Ala Leu Arg Glu His Pro Ala Val Gly Asp 355 360 365

Val Arg Gly Lys Gly Leu Leu lie Gly lie Glu Leu Val Lys Asp Lys 370 375 380

Leu Thr Lys Glu Pro Ala Asp Ala Ala Lys Val Asn Gin Val Val Ala 385 390 395 400

Ala Cys Lys Glu Lys Gly Leu lie lie Gly Lys Asn Gly Asp Thr Val 405 410 415

Ala Gly Tyr Asn Asn Val lie His Val Ala Pro Pro Phe Cys Leu Thr 420 425 430

Glu Glu Asp Leu Ser Phe lie Val Lys Thr Val Lys Glu Ser Phe Gin 435 440 445

Thr lie 450 &lt;210&gt; 66 &lt;211&gt; 1407

&Lt; 212 &gt; DNA &lt; 213 &gt; Pseudomonas aeruginosa (Pseudomonas aeruginosa) &lt; 400 &gt; 66 atgaacgcaa gactgcacgc cacgtccccc ctcggcgacg ccgacctggt ccgtgccgac 60 caggcccact acatgcacgg ctaccacgtg ttcgacgacc accgcgtcaa cggctcgctg 120 aacatcgccg ccggcgacgg cgcctatatc tacgacaccg ccggcaaccg ctacctcgac 180 gcggtgggcg gcatgtggtg caccaacatc ggcctggggc gcgaggaaat ggctcgcacc 240 gtggccgagc agacccgcct gctggcctat tccaatccct tctgcgacat ggccaacccg 300 cgcgccatcg aactctgccg caagctcgcc gagctggccc ccggcgacct cgaccacgtg 360 ttcctcacca ccggcggttc caccgccgtg gacaccgcga tccgcctcat gcactactac 420 cagaactgcc gcggcaagcg cgccaagaag cacgtcatca cgcggatcaa cgcctaccac 480 ggctcgacct tcctcggcat gtcgctgggc ggcaagagcg ccgaccggcc ggccgagttc 540 gacttcctcg acgagcgcat ccaccacctc gcctgtccct attactaccg cgctccggaa 600 99 660 201127961 gggctgggcg aagccgagtt cctcgatggc ctggtggacg Agttcgaacg caagatcctc gaactgggcg ccgaccgggt gggggcgttc atctccgagc cggtgttcgg ctccggcggc gtgatcgtcc cgcccgcggg ctaccacagg cggatgtggg agctgtgcca gcgctacga c gtgctgtaca tctccgacga agtggtgacc tccttcggcc gcctcggcca cttcttcgcc agccaggcgg tgttcggcgt acagccggac atcatcctca ccgccaaggg cctcacctcc ggctaccagc cgctgggcgc gtgcatcttc tcccggcgca tctgggaggt gatcgccgag ccggacaagg gccgctgctt cagccatggt ttcacctact ccggccaccc ggtggcctgc gcggcggcgc tgaagaacat cgagatcatc gagcgcgagg gcttgctcgc ccacgccgac gaggtcggcc gctacttcga ggagcgcctg caaagcctcc gcgacctgcc catcgtcggc gacgtgcgcg ggatgcgctt catggcctgt gtcgagttcg tcgccgacaa ggcgagcaag gcgctgtttc cggaaagcct gaacatcggc gagtgggtcc acctgcgggc gcagaagcgc ggcctgctgg Ttcgtccgat cgtccacctg aacgtgatgt cgccgccgct gatcctcacc cgcgaacagg tcgataccgt ggtccgggtg ctgcgcgaga gcatcgagga aaccgtggag gatcttgtcc gcgccggtca ccggtaa &lt;210&gt; 67 &lt;211&gt; 468

&lt;212&gt; PRT &lt;213&gt; Pseudomonas aeruginosa &lt;400&gt; 67

Met Asn Ala Arg Leu His Ala Thr Ser Pro Leu Gly Asp Ala Asp Leu 15 10 15

Val Arg Ala Asp Gin Ala His Tyr Met His Gly Tyr His Val Phe Asp 20 25 30

Asp His Arg Val Asn Gly Ser Leu Asn lie Ala Ala Gly Asp Gly Ala 35 40 45

Tyr He Tyr Asp Thr Ala Gly Asn Arg Tyr Leu Asp Ala Val Gly Gly 50 55 60

Met Trp Cys Thr Asn lie Gly Leu Gly Arg Glu Glu Met Ala Arg Thr 65 70 75 80

Val Ala Glu Gin Thr Arg Leu Leu Ala Tyr Ser Asn Pro Phe Cys Asp 85 90 95 720 780 840 900 960 1020 1080 1140 1200 1260 1320 1380 1407 100 201127961

Met Ala Asn Pro Arg Ala lie Glu Leu Cys Arg Lys Leu Ala Glu Leu 100 105 110

Ala Pro Gly Asp Leu Asp His Val Phe Leu Thr Thr Gly Gly Ser Thr 115 120 125

Ala Val Asp Thr Ala lie Arg Leu Met His Tyr Tyr Gin Asn Cys Arg 130 135 140

Gly Lys Arg Ala Lys Lys His Val lie Thr Arg lie Asn Ala Tyr His 145 150 155 160

Gly Ser Thr Phe Leu Gly Met Ser Leu Gly Gly Lys Ser Ala Asp Arg 165 170 175

Pro Ala Glu Phe Asp Phe Leu Asp Glu Arg lie His His Leu Ala Cys 180 185 190

Pro Tyr Tyr Tyr Arg Ala Pro Glu Gly Leu Gly Glu Ala Glu Phe Leu 195 200 205

Asp Gly Leu Val Asp Glu Phe Glu Arg Lys lie Leu Glu Leu Gly Ala 210 215 220

Asp Arg Val Gly Ala Phe lie Ser Glu Pro Val Phe Gly Ser Gly Gly 225 230 235 240

Val lie Val Pro Pro Ala Gly Tyr His Arg Arg Met Trp Glu Leu Cys 245 250 255

Gin Arg Tyr Asp Val Leu Tyr He Ser Asp Glu Val Val Thr Ser Phe 260 265 270

Gly Arg Leu Gly His Phe Phe Ala Ser Gin Ala Val Phe Gly Val Gin 275 280 285

Pro Asp lie lie Leu Thr Ala Lys Gly Leu Thr Ser Gly Tyr Gin Pro 290 295 300

Leu Gly Ala Cys He Phe Ser Arg Arg He Trp Glu Val He Ala Glu 305 310 315 320

Pro Asp Lys Gly Arg Cys Phe Ser His Gly Phe Thr Tyr Ser Gly His 325 330 335 101 201127961

Pro Val Ala Cys Ala Ala Ala Leu Lys Asn lie Glu lie lie Glu Arg 340 345 350

Glu Gly Leu Leu Ala His Ala Asp Glu Val Gly Arg Tyr Phe Glu Glu 355 360 365

Arg Leu Gin Ser Leu Arg Asp Leu Pro lie Val Gly Asp Val Arg Gly 370 375 380

Met Arg Phe Met Ala Cys Val Glu Phe Val Ala Asp Lys Ala Ser Lys 385 390 395 400

Ala Leu Phe Pro Glu Ser Leu Asn lie Gly Glu Trp Val His Leu Arg 405 410 415

His Leu Asn Val 430

Asp Thr Val Val 445

Ala Gin Lys Arg Gly Leu Leu Val Arg Pro lie Val 420 425

Met Ser Pro Pro Leu lie Leu Thr Arg Glu Gin Val 435 440

Arg Val Leu Arg Glu Ser lie Glu Glu Thr Val Glu Asp Leu Val Arg 450 455 460

Ala Gly His Arg 465 &lt;210&gt; 68 &lt;211&gt; 1335

&Lt; 212 &gt; DNA &lt; 213 &gt; Pseudomonas aeruginosa (Pseudomonas aeruginosa) &lt; 400 &gt; 68 atgacaatga atgacgagcc gcagtcgagc agcctcgaca acttctggat gcccttcacc 60 gccaaccgcc agttcaaggc gcggccgcgc ctgctggaaa gcgccgaagg catccactat 120 atcgcccagg gcgggcgccg catcctcgac ggcaccgccg gcctctggtg ctgcaatgcc 180 ggccacggcc ggcgcgagat cagcgaagcg gtggcccggc agatcgccac cctcgactac 240 gccccgccgt tccagatggg tcacccgctg ccgttcgaac tcgccgcgcg gctgacggaa 300 atcgccccgc cgagcctgaa caaagtattc ttcaccaact ccggctcgga atcggcggac 360 accgcgctga agatcgccct tgcctaccag cgcgccatcg gccagggcac ccgcacccgc 420 ctgatcggcc gcgaactggg ctaccacggg gtcggcttcg gcggcctgtc ggtaggcggt 480 102 201127961 atggtcaaca accgcaaggc cttctccgcc aacctgctgc cgggggtcga ccacctgccg 540 cacaccctgg acgtcgcccg caacgccttc accgtcggcc tgcccgagca tggcgtggaa 600 aaggccgagg agctggaacg cctggtgacc ctgcacggcg ccgagaatat Cgccgcggtg 660 atcgtcgagc cgatgtccgg ctcggccggc gtggtgctgc cgcccaaggg ctaccttcag 720 cggctgcggg agataacccg caagcatggc atcctgctga tcttcgacga agtgatcacc 780 ggtttcggcc gcgtcggcga agccttcgcc gcgcagcgct ggggcgtcgt cccggacctg 840 ctgacctgcg ccaaggggct gaccaacggc agcatcccga tgggcgccgt attcgtcgac 900 gagaagatcc atgctgcctt catgcaaggc ccgcagggcg ccatcgagtt cttccacggc 960 tatacctatt ccggccatcc ggtagcctgc gccgccgccc tggcgaccct ggacatctac 1020 cgtcgcgacg acctgttcca gcgggccgtc gaactggaag gctactggca ggacgcgctg 1080 ttcagcctgc gcgacctgcc caacgtggtc gacatccgcg ccgtaggcct ggtcggcggc 1140 gtgcaactgg cgccgcacgc ggacggcccc ggcaagcgcg gctacgacgt cttcgagcgc 1200 tgcttctggg agcacgacct gatggtccgg gtgaccggcg acatcatcgc catgtcgccg 1260 ccgctgatca tcgacaagcc ccacatcgac cagatcgtcg agcgcctggc ccaggccatc 1320 cgcgccagcg tctga 1335 &lt;210&gt; 69 &lt;211&gt; 444

&lt;212&gt; PRT &lt;213&gt; Pseudomonas aeruginosa &lt;400&gt; 69

Met Thr Met Asn Asp Glu Pro Gin Ser Ser Ser Leu Asp Asn Phe Trp 15 10 15

Met Pro Phe Thr Ala Asn Arg Gin Phe Lys Ala Arg Pro Arg Leu Leu 20 25 30

Glu Ser Ala Glu Gly lie His Tyr lie Ala Gin Gly Gly Arg Arg lie 35 40 45

Leu Asp Gly Thr Ala Gly Leu Trp Cys Cys Asn Ala Gly His Gly Arg 50 55 60

Arg Glu lie Ser Glu Ala Val Ala Arg Gin lie Ala Thr Leu Asp Tyr 65 70 75 80 103 201127961

Ala Pro Pro Phe Gin Met Gly His Pro Leu Pro Phe Glu Leu Ala Ala 85 90 95

Arg Leu Thr Glu lie Ala Pro Pro Ser Leu Asn Lys Val Phe Phe Thr 100 105 110

Asn Ser Gly Ser Glu Ser Ala Asp Thr Ala Leu Lys lie Ala Leu Ala 115 120 125

Tyr Gin Arg Ala lie Gly Gin Gly Thr Arg Thr Arg Leu lie Gly Arg 130 135 140

Glu Leu Gly Tyr His Gly Val Gly Phe Gly Gly Leu Ser Val Gly Gly 145 150 155 160

Met Val Asn Asn Arg Lys Ala Phe Ser Ala Asn Leu Leu Pro Gly Val 165 170 175

Asp His Leu Pro His Thr Leu Asp Val Ala Arg Asn Ala Phe Thr Val 180 185 190

Gly Leu Pro Glu His Gly Val Glu Lys Ala Glu Glu Leu Glu Arg Leu 195 200 205

Val Thr Leu His Gly Ala Glu Asn lie Ala Ala Val lie Val Glu Pro 210 215 220

Met Ser Gly Ser Ala Gly Val Val Leu Pro Pro Lys Gly Tyr Leu Gin 225 230 235 240

Arg Leu Arg Glu lie Thr Arg Lys His Gly lie Leu Leu lie Phe Asp 245 250 255

Glu Val lie Thr Gly Phe Gly Arg Val Gly Glu Ala Phe Ala Ala Gin 260 265 270

Arg Trp Gly Val Val Pro Asp Leu Leu Thr Cys Ala Lys Gly Leu Thr 275 280 285

Asn Gly Ser lie Pro Met Gly Ala Val Phe Val Asp Glu Lys lie His 290 295 300

Ala Ala Phe Met Gin Gly Pro Gin Gly Ala He Glu Phe Phe His Gly 305 310 315 320 104 201127961

Tyr Thr Tyr Ser Gly His Pro Val Ala Cys Ala Ala Ala Leu Ala Thr 325 330 335

Leu Asp lie Tyr Arg Arg Asp Asp Leu Phe Gin Arg Ala Val Glu Leu 340 345 350

Glu Gly Tyr Trp Gin Asp Ala Leu Phe Ser Leu Arg Asp Leu Pro Asn 355 360 365

Val Val Asp He Arg Ala Val Gly Leu Val Gly Gly Val Gin Leu Ala 370 375 380

Pro His Ala Asp Gly Pro Gly Lys Arg Gly Tyr Asp Val Phe Glu Arg 385 390 395 400

Cys Phe Trp Glu His Asp Leu Met Val Arg Val Thr Gly Asp lie lie 405 410 415

Ala Met Ser Pro Pro Leu lie lie Asp Lys Pro His lie Asp Gin lie 420 425 430

Val Glu Arg Leu Ala Gin Ala lie Arg Ala Ser Val 435 440 &lt;210&gt; 70 &lt;211&gt; 71 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; for amplifying Bacillus subtilis ( The forward primer of the aminotransferase of Bacillus subtilis (gil6〇7799l) &lt;400&gt; 70 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatggaga tgatggggat 60 ggaaaacatt c 71 &lt;210&gt; 71 &lt;211&gt; 65 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer for amplifying the aminotransferase of Bacillus subtilis (gil6〇77&quot;l)&lt;400&gt; 71 ggggaccact ttgtacaaga aagctgggtt tatatcgttt gaaagctttc tttcaccgtt 60 65 ttcac 105 201127961 &lt;210&gt; 72 &lt;211&gt; 66 &lt;212&gt; DNA &lt;;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; For amplifying the aminotransferase of Pseudomonas aeruginosa ( Gi&quot;51〇72) Forward introduction &lt;400&gt; 72 ggggacaagt ttgtacaaaa aagcaggcta ggaggaatta accatgaacg caagactgca 60 cgccac 66 &lt;210&gt; 73 &lt;211&gt; 48 &lt;212&gt; D NA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer for amplifying the aminotransferase of Pseudomonas aeruginosa (gi&quot;5l〇72) &lt;400&gt; 73 9999accact ttgtacaaga aagctgggtt Taccggtgac cggcgcgg 48 &lt;210&gt; 74 &lt;211&gt; 69 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; for amplifying the aminotransferase of Pseudomonas aeruginosa ( gi99Si63〇&gt; Forward introduction &lt;400&gt; 74 9999acaagt ttgtacaaaa aagcaggcta ggaggaatta accatgacaa tgaatgacga 6〇gccgcagtc 69 &lt;210&gt; 75 &lt;211&gt; 49 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer for amplifying the aminotransferase (gi995163〇) of Pseudomonas aeruginosa &lt;400&gt; 75 9999accact ttgtacaaga aagctgggtt cagacgctgg cgcggatgg 49 106 201127961 &lt;210&gt;&lt;211&gt;&lt;212&gt;&lt;213&gt; 76 57 DNA artificial sequence &lt;220&gt;&lt;223&gt; Forward introduction &lt;400&gt; 76 aaatttacta gtaagaattt ttgaggaggc aatataaatg aataaaccac Agtcttg &lt;210&gt;&lt;211&gt;&lt;212&gt;&lt;213&gt; 77 32 DNA artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer &lt;400&gt; 77 aaatttggat cctacaagaa agctgggttt ac &lt;210&gt;&lt;211&gt;&lt;;212&gt;&lt;213&gt; 78 58 DNA artificial sequence &lt;220&gt;&lt;223&gt; Forward introduction &lt;400&gt; 78 aaatttacta gtaagaattt ttgaggaggc aatataaatg aacagccaaa tcaccaac &lt;210&gt;&lt;211&gt;&lt;212&gt;&lt;213&gt; 7 9 37 DNA artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer &lt;400&gt; 79 aaatttggat ccactttgta caagaaagct gggttca &lt;210&gt;&lt;211&gt;&lt;212&gt;&lt;213&gt; 80 57 DNA artificial sequence &lt;220&gt;&lt;223&gt; Forward introduction &lt;400&gt; 80 aaatttggat ccgttgagga ggcctcaaaa atgtccgaga tcactctggg caaatac 201127961 &lt;210&gt; 81 &lt;211&gt; 35 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse primer &lt;400&gt; 81 aaatttggcg cgccattact gtttagcgtt agttg 35 &lt;210&gt; 82 &lt;211&gt; 52 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Forward introduction &lt;400&gt; 82 aaatttggat ccgttgagga ggcctcaaaa atgtatactg ttggtgatta tc 52 &lt;210&gt; 83 &lt;211&gt; 37 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Reverse introduction &lt;400&gt; 83 aaatttggcg cgccattact Tgttctgctc cgcaaac 37 &lt;210&gt; 84 &lt;211&gt; 1113 &lt;212&gt; DNA &lt;213 &gt; Artificial sequence &lt;220&gt;&lt;223&gt; Human (Homo sapiens) residue acid oxidase (glycolate oxidase) (HAOX-5B> &lt; 400 &gt; 84 atgctccccc ggctaatttg tatcaatgat tatgaacaac atgctaaatc agtacttcca 60 aagtctatat atgactatta caggtctggg gcaaatgatg aagaaacttt ggctgataat 120 attgcagcat tttccagatg gaagctgtat ccaaggatgc tccggaatgt tgctgaaaca 180 gatctgtcga cttctgtttt aggacagagg gtcagcatgc caatatgtgt gggggctacg 240 gccatgcagc gcatggctca tgtggacggc gagcttgcca ctgtgagagc ctgtcagtcc 300 ctgggaacgg gcatgatgtt gagttcctgg gccacctcct caattgaaga Agtggcggaa 360 108 201127961 gctggtcctg aggcacttcg ttggctgcaa ctgtatatct acaaggaccg agaagtcacc 420 aagaagctag tgcggcaggc agagaagatg ggctacaagg ccatattt gt gacagtggac 480 acaccttacc tgggcaaccg tctggatgat gtgcgtaaca gattcaaact gccgccacaa 540 ctcaggatga aaaattttga aaccagtact ttatcatttt ctcctgagga aaattttgga 600 gacgacagtg gacttgctgc atatgtggct aaagcaatag acccatctat cagctgggaa 660 gatatcaaat ggctgagaag actgacatca ttgccaattg ttgcaaaggg cattttgaga 720 ggtgatgatg ccagggaggc tgttaaacat ggcttgaatg ggatcttggt gtcgaatcat 780 ggggctcgac aactcgatgg ggtgccagcc actattgatg ttctgccaga aattgtggag 840 gctgtggaag ggaaggtgga agtcttcctg gacgggggtg tgcggaaagg cactgatgtt 900 ctgaaagctc tggctcttgg cgccaaggct gtgtttgtgg ggagaccaat cgtttggggc 960 ttagctttcc agggggagaa aggtgttcaa gatgtcctcg agatactaaa ggaagaattc 1020 cggttggcca tggctctgag tgggtgccag aatgtgaaag tcatcgacaa gacattggtg 1080 aggaaaaatc ctttggccgt ttccaagatc tga 1113 &lt; 210 &gt; 85 &lt; 211 &gt; 370

&lt;212&gt; PRT &lt;213&gt; HA0X1_human basal acid oxidase 1 - Homo sapiens (human) &lt;400&gt; 85

Met Leu Pro Arg Leu lie Cys lie Asn Asp Tyr Glu Gin His Ala Lys 1 5 10 15 .

Ser Val Leu Pro Lys Ser He Tyr Asp Tyr Tyr Arg Ser Gly Ala Asn 20 25 30

Asp Glu Glu Thr Leu Ala Asp Asn lie Ala Ala Phe Ser Arg Trp Lys 35 40 45

Leu Tyr Pro Arg Met Leu Arg Asn Val Ala Glu Thr Asp Leu Ser Thr 50 55 60

Ser Val Leu Gly Gin Arg Val Ser Met Pro lie Cys Val Gly Ala Thr 65 70 75 80

Ala Met Gin Arg Met Ala His Val Asp Gly Glu Leu Ala Thr Val Arg 85 90 95 109 £ 201127961

Ala Cys Gin Ser Leu Gly Thr Gly Met Met Leu Ser Ser Trp Ala Thr 100 105 110

Ser Ser lie Glu Glu Val Ala Glu Ala Gly Pro Glu Ala Leu Arg Trp 115 120 125

Leu Gin Leu Tyr lie Tyr Lys Asp Arg Glu Val Thr Lys Lys Leu Val 130 135 140

Arg Gin Ala Glu Lys Met Gly Tyr Lys Ala lie Phe Val Thr Val Asp 145 150 155 160

Thr Pro Tyr Leu Gly Asn Arg Leu Asp Asp Val Arg Asn Arg Phe Lys 165 170 175

Leu Pro Pro Gin Leu Arg Met Lys Asn Phe Glu Thr Ser Thr Leu Ser 180 185 190

Phe Ser Pro Glu Glu Asn Phe Gly Asp Asp Ser Gly Leu Ala Ala Tyr 195 200 205

Val Ala Lys Ala lie Asp Pro Ser lie Ser Trp Glu Asp lie Lys Trp 210 215 220

Leu Arg Arg Leu Thr Ser Leu Pro lie Val Ala Lys Gly lie Leu Arg 225 230 235 240

Gly Asp Asp Ala Arg Glu Ala Val Lys His Gly Leu Asn Gly lie Leu 245 250 255

Val Ser Asn His Gly Ala Arg Gin Leu Asp Gly Val Pro Ala Thr lie 260 265 270

Asp Val Leu Pro Glu lie Val Glu Ala Val Glu Gly Lys Val Glu Val 275 280 285

Phe Leu Asp Gly Gly Val Arg Lys Gly Thr Asp Val Leu Lys Ala Leu 290 295 300

Ala Leu Gly Ala Lys Ala Val Phe Val Gly Arg Pro lie Val Trp Gly 305 310 315 320

Leu Ala Phe Gin Gly Glu Lys Gly Val Gin Asp Val Leu Glu lie Leu 325 330 335 110 201127961

Lys Glu Glu Phe Arg Leu Ala Met Ala Leu Ser Gly Cys Gin Asn Val 340 345 350

Lys Val lie Asp Lys Thr Leu Val Arg Lys Asn Pro Leu Ala Val Ser 355 360 365

Lys lie 370 &lt;210&gt; 86 &lt;211&gt; 1113 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthetic DNA (human (Homo sapiens) hydroxyl oxidase (glycolate oxidase) (HAOX-5B), it codon optimized) &lt; 400 &gt; 86 atgctgccac gtctgatttg tattaacgat tacgaacaac acgcgaagag cgtactgccg 60 aaatccattt acgattatta ccgttctggt gcaaacgatg aagaaacgct ggctgataac 120 atcgccgctt tttcccgttg gaaactgtac ccacgtatgc tgcgtaacgt tgccgaaacc 180 gacctgtcca ccagcgtcct gggtcagcgt gtgtccatgc caatctgcgt gggtgcaacc 240 gcaatgeage gtatggcaca cgttgacggc gaactggcaa ccgtccgtgc gtgccagagc 300 ctgggtaccg gtatgatgct gagcagctgg gctacctcta gcatcgagga agtggcagaa 360 gctggtccgg aagcactgcg ctggctgcag ctgtacatct acaaagatcg cgaagtcact 420 aagaaactgg tgcgccaggc ggaaaagatg ggttacaagg caatctttgt gactgttgac 480 accccgtacc tgggtaaccg cctggatgac gttcgtaacc gcttcaagct gccgccgcag 540 ctgcgtatga agaactttga aaccagcacc ctgtcctttt ccccagaaga aaatttcggt 600 gatgactctg gtctggccgc gtacgtcgcg aaagctatcg atccgtcc at ctcctgggaa 660 gatatcaaat ggctgcgtcg tctgacttcc ctgccgatcg ttgctaaggg tattctgcgt 720 ggtgacgacg cgcgtgaagc tgttaaacat ggtctgaacg gcattctggt aagcaaccat 780 ggcgcacgcc agctggatgg tgtacctgct actattgatg tgctgccgga aatcgtggaa 840 gcggttgaag gtaaagttga agtgttcctg gacggtggtg tgcgcaaagg caccgatgta 900 ctgaaagcac tggcgctggg tgcgaaagcc gtctttgttg gccgtcctat tgtttggggt 960 ctggcattcc agggtgagaa aggtgtacag gacgttctgg agatcctgaa a9aggagttc 1020 cgcctggcta tggcgctgtc tggttgtcaa aacgtgaaag taatcgataa Acacctggta 1080 111 201127961 cgtaaaaacc ctctggcagt aagcaagatc taa 1113 &lt;210&gt; 87 &lt;211&gt; 1125 &lt;212&gt; DNA &lt;213 &gt; Artificial sequence &lt;220&gt;&lt;223&gt; Lactate oxidase-Aerococcus viridans (wt) (LAOX-8C) &lt;400&gt; 87 atgaataaca atgacattga atataatgca cctagtgaaa tcaagtacat tgatgttgtc 60 aatacttacg acttagaaga agaagcaagt aaagtggt cacatggtgg ttttaactat 120 attgccggtg catctggtga tgagtggact aaacgcgcta atgaccgtgc ttggaaacat 180 aaattactat acccacgtct agcgcaagat gtt gaagcgc ccgatacaag tactgaaatt 240 ttaggtcata aaattaaagc cccattcatc atggcaccaa ttgctgcaca tggtttagcc 300 cacactacta aagaagctgg tactgcacgt gcagtttcag aatttggtac aattatgtcc 360 atctcagctt attctggtgc aacatttgaa gaaatttctg aaggcttaaa tggcggaccc 420 cgttggttcc aaatctatat ggctaaagat gaccaacaaa accgtgatat cttagacgaa 480 gctaaatctg atggtgcaac tgctatcatc cttacagctg actcaactgt ttctggaaac 540 cgtgaccgtg atgtgaagaa taaattcgtt tacccatttg gtatgccaat tgttcaacgt 600 tacttacgtg gtacagcaga aggtatgtca ttaaacaata tctacggtgc ttcaaaacaa 660 aaaatctcac caagagatat tgaggaaatc gccggtcatt ctggattacc agtattcgtt 720 aaaggtattc aacacccaga agatgcagat atggcaatca aacgtggtgc atcaggtatc 780 tgggtatcta accacggtgc tcgtcaacta tatgaagctc caggttcatt tgacaccctt 840 ccagctattg ctgaacgtgt aaacaaacgt gtaccaatcg tctttgattc aggtgtacgt 900 cgtggtgaac acgttgccaa agcgctagct tcaggggcag acgttgttgc tttaggacgc 960 ccagtcttat ttggtttagc tttaggtggc tggcaaggtg cttactcagt acttgactac 1020 ttccaaaaag acttaacacg cgtaatgcaa ttaacaggtt cacaaaatgtGgaagacttg 1080 atttattcga taacccatac ggttatgaat actag 1125 &lt;210&gt; 88 &lt;211&gt; 374

&lt;212&gt; PRT &lt;213&gt; LAOX-8C lactate oxidase-green balloon ij (Aerococcus viridans) &lt;400&gt; 88

Met Asn Asn Asn Asp lie Glu Tyr Asn Ala Pro Ser Glu lie Lys Tyr 112 201127961 15 10 15 lie Asp Val Val Asn Thr Tyr Asp Leu Glu Glu Glu Ala Ser Lys Val 20 25 30

Val Pro His Gly Gly Phe Asn Tyr lie Ala Gly Ala Ser Gly Asp Glu 35 40 45

Trp Thr Lys Arg Ala Asn Asp Arg Ala Trp Lys His Lys Leu Leu Tyr 50 55 60

Pro Arg Leu Ala Gin Asp Val Glu Ala Pro Asp Thr Ser Thr Glu lie 65 70 75 80

Leu Gly His Lys lie Lys Ala Pro Phe lie Met Ala Pro lie Ala Ala 85 90 95

His Gly Leu Ala His Thr Thr Lys Glu Ala Gly Thr Ala Arg Ala Val 100 105 110

Ser Glu Phe Gly Thr lie Met Ser lie Ser Ala Tyr Ser Gly Ala Thr 115 120 125

Phe Glu Glu lie Ser Glu Gly Leu Asn Gly Gly Pro Arg Trp Phe Gin 130 135 140 lie Tyr Met Ala Lys Asp Asp Gin Gin Asn Arg Asp lie Leu Asp Glu 145 150 155 160

Ala Lys Ser Asp Gly Ala Thr Ala lie lie Leu Thr Ala Asp Ser Thr 165 170 175

Val Ser Gly Asn Arg Asp Arg Asp Val Lys Asn Lys Phe Val Tyr Pro 180 185 190

Phe Gly Met Pro lie Val Gin Arg Tyr Leu Arg Gly Thr Ala Glu Gly 195 200 205

Met Ser Leu Asn Asn lie Tyr Gly Ala Ser Lys Gin Lys lie Ser Pro 210 215 220

Arg Asp lie Glu Glu lie Ala Gly His Ser Gly Leu Pro Val Phe Val 225 230 235 240 113 201127961

Lys Gly lie Gin His Pro Glu Asp Ala Asp Met Ala lie Lys Arg Gly 245 250 255

Ala Ser Gly lie Trp Val Ser Asn His Gly Ala Arg Gin Leu Tyr Glu 260 265 270

Ala Pro Gly Ser Phe Asp Thr Leu Pro Ala lie Ala Glu Arg Val Asn 275 280 285

Lys Arg Val Pro lie Val Phe Asp Ser Gly Val Arg Arg Gly Glu His 290 295 300

Val Ala Lys Ala Leu Ala Ser Gly Ala Asp Val Val Ala Leu Gly Arg 305 310 315 320

Pro Val Leu Phe Gly Leu Ala Leu Gly Gly Trp Gin Gly Ala Tyr Ser 325 330 335

Val Leu Asp Tyr Phe Gin Lys Asp Leu Thr Arg Val Met Gin Leu Thr 340 345 350

Gly Ser Gin Asn Val Glu Asp Leu Lys Gly Leu Asp Leu Phe Asp Asn 355 360 365

&lt;210&gt;&lt;211&gt; 400 &gt; 89 60 120 180 240 300 360 atgaacaaca acgacatcga atataacgct ccttctgaaa tcaaatatat cgacgtggtt aacacctatg acctggagga agaagcgtct aaggtcgtac cgcacggtgg tttcaattac attgcaggtg cctctggtga tgaatggacc aaacgcgcaa acgatcgtgc atggaaacac aaactgctgt atccgcgcct ggcccaggat gtggaagcac cggatacttc cactgaaatc ctgggtcaca aaatcaaggc accgtttatt atggctccga tcgcagcgca cggcctggca cacaccacca aagaagctgg caccgctcgt gcggtttctg agttcggcac cattatgtct atctctgcgt atagcggtgc cactttcgag gaaatttccg agggcctgaa cggtggcccg 114 420 201127961 cgttggtttc agatttacat ggcgaaagat gaccagcaga accgcgatat cctggatgaa 480 gccaaatctg acggcgcgac tgctatcatc ctgaccgcgg actctaccgt atccggtaac 540 cgtgaccgtg atgtgaagaa caagttcgtc tatcctttcg gtatgccgat tgttcagcgc 600 tatctgcgcg gtaccgctga gggtatgagc ctgaacaaca tctatggtgc gtccaaacag 660 aaaatcagcc cacgtgacat cgaa gaaatt gctggtcata gcggtctgcc ggtgtttgtg 720 aaaggtatcc agcatccaga agatgcggac atggcaatca aacgtggtgc gtctggcatc 780 tgggttagca accacggtgc gcgtcagctg tacgaagctc cgggtagctt cgataccctg 840 ccggccatcg cggaacgtgt gaataaacgc gtgccgatcg ttttcgattc cggtgtgcgt 900 cgtggtgaac atgtggcaaa agcactggcg tctggcgctg atgtcgtagc actgggccgt 960 ccagtgctgt tcggtctggc tctgggtggc tggcagggcg cttactccgt cctggattac 1020 tttcagaaag acctgacccg tgttatgcag ctgaccggtt cccagaacgt agaggacctg 1080 aaaggcctgg acctgttcga caacccttac Ggttacgaat actaa 1125 &lt;2X0&gt; 90 &lt;211&gt; 314

&lt;212&gt; PRT &lt;213&gt; EC 1.1.1.27 - L-lactate dehydrogenase &gt; 〇8171^0_Corynebacterium glutamicum &lt;400&gt; 90

Met Lys Glu Thr Val Gly Asn Lys lie Val Leu lie Gly Ala Gly Asp 15 10 15

Val Gly Val Ala Tyr Ala Tyr Ala Leu lie Asn Gin Gly Met Ala Asp 20 25 30

His Leu Ala lie lie Asp lie Asp Glu Lys Lys Leu Glu Gly Asn Val 35 40 45

Met Asp Leu Asn His Gly Val Val Trp Ala Asp Ser Arg Thr Arg Val 50 55 60

Thr Lys Gly Thr Tyr Ala Asp Cys Glu Asp Ala Ala Met Val Val lie 65 70 75 80

Cys Ala Gly Ala Ala Gin Lys Pro Gly Glu Thr Arg Leu Gin Leu Val 85 90 95

Asp Lys Asn Val Lys lie Met Lys Ser lie Val Gly Asp Val Met Asp 100 105 110 115 201127961

Ser Gly Phe Asp Gly lie Phe Leu Val Ala Ser Asn Pro Val Asp lie 115 120 125

Leu Thr Tyr Ala Val Trp Lys Phe Ser Gly Leu Glu Trp Asn Arg Val 130 135 140 lie Gly Ser Gly Thr Val Leu Asp Ser Ala Arg Phe Arg Tyr Met Leu 145 150 155 160

Gly Glu Leu Tyr Glu Val Ala Pro Ser Ser Val His Ala Tyr lie lie 165 170 175

Gly Glu His Gly Asp Thr Glu Leu Pro Val Leu Ser Ser Ala Thr lie 180 185 190

Ala Gly Val Ser Leu Ser Arg Met Leu Asp Lys Asp Pro Glu Leu Glu 195 200 205

Gly Arg Leu Glu Lys lie Phe Glu Asp Thr Arg Asp Ala Ala Tyr His 210 215 220 . lie lie Asp Ala Lys Gly Ser Thr Ser Tyr Gly lie Gly Met Gly Leu 225 230 235 240

Ala Arg lie Thr Arg Ala lie Leu Gin Asn Gin Asp Val Ala Val Pro 245 250 255

Val Ser Ala Leu Leu His Gly Glu Tyr Gly Glu Glu Asp He Tyr lie 260 265 270

Gly Thr Pro Ala Val Val Asn Arg Arg Gly lie Arg Arg Val Val Glu 275 280 285

Leu Glu lie Thr Asp His Glu Met Glu Arg Phe Lys His Ser Ala Asn 290 295 300

Thr Leu Arg Glu lie Gin Lys Gin Phe Phe 305 310 &lt;210&gt; 91 &lt;211&gt; 329

&lt;212&gt; PRT &lt;213&gt; EC 1.1.1.28 - D-lactate dehydrogenase s &gt; 卩52643_Escherichia coli 116 201127961 &lt;400&gt; 91

Met Lys Leu Ala Val Tyr Ser Thr Lys Gin Tyr Asp Lys Lys Tyr Leu 15 10 15

Gin Gin Val Asn Glu Ser Phe Gly Phe Glu Leu Glu Phe Phe Asp Phe 20 25 30

Leu Leu Thr Glu Lys Thr Ala Lys Thr Ala Asn Gly Cys Glu Ala Val 35 40 45

Cys lie Phe Val Asn Asp Asp Gly Ser Arg Pro Val Leu Glu Glu Leu 50 55 60

Lys Lys His Gly Val Lys Tyr lie Ala Leu Arg Cys Ala Gly Phe Asn 65 70 75 80

Asn Val Asp Leu Asp Ala Ala Lys Glu Leu Gly Leu Lys Val Val Arg 85 90 95

Val Pro Ala Tyr Asp Pro Glu Ala Val Ala Glu His Ala lie Gly Met 100 105 110

Met Met Thr Leu Asn Arg Arg lie His Arg Ala Tyr Gin Arg Thr Arg 115 120 125

Asp Ala Asn Phe Ser Leu Glu Gly Leu Thr Gly Phe Thr Met Tyr Gly 130 135 140

Lys Thr Ala Gly Val lie Gly Thr Gly Lys lie Gly Val Ala Met Leu 145 150 155 160

Arg lie Leu Lys Gly Phe Gly Met Arg Leu Leu Ala Phe Asp Pro Tyr 165 170 175

Pro Ser Ala Ala Ala Leu Glu Leu Gly Val Glu Tyr Val Asp Leu Pro 180 185 190

Thr Leu Phe Ser Glu Ser Asp Val lie Ser Leu His Cys Pro Leu Thr 195 200 205

Pro Glu Asn Tyr His Leu Leu Asn Glu Ala Ala Phe Glu Gin Met Lys 210 215 220

Asn Gly Val Met lie Val Asn Thr Ser Arg Gly Ala Leu lie Asp Ser 117 201127961 225 230 235 240 Gin Ala Ala He Glu 245 Ala Leu Lys Asn Gin 250 Lys lie Gly Ser Leu 255 Gly

Met Asp Val Tyr Glu Asn Glu Arg Asp Leu Phe Phe Glu Asp Lys Ser 260 265 270

Asn Asp Val lie Gin Asp Asp Val Phe Arg Arg Leu Ser Ala Cys His 275 280 285

Asn Val Leu Phe Thr Gly His Gin Ala Phe Leu Thr Ala Glu Ala Leu 290 295 300

Thr Ser lie Ser Gin Thr Thr Leu Gin Asn Leu Ser Asn Leu Glu Lys 305 310 315 320

Gly Glu Thr Cys Pro Asn Glu Leu Val 325 &lt;210&gt; 92 &lt;211&gt; 312

&lt;212&gt; PRT &lt;213&gt; EC 1.1.1.37 - Malate dehydrogenase &gt; P61889-Escherichia coli &lt;400&gt; 92

Met Lys Val Ala Val Leu Gly Ala Ala Gly Gly lie Gly Gin Ala Leu 15 10 15

Ala Leu Leu Leu Lys Thr Gin Leu Pro Ser Gly Ser Glu Leu Ser Leu 20 25 30

Tyr Asp lie Ala Pro Val Thr Pro Gly Val Ala Val Asp Leu Ser His 35 40 45 lie Pro Thr Ala Val Lys lie Lys Gly Phe Ser Gly Glu Asp Ala Thr 50 55 60

Pro Ala Leu Glu Gly Ala Asp Val Val Leu lie Ser Ala Gly Val Ala 65 70 75 80

Arg Lys Pro Gly Met Asp Arg Ser Asp Leu Phe Asn Val Asn Ala Gly 85 90 95

He Val Lys Asn Leu Val Gin Gin Val Ala Lys Thr Cys Pro Lys Ala 118 201127961 100 105 110 Cys lie Gly lie lie Thr Asn Pro Val Asn Thr Thr Val Ala lie Ala 1X5 120 125 Ala Glu Val Leu Lys Lys Ala Gly Val Tyr Asp Lys Asn Lys Leu Phe 130 135 140 Gly Val Thr Thr Leu Asp He lie Arg Ser Asn Thr Phe Val Ala Glu 145 150 155 160

Leu Lys Gly Lys Gin Pro Gly Glu Val Glu Val Pro Val lie Gly Gly 165 170 175

His Ser Gly Val Thr lie Leu Pro Leu Leu Ser Gin Val Pro Gly Val 180 185 190

Ser Phe Thr Glu Gin Glu Val Ala Asp Leu Thr Lys Arg lie Gin Asn 195 200 205

Ala Gly Thr Glu Val Val Glu Ala Lys Ala Gly Gly Gly Ser Ala Thr 210 215 220

Leu Ser Met Gly Gin Ala Ala Ala Arg Phe Gly Leu Ser Leu Val Arg 225 230 235 240

Ala Leu Gin Gly Glu Gin Gly Val Val Glu Cys Ala Tyr Val Glu Gly 245 250 255

Asp Gly Gin Tyr Ala Arg Phe Phe Ser Gin Pro Leu Leu Leu Gly Lys 260 265 270

Asn Gly Val Glu Glu Arg Lys Ser He Gly Thr Leu Ser Ala Phe Glu 275 280 285

Gin Asn Ala Leu Glu Gly Met Leu Asp Thr Leu Lys Lys Asp lie Ala 290 295 300

Leu Gly Glu Glu Phe Val Asn Lys 305 310 &lt;210&gt; 93 &lt;211&gt; 312

&lt;212&gt; PRT &lt;213&gt; P49814_Bacinus subtilis 119 201127961 &lt;400&gt; 93

Met Gly Asn Thr Arg Lys Lys Val Ser Val lie Gly Ala Gly Phe Thr 15 10 15

Gly Ala Thr Thr Ala Phe Leu lie Ala Gin Lys Glu Leu Ala Asp Val 20 25 30

Val Leu Val Asp lie Pro Gin Leu Glu Asn Pro Thr I^ys Gly Lys Ala 35 40 45

Leu Asp Met Leu Glu Ala Ser Pro Val Gin Gly Phe Asp Ala Lys lie 50 55 60

Thr Gly Thr Ser Asn Tyr Glu Asp Thr Ala Gly Ser Asp lie Val Val 65 70 75 80 lie Thr Ala Gly lie Ala Arg Lys Pro Gly Met Ser Arg Asp Asp Leu 85 90 95

Val Ser Thr Asn Glu Lys lie Met Arg Ser Val Thr Gin Glu lie Val 100 105 110

Lys Tyr Ser Pro Asp Ser lie lie Val Val Leu Thr Asn Pro Val Asp 115 120 125

Ala Met Thr Tyr Ala Val Tyr Lys Glu Ser Gly Phe Pro Lys Glu Arg 130 135 140

Val lie Gly Gin Ser Gly Val Leu Asp Thr Ala Arg Phe Arg Thr Phe 145 150 155 160

Val Ala Glu Glu Leu Asn Leu Ser Val Lys Asp Val Thr Gly Phe Val 165 170 175

Leu Gly Gly His Gly Asp Asp Met Val Pro Leu Val Arg Tyr Ser Tyr 180 185 190

Ala Gly Gly lie Pro Leu Glu Thr Leu lie Pro Lys Glu Arg lie Asp 19S 200 205

Ala lie Val Glu Arg Thr Arg Lys Gly Gly Gly Glu lie Val Asn Leu 210 215 220 120 201127961

Leu Gly Asn Gly Ser Ala Tyr Tyr Ala Pro Ala Ala Ser Leu Thr Glu 225 230 235 240

Met Val Glu Ala lie Leu Lys Asp Gin Arg Arg Val Leu Pro Thr lie 245 250 255

Ala Tyr Leu Glu Gly Glu Tyr Gly Tyr Glu Gly lie Tyr Leu Gly Val 260 265 270

Pro Thr lie Val Gly Gly Asn Gly Leu Glu Gin lie lie Glu Leu Glu 275 280 285

Leu Thr Asp Tyr Glu Arg Ala Gin Leu Asn Lys Ser Val Glu Ser Val 290 295 300

Lys Asn Val Met Lys Val Leu Ser 305 310 &lt;210&gt; 94 &lt;211&gt; 365

&lt;212&gt; PRT &lt;213&gt; EC 1.1.1.81 - hydroxypyruvate reductase &gt; 厶3 乙1^9_Pichia stipitis &lt;400&gt; 94

Met Thr Leu Lys Gin Gin Val Leu Phe Val Gly Lys Pro Asn Thr Asn 15 10 15

Thr Glu Ala Tyr Lys Lys Phe Ser Ala Asn Phe Glu Val lie Asn Tyr 20 25 30

Lys lie Thr Ser Lys Ser Gin Leu lie Glu Asp Phe Glu Gly Arg Leu 35 40 45

Arg Tyr lie Glu Ala lie Tyr Ala Gly Trp Gly Gly Phe Asp Gly Val 50 55 60

Gly Gly Phe Gin Gly Glu Val Leu Arg His Cys Pro Pro Asn Val Lys 65 70 75 80

Val Val Ala lie Cys Ser lie Gly His Asp Gly Tyr Asp Thr Glu Gly 85 90 95

Met Ser Lys Arg Gly lie Thr Leu Thr Asn Val Pro Ser Val lie Ala 100 105 110

S 121 201127961

Ser Glu Ala Val Ala Asp Leu Val Leu Tyr Asn Thr Leu Ser Ser Phe 115 120 125

Arg Asn Phe Lys Met Phe Glu Lys Asn Leu Gly Gly Lys Leu Thr Asn 130 135 140

Thr Gly Ala Leu Arg Thr Ala Leu Val Arg Gly Glu Phe Asp Gin Phe 145 150 155 160

Asn Gly Val Pro Val lie Lys Pro Thr Val Gly Gly Ala Phe Ala Ser 165 170 175

Ser Cys Cys Gly Arg Asp lie Leu Ser Pro Arg Gly His Asn Val Val 180 185 190 lie Val Gly Phe Gly Ser lie Gly Lys Leu lie Gly Glu Arg Leu Ala 195 200 205

Cys lie Gly Met Asn lie His Tyr Val Lys Arg Ser Lys Leu Ser Glu 210 215 220

Gin Glu Glu Ala Ser Leu Gly Tyr Lys Val Thr Tyr His Ala Thr Leu 225 230 235 240

Lys Asp Thr Lys Asn lie Ala Asp Leu Val Val lie Ala Cys Pro Gly 245 250 255

Thr Ala His Thr Arg His Met Val Asn Glu Glu Met lie Asn Asp Phe 260 265 270

Ala Lys Pro Phe Arg Leu lie Asn lie Gly Arg Gly Tyr Val Val Asp 275 280 285

Glu Lys Ala Leu Val Asn Gly Leu Gin Ser Gly Lys lie Leu Phe Ala 290 295 300

Gly Leu Asp Val Phe Glu Asn Glu Pro Ser lie Asn Pro Asp Leu Leu 305 310 315 320

Asn Arg Gin Asp Val Val Leu Thr Pro His lie Gly Ser Ser Thr Thr 325 330 335

Glu Asn Phe Asn Tyr Thr Ala Ala Ala Ala Met Phe Asn lie Glu Thr 340 345 350 122 201127961

Val Leu Tyr Asp Arg Glu Asp Thr lie Thr Arg Val Asn 355 360 365 &lt;210&gt; 95 &lt;211&gt; 424

&lt;212&gt; PRT &lt;213&gt;&gt;Q88F00 Pseudomonas putida &lt;400&gt; 95

Met Ser Val Asp Pro Gin Lys Leu Leu Arg Glu Leu Phe Asp Thr Ala 15 10 15 lie Ala Ala Ala His Pro Arg Gin Val Leu Glu Pro Tyr Leu Pro Ala 20 25 30

Asp Arg Ser Gly Arg Val lie Val 35 40 lie Gly Ala Gly Lys Ala Ala Ala 45

Ala Met Ala Glu Val Val Glu Lys 50 55

Ser Trp Gin Gly Glu Val 60

Ser Gly

Leu Val Val Thr Arg Tyr Gly His Gly Ala Asn Cys Gin Lys lie Glu 65 70 75 80

Val Val Glu Ala Ala His Pro Val Pro Asp Ala Ala Gly Leu Ala Val 85 90 95

Ala Lys Arg Val Leu Glu Leu Val Ser Asn Leu Asn Glu Glu Asp Arg 100 105 110

Val lie Phe Leu Leu Ser Gly Gly Gly Ser Ala Leu Leu Ala Leu Pro 115 120 125

Ala Glu Gly Leu Thr Leu Ala Asp Lys Gin Gin lie Asn Lys Ala Leu 130 135 140

Leu Lys Ser Gly Ala Thr lie Gly Glu Met Asn Cys Val Arg Lys His 145 150 155 160

Leu Ser Ala lie Lys Gly Gly Arg Leu Ala Lys Ala Cys Trp Pro Ala 165 170 175

Thr Val Tyr Thr Tyr Ala lie Ser Asp Val Pro Gly Asp Leu Ala Thr 180 185 190 123 S. 201127961

Val lie Ala Ser Gly Pro Thr Val Ala Asp Pro Ser Thr Ser Ala Asp 195 200 205

Ala Leu Ala lie Leu Lys Arg Tyr Asn lie Glu Ala Pro Lys Ala Val 210 215 220 lie Asp Trp Leu Asn Asn Pro Ala Ser Glu Thr Val Lys Ala Asp Asp 225 230 235 240

Pro Ala Leu Ala Arg Ser His Phe Gin Leu lie Ala Lys Pro Gin Gin 245 250 255

Ser Leu Glu Ala Ala Ala Val Lys Ala Arg Gin Ala Gly Phe Ser Pro 260 265 270

Leu lie Leu Gly Asp Leu Glu Gly Glu Ser Arg Glu Val Ala Lys Val 275 280 285

His Ala Gly lie Ala Arg Gin lie Val Gin His Gly Gin Pro Leu Lys 290 295 300

Ala Pro Cys Val lie Leu Ser Gly Gly Glu Thr Thr Val Thr Val Arg 305 310 315 320

Gly Asn Gly Arg Gly Gly Arg Asn Ala Glu Phe Leu Leu Ser Leu Thr 325 330 335

Glu Ser Leu Lys Gly Leu Pro Gly Val Tyr Ala Leu Ala Gly Asp Thr 340 345 350

Asp Gly lie Asp Gly Ser Glu Glu Asn Ala Gly Ala Phe Met Thr Pro 355 360 365

Ala Ser Tyr Ala Ser Ala Glu Ala Leu Gly Leu Ser Ala Ser Asp Glu 370 375 380

Lau Asp Asn Asn Asn Gly Tyr Gly Tyr Phe Ala Ala Leu Asp Ala Leu 385 390 395 400 lie Val Thr Glu Pro Thr Arg Thr Asn Val Asn Asp Phe Arg Ala lie 405 410 415

Leu lie Leu Glu Thr Ala Gin Ser 420 124 201127961 &lt;210&gt; 96 &lt;211&gt; 347

&lt;212&gt; PRT &lt;213&gt; EC 1.1, 1.82 - Pinacid dehydrogenase [NADP+] &gt;〇8耶1&lt;:9_Corynebacterium glutamicum &lt;400&gt; 96

Met Pro Glu Val Thr Val Asn Ala Gin Gin Leu Thr Val Leu Cys Thr 1 5 10 15

Asp lie Leu Thr Lys Thr Gly Val Pro Ala Ala Asp Ala His Leu Val 20 25 30

Gly Asp Ser Leu Val Gin Ala Asp Leu Trp Gly His Pro Ser His Gly 35 40 45

Val Leu Arg Leu Pro Trp Tyr Val Arg Arg Leu His Ser Gly Ala Met 50 55 60

Thr Thr His Ala His Val Glu Val Leu Asn Asp Leu Gly Ala Val Leu 65 70 75 80

Ala Leu Asp Gly His Asn Gly lie Gly Gin Val Leu Ala Asp His Ala 85 90 95

Arg Lys Glu Ala Val Thr Arg Ala Met Met Phe Gly lie Gly Ala Val 100 105 110

Ser Val Arg Asn Ser Asn His Phe Gly Thr Ala Met Tyr Tyr Thr Arg 115 120 125

Lys Ala Ala Ala Gin Gly Cys Val Ser lie Leu Thr Thr Asn Ala Ser 130 135 140

Pro Ala Met Ala Pro Trp Gly Gly Arg Glu Lys Arg lie Gly Thr Asn 145 150 155 160

Pro Trp Ser lie Ala Ala Pro Phe Gly Glu Thr Ala Thr Val Val Asp 165 170 175

He Ala Asn Thr Ala Val Ala Arg Gly Lys lie Tyr His Ala Arg Gin 180 185 190

Thr Asn Met Pro lie Pro Glu Thr Trp Ala lie Thr Ser Glu Gly Ala e 125 201127961 195 200 205 Pro Thr Thr Asp Pro Ala Glu Ala He Asn Gly Val Val Leu Pro Met 210 215 220

Ala Gly His Lys Gly Tyr Ala lie Ser Phe Met Met Asp Val Leu Ser 225 230 235 240

Gly Val Leu Thr Gly Ser Gin His Ser Thr Lys Val His Gly Pro Tyr 245 250 255

Asp Pro Thr Pro Pro Gly Gly Ala Gly His Leu Phe lie Ala Leu Asp 260 265 270

Val Ala Ala Phe Arg Asp Pro Gin Asp Phe Asp Asp Ala Leu Ser Asp 275 280 285

Leu Val Gly Glu Val Lys Ser Thr Pro Lys Ala Gin Asn Thr Glu Glu 290 295 300 lie Phe Tyr Pro Gly Glu Ser Glu Asp Arg Ala His Arg Lys Asn Ser 305 310 315 320

Ala His Gly lie Ser Leu Pro Glu Lys Thr Trp Met Glu Leu Gin Glu 325 330 335

Leu Ala lie Glu Asn His Val Val Thr His Arg 340 345 &lt;210&gt; 97 &lt;211&gt; 315

&lt;212&gt; PRT &lt;213&gt;&gt;Q5E5E9j Vibrio fischeri &lt;400&gt; 97

Met Lys Val Ser Tyr Tyr Glu Val Lys Glu Arg Leu lie Arg Lys Phe 15 10 15 lie Ala Ser Gly Leu Ala Trp Asp Asp Ala Asn Trp Val Thr Asp Val 20 25 30

Leu lie Ser Ser Glu Gin Arg Gly Asp Lys Ser His Gly lie Lys His 35 40 45

Ala Lys Asn He Phe Asp Val lie Asn Ser Glu Cys Tyr lie Ala Gin 126 201127961 50 55 60 Ala Pro lie lie His Asp Glu Arg Ser lie Thr lie Leu Asp Gly Gin 65 70 75 80

Asn Ser lie Gly Pro lie Val Ala Lys Gin Ala lie Asp lie Ala lie 85 90 95

Lys Lys Ala Lys Lys Tyr Gly Thr Ala Ala lie Ser Leu Arg Ser Ser 100 105 no

Asn His Leu Phe Ser Leu Ser His Tyr Val Arg Tyr lie Ala Asn Asn 115 120 125

Asn Met lie Gly Phe lie Cys Ser Ser Ser Ser Ala Met Ala Ala 130 135 140

Pro Asn Ser Leu Asn Ala Thr lie Gly Thr Asn Pro Phe Ala Phe Gly 145 150 155 160

Ala Pro Ser Ser Lys Asp Pro lie Val lie Asp Met Ser Ser As As 165 170 175

Val Ala Arg Gly Lys lie Lys Glu Tyr Lys Asp Ala Glu Leu Asp lie 180 185 190

Pro Val Ser Trp Ala Leu Asp Glu Tyr Gly Asn Pro Thr Thr Cys Ala 195 200 205 lie Glu Ala Leu Lys Gly Thr Leu Ser Pro Leu Gly Gly Tyr Lys Gly 210 215 220

Phe Ala Leu Gly Cys Met lie Asp lie Phe Ser Ser Val Leu Ser Gly 225 230 235 240

Ser Ala Phe Ser Thr Gin lie Thr Gly Thr Ser Leu His Met Glu Glu 245 250 255

Ala Asp Val Asn Lys Lys Gly Asp Phe Leu Phe Val Leu Asp lie Ser 260 265 270

Lys Phe lie Gin Leu Ser Glu Phe Lys lie Arg Met Asp Glu Phe lie 275 280 285 127 £ 201127961

His lie lie Glu Ser Asn Gly Gly Tyr lie Pro Gly Thr Asn Tyr lie 29. 295 300

Asn Asn Gin Phe Ala Asp lie Glu lie Leu Asn 305 310 315 &lt;210&gt; 98 &lt;211&gt; 354

&lt;212&gt; PRT &lt;213&gt; EC 1.1.1.85 - 3-isopropylmalate dehydrogenase&gt; A9VLG8-Bacillus weihenstephanensis &lt;400&gt; 98

Met Glu Lys Arg lie Val Cys Leu Ala Gly Asp Gly Val Gly Pro Glu 15 10 15 lie Met Glu Ser Ala Lys Glu Val Leu His Met Val Glu Arg Leu Tyr 20 25 30

Gly His His Phe His Leu Gin Asp Glu Tyr Phe Gly Gly Ala Ala lie 35 40 45

Asp Leu Asn Gly Gin Pro Leu Pro Gin Arg Thr Leu Ala Ala Cys Leu 50 55 60

Ala Ser Asp Ala Val Leu Leu Gly Ala Val Gly Gly Pro Arg Trp Asp 65 70 75 80

Asp Ala Lys Glu Arg Pro Glu Lys Gly Leu Leu Ala Leu Arg Lys Gly 85 90 95

Leu Gly Val Phe Ala Asn Val Arg Pro Val Thr Val Glu Ser Ala Thr 100 105 110

Ala His Leu Ser Pro Leu Lys Asn Ala Asp Glu lie Asp Phe Val Val 115 120 125

Val Arg Glu Leu Thr Gly Gly lie Tyr Phe Ser Tyr Pro Lys Glu Arg 130 135 140

Thr Glu Glu Ser Ala Thr Asp Thr Leu Thr Tyr His Arg His Glu lie 145 150 155 160

Glu Arg lie Val Ser Tyr Ala Phe Gin Leu Ala Ser Lys Arg Glu Lys 165 170 175 128 201127961

Lys Val Thr Ser lie Asp Lys Ala Asn Val Leu Glu Ser Ser Lys Leu 180 185 190

Trp Arg Ala Val Thr Glu Glu Val Ala Leu Arg Tyr Pro Asn Val Glu 195 200 205

Leu Glu His lie Leu Val Asp Ala Ala Ala Met Glu Leu lie Arg Asn 210 215 220

Pro Arg Arg Phe Asp Val lie Val Thr Glu Asn Leu Phe Gly Asp lie 225 230 235 240

Leu Ser Asp Glu Ala Ser Val Leu Ala Gly Ser Leu Gly Met Leu Pro 245 250 255

Pro lie His 270

Ser Ala Ser His Ala Glu Asn Gly Pro Ser Leu Tyr Glu 260 265

Gly Ser Ala Pro Asp lie Ala Gly Lys Asn Lys Ala Asn Pro lie Ala 275 280 285

Met Met Arg Ser Val Ala Met Met Leu Gly Gin Ser Phe Gly Leu Thr 290 295 300

Arg Glu Gly Tyr Ala He Glu Glu Ala He Ser Ala Val Leu Gin Ser 305 310 315 320

Gly Lys Cys Thr Ala Asp He Gly Gly Asn Glu Thr Thr Thr Ser Phe 325 330 335

Thr Arg Ala Val lie Gin Glu Met Glu Glu Gin Ala Leu Val Gly Arg 340 345 350

Gly Arg &lt;210&gt; 99 &lt;211&gt; 349

&lt;212&gt; PRT &lt;213&gt;&gt; Q5NPQ9 zymomonas mobilis &lt;400&gt; 99

Met Arg lie Ala Leu Leu Ala Gly Asp Gly lie Gly Pro Glu lie Thr 1 5 i〇 15 129 201127961

Ala Glu Ala Val Lys lie Leu Lys Ala Val Val Gly Gin Glu lie Glu 20 25 30

Phe Asp Glu Ala Leu lie Gly Gly Ala Ala Trp Lys Val Thr Gly Ser 35 40 45

Pro Leu Pro Glu Glu Thr Leu Lys Leu Cys Lys Asn Ser Asp Ala lie 50 55 60

Leu Phe Gly Ser Val Gly Asp Pro Glu Cys Asp His Leu Glu Arg Ala 65 70 75 80

Leu Arg Pro Glu Gin Ala lie Leu Gly Leu Arg Lys Glu Leu Asp Leu 85 90 95

Phe Ala Asn Leu Arg Pro Ala Arg Leu Phe Pro Glu Leu Gin Ala Glu 100 105 * 110

Ser Pro Leu Lys Glu Asn lie Val Thr Gly Thr Asp Leu Met lie Val 115 120 125

Arg Glu Leu Thr Gly Asp Val Tyr Phe Gly Thr Pro Arg Gly Gin Arg 130 135 140

Lys Asp Asp Gin Asn Arg Arg Glu Gly Phe Asp Thr Met Arg Tyr Asn 145 150 155 160

Glu Asp Glu Val Lys Arg lie Ala Arg lie Gly Phe Glu Thr Ala Arg 165 170 175

Ser Arg Ser Gly Asn Leu Cys Ser lie Asp Lys Ser Asn Val Leu Glu 180 185 190

Thr Ser Gin Leu Trp Arg Thr Val Val Leu Glu lie Ala Gin Glu Tyr 195 200 205

Pro Asp Val Glu Leu Ser His Met Tyr Val Asp Asn Ala Ala Met Gin 210 215 220

Leu Val Arg Ala Pro Asp Gin Phe Asp Val lie Val Thr Gly Asn Leu 225 230 235 240

Phe Gly Asp lie Leu Ser Asp Leu Ala Ser Ala Cys Val Gly Ser lie 245 250 255 130 201127961

Gly Leu Leu Pro Ser Ala Ser Leu Asn Ser Glu Gly Lys Gly Leu Tyr 260 265 270

Glu Pro lie His Gly Ser Ala Pro Asp He Ala Gly Leu Gly Lys Ala 275 280 285

Asn Pro Leu Ala Thr lie Leu Ser Gly Ala Met Met Leu Arg Tyr Ser 290 295 300

Leu Lys Arg Glu Ala Asp Ala Asp Arg lie Glu Lys Ala Val Ser Thr 305 310 315 320

Ala Leu Glu Lys Gly Ala Arg Thr Ala Asp Leu Gly Gly Lys Met Thr 325 330 335

Thr Ser Glu Met Gly Asn Ala Val Leu Ala Ala Leu Asn 340 345 &lt;210&gt; 100 &lt;211&gt; 361

&lt;212&gt; PRT &lt;213&gt; EC 1.1.1.93 - tartaric acid dehydrogenase &gt; P76251__Escherichia coli &lt;400&gt;

Met Met Lys Thr Met Arg lie Ala Ala lie Pro Gly Asp Gly lie Gly 1 5 10 15

Lys Glu Val Leu Pro Glu Gly lie Arg Val Leu Gin Ala Ala Ala Glu 20 25 30

Arg Trp Gly Phe Ala Leu Ser Phe Glu Gin Met Glu Trp Ala Ser Cys 35 40 45

Glu Tyr Tyr Ser His His Gly Lys Met Met Pro Asp Asp Trp His Glu 50 55 60

Gin Leu Ser Arg Phe Asp Ala lie Tyr Phe Gly Ala Val Gly Trp Pro 65 70 75 80

Asp Thr Val Pro Asp His lie Ser Leu Trp Gly Ser Leu Leu Lys Phe 85 90 95

Arg Arg Glu Phe Asp Gin Tyr Val Asn Leu Arg Pro Val Arg Leu Phe 100 105 no 131 201127961

Pro Gly Val Pro Cys Pro Leu Ala Gly Lys Gin Pro Gly Asp lie Asp 115 120 125

Phe Tyr Val Val Arg Glu Asn Thr Glu Gly Glu Tyr Ser Ser Leu Gly 130 135 140

Gly Arg Val Asn Glu Gly Thr Glu His Glu Val Val He Gin Glu Ser 145 150 155 160

Val Phe Thr Arg Arg Gly Val Asp Arg lie Leu Arg Tyr Ala Phe Glu 165 170 175

Leu Ala Gin Ser Arg Pro Arg Lys Thr Leu Thr Ser Ala Thr Lys Ser 180 185 190

Asn Gly Leu Ala lie Ser Met Pro Tyr Trp Asp Glu Arg Val Glu Ala 195 200 205

Met Ala Glu Asn Tyr Pro Glu lie Arg Trp Asp Lys Gin His lie Asp 210 215 220 lie Leu Cys Ala Arg Phe Val Met Gin Pro Glu Arg Phe Asp Val Val 225 230 235 240

Val Ala Ser Asn Leu Phe Gly Asp lie Leu Ser Asp Leu Gly Pro Ala 245 250 255

Cys Thr Gly Thr lie Gly lie Ala Pro Ser Ala Asn Leu Asn Pro Glu 260 265 270

Arg Thr Phe Pro Ser Leu Phe Glu Pro Val His Gly Ser Ala Pro Asp 275 280 285 lie Tyr Gly Lys Asn lie Ala Asn Pro lie Ala Thr lie Trp Ala Gly 290 295 300

Ala Met Met Leu Asp Phe Leu Gly Asn Gly Asp Glu Arg Phe Gin Gin 305 310 315 320

Ala His Asn Gly lie Leu Ala Ala lie Glu Glu Val lie Ala His Gly 325 330 335

Pro Lys Thr Pro Asp Met Lys Gly Asn Ala Thr Thr Pro Gin Val Ala 132 201127961 340 345 350

Asp Ala lie Cys Lys lie lie Leu Arg 355 360 &lt;210&gt; 101 &lt;211&gt; 362

&lt;212&gt; PRT &lt;213&gt;&gt;A2Q846"#, Aspergillus niger &lt;400&gt; 101

Met Thr Thr Glu Thr Thr Thr Tyr Arg lie Ala Ser lie Pro Gly Asp 15 10 15

Gly lie Gly Glu Glu Val Val Arg Ala Thr lie Glu Val lie Asn Lys 20 25 30

Leu Ala Gin Thr Leu Asn Thr Phe Asn lie Glu Phe Thr His Leu Pro 35 40 45

Trp Gly Thr Glu Tyr Tyr Lys Gin His Gly Arg Tyr Val Ser Glu Gly 50 55 60

Tyr Leu Asp Thr Leu Arg Gin Phe Asp Ala Gly Leu Phe Gly Ser Val 65 70 75 80

Gly His Pro Asp Val Pro Asp His Val Ser Leu Trp Gly Leu Leu Leu 85 90 95

Ala Leu Arg Ser Pro Leu Gin Leu Tyr Ala Asn Val Arg Pro Val Arg 100 105 110

Thr Phe Pro Gly Thr Lys Ser Pro Leu Thr Thr Ala Val Asn Gly lie 115 120 125

Asp Trp Val Leu Val Arg Glu Asn Ser Glu Gly Glu Tyr Cys Gly Gin 130 135 140

Gly Gly Arg Ser His Thr Gly Gin Pro Trp Glu Ala Ala Thr Glu Val 145 150 155 160

Ala lie Phe Thr Arg Val Gly Val Glu Arg lie Met Arg Phe Ala Phe 165 170 175

Glu Thr Ala Arg Ser Arg Pro Arg Arg His Leu Thr Val Val Thr Lys 133 201127961 180 185 Ser Asn Ala Met Arg His Gly Met Val Leu Trp Asp 195 200 190

Glu Val Ala Glu 205

Glu Val Ala Lys Asp Phe Pro Asp Val Thr Trp Asp 210 215 220

Lys Met Leu Val

Asp Ala Met Thr Leu Arg Met lie Ser Lys Pro Glu 225 230 235

Ser Leu Asp Thr 240 lie Val Gly Thr Asn Leu His Met Asp lie Leu Ser 245 250

Asp Leu Ala Ala 255

Gly Leu Ala Gly Ser lie Gly Val Ala Pro Ser Ser 260 265

Asn Leu Asp Pro 270

Thr Arg Lys Asn Pro Ser Leu Phe Glu Pro Val His 275 280

Gly Ser Ala Phe 285

Asp lie Met Gly Lys Gly Val Ala Asn Pro Val Ala 290 295 300

Thr Phe Trp Ser

Ala Ala Glu Met Leu Ala Trp Leu Gly Glu Lys Asp 305 310 315

Ala Ala Lys Lys 320

Leu Met Asp Cys Val Glu Lys Val Cys Ala Ala Gly 325 330 lie Leu Thr Pro 335

Asp Leu Gly Gly Ser Ala Asn Thr Gin Gly Val Val 340 345

Asp Ala Val Cys 350

Lys Glu lie Glu Gin Gin Leu Ala Ser Ser 355 360 &lt;210&gt; 102 &lt;211&gt; 591 &lt;212&gt; PRT &lt;213&gt; EC 1.] -.2.3 - L-lactate dehydrogenase (cytochrome) &gt ;P00175 cerevisiae) &lt;400&gt; 102 Let Saccharomyces

Met Leu Lys Tyr Lys Pro Leu Leu Lys lie Ser Lys 15 l〇

Asn Cys Glu Ala 15 134 201127961

Ala lie Leu Arg Ala Ser Lys Thr Arg Leu Asn Thr lie Arg Ala Tyr 20 25 30

Gly Ser Thr Val Pro Lys Ser Lys Ser Phe Glu Gin Asp Ser Arg Lys 35 40 45

Arg Thr Gin Ser Trp Thr Ala Leu Arg Val Gly Ala lie Leu Ala Ala 50 55 60

Thr Ser Ser Val Ala Tyr Leu Asn Trp His Asn Gly Gin lie Asp Asn 65 70 75 80

Glu Pro Lys Leu Asp Met Asn Lys Gin Lys lie Ser Pro Ala Glu Val 85 90 95

Ala Lys His Asn Lys Pro Asp Asp Cys Trp Val Val lie Asn Gly Tyr 100 105 110

Val Tyr Asp Leu Thr Arg Phe Leu Pro Asn His Pro Gly Gly Gin Asp 115 120 125

Val lie Lys Phe Asn Ala Gly Lys Asp Val Thr Ala lie Phe Glu Pro 130 135 140

Leu His Ala Pro Asn Val lie Asp Lys Tyr lie Ala Pro Glu Lys Lys 145 150 155 160

Leu Gly Pro Leu Gin Gly Ser Met Pro Pro Glu Leu Val Cys Pro Pro 165 170 175

Tyr Ala Pro Gly Glu Thr Lys Glu Asp lie Ala Arg Lys Glu Gin Leu 180 185 190

Lys Ser Leu Leu Pro Pro Leu Asp Asn lie lie Asn Leu Tyr Asp Phe 195 200 205

Glu Tyr Leu Ala Ser Gin Thr Leu Thr Lys Gin Ala Trp Ala Tyr Tyr 210 215 220

Ser Ser Gly Ala Asn Asp Glu Val Thr His Arg Glu Asn His Asn Ala 225 230 235 240

Tyr His Arg lie Phe Phe Lys Pro Lys lie Leu Val Asp Val Arg Lys 245 250 255 135 201127961

Val Asp lie Ser Thr Asp Met Leu Gly Ser His Val Asp Val Pro Phe 260 265 270

Tyr Val Ser Ala Thr Ala Leu Cys Lys Leu Gly Asn Pro Leu Glu Gly 275 280 285

Glu Lys Asp Val Ala Arg Gly Cys Gly Gin Gly Val Thr Lys Val Pro 290 295 300

Gin Met lie Ser Thr Leu Ala Ser Cys Ser Pro Glu Glu lie lie Glu 305 310 315 320

Ala Ala Pro Ser Asp Lys Gin lie Gin Trp Tyr Gin Leu Tyr Val Asn 325 330 335

Ser Asp Arg Lys lie Thr Asp Asp Leu Val Lys Asn Val Glu Lys Leu 340 345 350

Gly Val Lys Ala Leu Phe Val Thr Val Asp Ala Pro Ser Leu Gly Gin 355 360 365

Arg Glu Lys Asp Met Lys Leu Lys Phe Ser Asn Thr Lys Ala Gly Pro 370 375 380

Lys Ala Met Lys Lys Thr Asn Val Glu Glu Ser Gin Gly Ala Ser Arg 385 390 395 400

Ala Leu Ser Lys Phe lie Asp Pro Ser Leu Thr Trp Lys Asp lie Glu 405 410 415

Glu Leu Lys Lys Lys Thr Lys Leu Pro lie Val lie Lys Gly Val Gin 420 425 430

Arg Thr Glu Asp Val He Lys Ala Ala Glu lie Gly Val Ser Gly Val 435 440 445

Val Leu Ser Asn His Gly Gly Arg Gin Leu Asp Phe Ser Arg Ala Pro 450 455 460 lie Glu Val Leu Ala Glu Thr Met Pro lie Leu Glu Gin Arg Asn Leu 465 470 475 480

Lys Asp Lys Leu Glu Val Phe Val Asp Gly Gly Val Arg Arg Gly Thr 485 490 495 136 201127961

Asp Val Leu Lys Ala Leu Cys Leu Gly Ala Lys Gly Val Gly Leu Gly 500 505 510

Arg Pro Phe Leu Tyr Ala Asn Ser Cys Tyr Gly Arg Asn Gly Val Glu 515 520 525

Lys Ala lie Glu lie Leu Arg Asp Glu lie Glu Met Ser Met Arg Leu 530 535 540

Leu Gly Val Thr Ser lie Ala Glu Leu Lys Pro Asp Leu Leu Asp Leu 545 550 555 560

Ser Thr Leu Lys Ala Arg Thr Val Gly Val Pro Asn Asp Val Leu Tyr 565 570 575

Asn Glu Val Tyr Glu Gly Pro Thr Leu Thr Glu Phe Glu Asp Ala 580 585 590 &lt;210&gt; 103 &lt;211&gt; 396

&lt;212&gt; PRT &lt;213&gt;&gt;?33232_E. coli (Escherichia coli} &lt;400&gt; 103

Met lie lie Ser Ala Ala Ser Asp Tyr Arg Ala Ala Ala Gin Arg lie 15 10 15

Leu Pro Pro Phe Leu Phe His Tyr Met Asp Gly Gly Ala Tyr Ser Glu 20 25 30

Tyr Thr Leu Arg Arg Asn Val Glu Asp Leu Ser Glu Val Ala Leu Arg 35 40 45

Gin Arg lie Leu Lys Asn Met Ser Asp Leu Ser Leu Glu Thr Thr Leu 50 55 60

Phe Asn Glu Lys Leu Ser Met Pro Val Ala Leu Ala Pro Val Gly Leu 65 70 75 80

Cys Gly Met Tyr Ala Arg Arg Gly Glu Val Gin Ala Ala Lys Ala Ala 85 90 95

Asp Ala His Gly lie 100

Pro Phe Thr Leu Ser Thr Val Ser Val Cys Pro 105 110 137 201127961

He Glu Glu Val Ala Pro Ala lie Lys Arg Pro Met Trp Phe Gin Leu 115 120 125

Tyr Val Leu Arg Asp Arg Gly Phe Met Arg Asn Ala Leu Glu Arg Ala 130 135 140

Lys Ala Ala Gly Cys Ser Thr Leu Val Phe Thr Val Asp Met Pro Thr 145 150 155 160

Pro Gly Ala Arg Tyr Arg Asp Ala His Ser Gly Met Ser Gly Pro Asn 165 170 175

Ala Ala Met Arg Arg Tyr Leu Gin Ala Val Thr His Pro Gin Trp Ala 180 185 190

Trp Asp Val Gly Leu Asn Gly Arg Pro His Asp Leu Gly Asn lie Ser 195 200 205

Ala Tyr Leu Gly Lys 210

Pro Thr Gly Leu Glu Asp Tyr lie Gly Trp Leu 215 220

Gly Asn Asn Phe Asp Pro Ser lie Ser Trp Lys Asp Leu Glu Trp lie 225 230 235 240

Arg Asp Phe Trp Asp Gly Pro Met Val lie Lys Gly lie Leu Asp Pro 245 250 255

Glu Asp Ala Arg Asp Ala Val Arg Phe Gly Ala Asp Gly lie Val Val 260 265 270

Ser Asn His Gly Gly Arg Gin Leu Asp Gly Val Leu Ser Ser Ala Arg 275 280 285

Ala Leu Pro Ala lie Ala Asp Ala Val Lys Gly Asp lie Ala lie Leu 290 295 300

Ala Asp Ser Gly lie Arg Asn Gly Leu Asp Val Val Arg Met lie Ala 305 310 315 320

Leu Gly Ala Asp Thr Val Leu Leu Gly Arg Ala Phe Leu Tyr Ala Leu 325 330 335

Ala Thr Ala Gly Gin Ala Gly Val Ala Asn Leu Leu Asn Leu lie Glu 138 201127961 340 345 350

Lys Glu Met Lys Val Ala Met Thr Leu Thr Gly Ala 355 360

Lys Ser lie Ser 365

Glu lie Thr Gin Asp Ser Leu Val Gin Gly Leu Gly 370 375 380

Lys Glu Leu Pro

Ala Ala Leu Ala Pro Met Ala Lys Gly Asn Ala Ala 385 390 395 &lt;210&gt; 104 &lt;211&gt; 587 &lt;212&gt; PRT &lt;213&gt; EC 1.1.2.4 - D-Lactate Dehydrogenase (Cytochrome)&gt;;P32891 cerevisiae) &lt;400&gt; 104 Sour wine yeast (Saccharomyces

Met Leu Trp Lys Arg Thr Cys Thr Arg Leu lie Lys 1 5 10

Pro lie Ala Gin 15

Pro Arg Gly Arg Leu Val Arg Arg Ser Cys Tyr Arg 20 25

Tyr Ala Ser Thr 30

Gly Thr Gly Ser Thr Asp Ser Ser Ser Gin Trp Leu 35 40

Lys Tyr Ser Val 45 lie Ala Ser Ser Ala Thr Leu Phe Gly Tyr Leu Phe 50 55 60

Ala Lys Asn Leu

Tyr Ser Arg Glu Thr Lys Glu Asp Leu He Glu Lys 65 70 75

Leu Glu Met Val 80

Lys Lys lie Asp Pro Val Asn Ser Thr Leu Lys Leu 85 90

Ser Ser Leu Asp 95

Ser Pro Asp Tyr Leu His Asp Pro Val Lys lie Asp 100 105

Lys Val Val Glu 110

Asp Leu Lys Gin Val Leu Gly Asn Lys Pro Glu Asn 115 120

Tyr Ser Asp Ala 125

Lys Ser Asp Leu Asp Ala His Ser Asp Thr Tyr Phe 130 135 140

Asn Thr His His 139 201127961

Pro Ser Pro Glu Gin Arg Pro Arg lie lie Leu Phe Pro His Thr Thr 145 150 155 160

Glu Glu Val Ser Lys lie Leu Lys lie Cys His Asp Asn Asn Met Pro 165 170 175

Val Val Pro Phe Ser Gly Gly Thr Ser Leu Glu Gly His Phe Leu Pro 180 185 190

Thr Arg lie Gly Asp Thr lie Thr Val Asp Leu Ser Lys Phe Met Asn 195 200 205

Asn Val Val Lys Phe Asp Lys Leu Asp Leu Asp lie Thr Val Gin Ala 210 215 220

Gly Leu Pro Trp Glu Asp Leu Asn Asp Tyr Leu Ser Asp His Gly Leu 225 230 235 240

Met Phe Gly Cys Asp Pro Gly Pro Gly Ala Gin lie Gly Gly Cys lie 245 250 255

Ala Asn Ser Cys Ser Gly Thr Asn Ala Tyr Arg Tyr Gly Thr Met Lys 260 265 270

Glu Asn lie lie Asn Met Thr lie Val Leu Pro Asp Gly Thr lie Val 275 280 285

Lys Thr Lys Lys Arg Pro Arg Lys Ser Ser Ala Gly Tyr Asn Leu Asn 290 295 300

Gly Leu Phe Val Gly Ser Glu Gly Thr Leu Gly lie Val Thr Glu Ala 305 310 315 320

Thr Val Lys Cys His Val Lys Pro Lys Ala Glu Thr Val Ala Val Val 325 330 335

Ser Phe Asp Thr lie Lys Asp Ala Ala Ala Cys Ala Ser Asn Leu Thr 340 345 350

Gin Ser Gly lie His Leu Asn Ala Met Glu Leu Leu Asp Glu Asn Met 355 360 365

Met Lys Leu lie Asn Ala Ser Glu Ser Thr Asp Arg Cys Asp Trp Val 370 375 380 140 201127961

Glu Lys Pro Thr Met Phe Phe Lys lie Gly Gly Arg Ser Pro Asn lie 385 390 395 400

Val Asn Ala Leu Val Asp Glu Val Lys Ala Val Ala Gin Leu Asn His 405 410 415

Cys Asn Ser Phe Gin Phe Ala Lys Asp Asp Asp Glu Lys Leu Glu Leu 420 425 430

Trp Glu Ala Arg Lys Val Ala Leu Trp Ser Val Leu Asp Ala Asp Lys 435 440 445

Ser Lys Asp Lys Ser Ala Lys lie Trp Thr Thr Asp Val Ala Val Pro 450 455 460

Val Ser Gin Phe Asp Lys Val lie His Glu Thr Lys Lys Asp Met Gin 465 470 475 480

Ala Ser Lys Leu lie Asn Ala lie Val Gly His Ala Gly Asp Gly Asn 485 490 495

Phe His Ala Phe lie Val Tyr Arg·Thr Pro Glu Glu His Glu Thr Cys 500 505 510

Ser Gin Leu Val Asp Arg Met Val Lys Arg Ala Leu Asn Ala Glu Gly 515 520 525

Thr Cys Thr Gly Glu His Gly Val Gly lie Gly Lys Arg Glu Tyr Leu 530 535 540

Leu Glu Glu Leu Gly Glu Ala Pro Val Asp Leu Met Arg Lys lie Lys 545 550 B55 560

Leu Ala lie Asp Pro Lys Arg lie Met Asn Pro Asp Lys lie Phe Lys 565 570 575

Thr Asp Pro Asn Glu Pro Ala Asn Asp Tyr Arg 580 585

&lt;210&gt; 105 &lt;211&gt; 477 &lt;212&gt; PRT &lt;213&gt;&gt; Q5FP89 - Gluconobacter oxydans &lt;400&gt; 105 141 201127961

Met Pro Glu Pro Val Met Thr Ala Ser Ser Ala Ser Ala Pro Asp Arg 1 5 10 15

Leu Gin Ala Val Leu Lys Ala Leu Gin Pro Val Met Gly Glu Arg lie 20 25 30

Ser Thr Ala Pro Ser Val Arg Glu Glu His Ser His Gly Glu Ala Met 35 40 45

Asn Ala Ser Asn Leu Pro Glu Ala Val Val Phe Ala Glu Ser Thr Gin 50 55 60

Asp Val Ala Thr Val Leu Arg His Cys His Glu Trp Arg Val Pro Val 65 70 75 80

Val Ala Phe Gly Ala Gly Thr Ser Val Glu Gly His Val Val Pro Pro 85 90 95

Glu Gin Ala lie Ser Leu Asp Leu Ser Arg Met Thr Gly lie Val Asp 100 105 110

Leu Asn Ala Glu Asp Leu Asp Cys Arg Val Gin Ala Gly lie Thr Arg 115 120 125

Gin Thr Leu Asn Val Glu lie Arg Asp Thr Gly Leu Phe Phe Pro Val 130 135 140

Asp Pro Gly Gly Glu Ala Thr lie Gly Gly Met Cys Ala Thr Arg Ala 145 150 155 160

Ser Gly Thr Ala Ala Val Arg Tyr Gly Thr Met Lys Glu Asn Val Leu 165 170 175

Gly Leu Thr Val Val Leu Ala Thr Gly Glu lie lie Arg Thr Gly Gly 180 185 190

Arg Val Arg Lys Ser Ser Thr Gly Tyr Asp Leu Thr Ser Leu Phe Val 195 200 205

Gly Ser Glu Gly Thr Leu Gly lie He Thr Glu Val Gin Leu Arg Leu 210 215 220

His Gly Arg Pro Asp Ser Val Ser Ala Ala lie Cys Gin Phe Glu Ser 225 230 235 240 142 201127961

Leu His Asp Ala lie Gin Thr Ala Met Glu lie lie Gin Cys Gly lie 245 250 255

Pro lie Thr Arg Val Glu Leu Met Asp Ser Val Gin Met Ala Ala Ser 260 265 270 lie Gin Tyr Ser Gly Leu Asn Glu Tyr Gin Pro Leu Thr Thr Leu Phe 275 280 285

Phe Glu Phe Thr Gly Ser Pro Ala Ala Val Arg Glu Gin Val Glu Thr 290 295 300

Thr Glu Ala lie Ala Ser Gly Asn Asn Gly Leu Gly Phe Ala Trp Ala 305 310 315 320

Glu Ser Pro Glu Asp Arg Thr Arg Leu Trp Lys Ala Arg His Asp Ala 325 330 335

Tyr Trp Ala Ala Lys Ala lie Val Pro Asp Ala Arg Val lie Ser Thr 340 345 350

Asp Cys lie Val Pro lie Ser Arg Leu Gly Glu Leu lie Glu Gly Val 355 360 365

His Arg Asp lie Glu Ala Ser Gly Leu Arg Ala Pro Leu Leu Gly His 370 375 380

Val Gly Asp Gly Asn Phe His Thr Leu lie lie Thr Asp Asp Thr Pro 385 390 395 400

Glu Gly His Gin Gin Ala Leu Asp Leu Asp Arg Lys lie Val Ala Arg 405 410 415

Ala Leu Ser Leu Asn Gly Ser Cys Ser Gly Glu His Gly Val Gly Met 420 425 430

Gly Lys Leu Glu Phe Leu Glu Thr Glu His Gly Pro Gly Ser Leu Ser 435 440 445

Val Met Arg Ala Leu Lys Asn Thr Met Asp Pro His His lie Leu Asn 450 455 460

Pro Gly Lys Leu Leu Pro Pro Gly Ala Val Tyr Thr Gly 143 201127961 465 470 475 &lt;210&gt; 106 &lt;211&gt; 433

&lt;212&gt; PRT &lt;213&gt; EC 1.1.99.2 2-Pentylglutarate dehydrogenase &gt; Q9N420 - Nematode (Caenorhabditis elegans) &lt;400&gt; 106

Met Leu Asn Arg Gly Thr Phe Gin Val Phe Arg Gly lie Ser Gly Pro 15 10 15

Pro Lys Lys Ser Val Asp Leu Pro Lys Tyr Asp Leu Val lie Val Gly 20 25 30

Gly Gly lie Val Gly Cys Ala Thr Ala Arg Gin Leu Leu lie Glu Lys 35 40 45

Pro Gin Leu Lys Val Ala Leu lie Glu Lys Glu Lys Glu Leu Ala Val 50 55 60

His Gin Ser Gly His Asn Ser Gly Val lie His Ala Gly lie Tyr Tyr 65 70 75 80

Thr Pro Gly Ser Leu Lys Ala Lys Leu Cys Val Glu Gly Leu Asp Leu 85 90 95

Ser Tyr Glu Phe Phe Asp Lys Glu Lys Val Pro Tyr Lys Lys Thr Gly 100 105 110

Lys Leu lie Val Ala Val Glu Pro Glu Glu Val Pro Arg Leu Asp Ala 115 120 125

Leu Phe Ser Arg Ala Gin Thr Asn Gly Cys Arg Asp lie Glu Met lie 130 135 140

Asp Ser Ser Lys lie Thr Glu Leu Glu Pro His Cys Arg Gly Leu Lys 145 150 155 160

Ala Leu Trp Ser Pro His Thr Gly lie Val Asp Trp Gly Tyr Val Thr 165 170 175

Lys Arg Phe Gly Glu Asp Phe Glu Lys Arg Gly Gly Lys lie Tyr Thr 180 185 190

Ser Tyr Pro Leu Glu Lys lie Ser Asp Asn His Asp Pro Gly Tyr Pro 144 201127961 195 200 205 lie Arg Val Ser Ser Gly Pro Ala Leu Ala Glu Phe Glu Thr Lys Asn 210 215 220

Leu lie Thr Cys Ala Gly Leu Gin Ser Asp Arg Val Ala Ala Leu Ser 225 230 235 240

Gly Cys Ser Thr Asp Pro Lys lie Val Pro Phe Arg Gly Glu Tyr Leu 245 250 255

Leu Leu Lys Pro Glu Lys Arg His Leu Val Lys Thr Asn lie Tyr Pro 260 265 270

Val Pro Asp Pro Arg Phe Pro Phe Leu Gly Val His Phe Thr Pro Arg 275 280 285

Met Asn Gly Asp lie Trp Leu Gly Pro Asn Ala Val Leu Ala Tyr Lys 290 295 300

Arg Glu Gly Tyr Ser Tyr Phe Ser lie Ser Pro Ser Asp Leu Leu Glu 305 310 315 320

Ser Leu Ser Tyr Ser Gly Met Gin Lys Leu Val Lys Lys His Phe Thr 325 330 335

Phe Gly lie Lys Glu Leu Tyr Arg Gly Val Trp lie Ala Ala Gin Val 340 345 350

Lys Gin Leu Gin Arg Phe lie Pro Glu Leu Lys Leu Ser Asp Val Thr 355 360 365

Arg Gly Pro Ala Gly Val Arg Ala Gin Ala Met Asp Ser Ala Gly Asn 370 375 380

Leu Val Asp Asp Phe Val Phe Asp Ser Gly Thr Gly Lys Leu Ser Pro 385 390 395 400

Leu Leu Met His Val Arg Asn Ala Pro Ser Pro Ala Ala Thr Ser Ser 405 410 415

Leu Ala lie Ala Lys Met lie Thr Ser Glu Ala lie Asn Arg Phe Lys 420 425 430 ε 145 201127961

Leu &lt;210&gt; 107 &lt;211&gt; 455

&lt;212&gt; PRT &lt;213&gt;&gt; Q9VJ28 - Drosophila melanogaster &lt;400&gt; 107

Met Ala Gin Val Arg Leu Leu Val Gin Gly Leu Arg Arg Ser Leu Leu 15 10 IS

Asn Val Gly Val Ala Ala Pro Asn Glu Ser Thr Ala Thr His Lys Arg 20 25 30

Ser Gin His Ser Ser Ser Sers Cys Gly Asp Tyr Asp Leu Val Val Val 35 40 45

Gly Gly Gly lie Val Gly Ala Ala Ser Ala Arg Glu lie Val Leu Arg 50 55 60

His Pro Ser Leu Lys Val Ala Val Leu Glu Lys Glu Cys Lys Leu Ala 65 70 75 80

Lys His Gin Ser Gly His Asn Ser Gly Val lie His Ala Gly lie Tyr 85 90 95

Tyr Lys Pro Gly Thr Leu Lys Ala Arg Leu Cys Val Glu Gly Met His 100 105 110

Leu Ala Tyr Ala Tyr Leu Asp Glu Lys Lys lie Pro Tyr Lys Lys Thr 115 120 125

Gly Lys Leu lie Val Ala Thr Asp Glu Lys Glu Val Lys Leu Leu Lys 130 135 140

Asp Leu Glu Lys Arg Gly He Ala Asn Asn Val Pro Asp Leu Arg Met 145 150 155 160 lie Glu Gly Ser Glu lie Gin Glu lie Glu Pro Tyr Cys Gin Gly Val 165 170 175

Met Ala Leu His Ser Pro His Thr Gly lie Val Asp Trp Gly Leu Val 180 185 190

Thr Glu His Tyr Gly Gin Asp Phe Lys Gin Cys Gly Gly Asp lie Tyr 146 201127961 195 200 205

Leu Asp Phe Asn Val Ser Lys Phe Thr Glu Thr Lys Glu Gly Thr Asp 210 215 220

Tyr Pro Val Thr lie His Gly Ala Lys Pro Gly Gin Thr Val Arg Thr 225 230 235 240

Lys Asn Val Leu Thr Cys Gly Gly Leu Gin Ser Asp Leu Leu Ala Glu 245 250 255

Lys Thr Gly Cys Pro Arg Asp Pro Arg lie Val Pro Phe Arg Gly Glu 260 265 270

Tyr Leu Leu Leu Thr Lys Glu Lys Gin His Met Val Lys Gly Asn lie 275 280 285

Tyr Pro Val Pro Asp Pro Arg Phe Pro Phe Leu Gly Val His Phe Thr 290 295 300

Pro Arg Met Asp Gly Ser lie Trp Leu Gly Pro Asn Ala Val Leu Ala 305 310 315 320

Leu Lys Arg Glu Gly Tyr Thr Trp Gly Asp lie Asn Leu Phe Glu Leu 325 330 335

Phe Asp Ala Leu Arg Tyr Pro Gly Phe Val Lys Met Ala Ser Lys Tyr 340 345 350

Lie Gly Phe Gly Leu Ser Glu Met Ser Lys Ser Trp Phe lie Asn Leu 355 360 36B

Gin lie Lys Ala Leu Gin Lys Tyr lie Pro Asp lie Thr Glu Tyr Asp 370 375 380 lie Gin Arg Gly Pro Ala Gly Val Arg Ala Gin Ala Met Asp Leu Asp 385 390 395 400

Gly Asn Leu Val Asp Asp Phe Val Phe Asp Arg Gly Gin Gly Ser Gly 405 410 415

Ala Leu Ala Lys Arg Val Leu His Cys Arg Asn Ala Pro Ser Pro Gly 420 425 430 147 201127961

Ala Thr Ser Ser Leu Ala lie Ala Lys Met lie Ala Asp Lys lie Glu 435 440 445

Asn Glu Phe Ser lie Gly Lys 450 455 &lt;210&gt; 108 &lt;211&gt; 321

&lt;2X2&gt; PRT &lt;213&gt; EC 1.1.1.27 - I*-lactate dehydrogenase &gt; P13714 - Bacillus subtilis &lt;400&gt;

Met Met Asn Lys His Val Asn Lys Val Ala Leu lie Gly Ala Gly Phe 15 l〇 15

Val Gly Ser Ser Tyr Ala Phe Ala Leu lie Asn Gin Gly lie Thr Asp 20 25 30

Glu Leu Val Val lie Asp Val Asn Lys Glu Lys Ala Met Gly Asp Val 35 40 45

Met Asp Leu Pro His Gly Lys Ala Phe Gly Leu Gin Pro Val Lys Thr 50 55 60

Ser Tyr Gly Thr Tyr Glu Asp Cys Lys Asp Ala Asp lie Val Cys lie 65 70 75 80

Cys Ala Gly Ala Asn Gin Lys Pro Gly Glu Thr Arg Leu Glu Leu Val 85 90 95

Glu Lys Asn Leu Lys lie Phe Lys Gly lie Val Ser Glu Val Met Ala 100 105 110

Ser Gly Phe Asp Gly lie Phe Leu Val Ala Thr Asn Pro Val Asp lie 115 120 125

Leu Thr Tyr Ala Thr Trp Lys Phe Ser Gly Leu Pro Lys Glu Arg Val 130 135 140 lie Gly Ser Gly Thr Thr Leu Asp Ser Ala Arg Phe Arg Phe Met Leu 145 150 155 160

Ser Glu Tyr Phe Gly Ala Ala Pro Gin Asn Val His Ala His lie lie 165 170 175 148 201127961

Gly Glu His

Gly Asp Thr Glu Leu Pro Val Trp Ser His Ala Asn Val 180 185 190

Gly Gly Val 195

Pro Val Ser Glu Leu Val Glu Lys Asn Asp Ala Tyr Lys 200 205

Gin Glu Glu 210

Leu Asp Gin lie Val Asp Asp Val Lys Asn Ala Ala Tyr 215 220

His lie lie 225

Glu Lys Lys Gly Ala Thr Tyr Tyr Gly Val Ala Met Ser 230 235 240

Leu Ala Arg work le Thr Lys Ala lie Leu His Asn Glu Asn Ser lie Leu 245 250 255

Thr Val Ser

Thr Tyr Leu Asp Gly Gin Tyr Gly Ala Asp Asp Val Tyr 260 265 270 lie Gly Val 275

Pro Ala Val Val Asn Arg Gly Gly lie Ala Gly lie Thr 280 285

Glu Leu Asn 290

Leu Asn Glu Lys Glu Lys Glu Gin Phe Leu His Ser Ala 295 300

Gly Val Leu 305

Lys Asn lie Leu Lys Pro His Phe Ala Glu Gin Lys Val 310 315 320

Asn &lt;210&gt; 109 &lt;211&gt; 342 &lt;212&gt; PRT &lt;213&gt; EC 1 &lt;400&gt; 109 Met Thr His 1 1·1.28 - D-lactate dehydrogenase: &gt;Q88MCM_ Pseudomonas putida

Pro Arg His Ala Leu Gin Arg Ser Ser Thr Met Arg Ala 5 10 15

Leu Leu Phe

Ser Ser Gin His Tyr Asp Gin Glu Ser Phe Thr Lys Ala 20 25 30

Ala Gly Gly 35

Thr Ala Leu Glu Leu His Phe Gin Pro Ala Arg Leu Thr 40 45 149 201127961

Leu Asp Thr Ala Ala Leu Ala Asp Gly Phe Glu Val Val Cys Ala Phe 50 55 60 lie Asn Asp Glu Leu Asp Ala Pro Val Leu Gin Arg Leu Ala Ala Ala 65 70 75 80

Gly Thr Arg Leu lie Ala Leu Arg Ser Ala Gly Tyr Asn His Val Asp 85 90 95

Val Arg Val Pro Ala 110

Ala Leu lie Leu Ala 125

Leu Ala Ala Ala Gin Arg Leu Gly Leu Ala Val 100 105

Tyr Ser Pro His Ala Val Ala Glu His Ala Val 115 120

Leu Asn Arg Arg Leu His Arg Ala Tyr Asn Arg Thr Arg Glu Gly Asp 130 135 140

Phe Thr Leu His Gly Leu Thr Gly Phe Asp Leu His Gly Lys Thr Val 145 150 155 160

Gly Val Val Gly Thr Gly Gin lie Gly Val Ala Phe Ala Arg lie Met 165 170 17S

Ala Gly Phe Gly Cys Gin Leu Leu Ala Tyr Asp Pro Tyr Pro Asn Pro 180 185 190

Glu Leu Leu Ala Leu Gly Ala Arg Tyr Leu Pro Leu Pro Glu Leu Leu 195 200 205

Arg Glu Ala Arg lie lie Ser Leu His Cys Pro Leu Thr Glu His Thr 210 215 220

Arg His Leu lie Asn Ala Gin Ser Leu Ala Gin Leu Gin Pro Gly Ala 225 230 235 240

Met Leu lie Asn Thr Gly Arg Gly Ala Leu Val Asp Thr Pro Ala Leu 245 250 255 lie Asp Ala Leu Lys Ser Gly Gin Leu Gly Tyr Leu Gly Leu Asp Val 260 265 270

Tyr Glu Glu Glu Ala Gin Leu Phe Phe Glu Asp Arg Ser Asp Leu Pro 275 280 285 150 201127961

Leu Ala Arg Leu Leu Thr 295

Leu Gin Asp Asp Val 290

Phe Pro Asn Val lie 300 lie Thr Ala His Gin Ala Phe 305 310

Leu Thr Arg Glu Ala 315

Leu Asp Ala lie 320

Ala Ala Thr Thr Leu Asp Asn lie 325

Asn Arg 330

Trp Ala Ala Gly Asn Pro 335

Gin Asn Leu Val Met Gly 340 [SEQ ID NO: 110] Methanocaldococcus jannascMi (motif) aksA wild-type gene

ATGACAAAAGTGCTGGTGATGTTTATGGATTTCTTATTTGAGAACAGCTGGAAAGCAGTTTGTCCCTACAA

TCCAAAGTTGGATTTAAAGGACATTTATATTTATGACACAACCCTAAGAGATGGAGAGCAAACCCCAGGAG

TTTGCTTTACCAAAGAACAAAAATTGGAGATTGCAAGGAAGTTGGATGAACTTGGATTAAAGCAGATTGAA

GCTGGCTTCCCAATAGTATCTGAAAGAGAAGCAGATATAGTTAAAACAATTGCTAATGAAGGGCTAAATGC

TGATATCTTAGCTTTATGCAGGGCTTTAAAGAAAGATATAGATAAAGCAATAGAGTGCGATGTAGATGGGA

TTATTACCTTCATAGCAACATCTCCTCTCCACTTAAAATATAAATTCAACAACAAAAGCTTAGATGAAATA

TTAGAGATGGGAGTTGAGGCAGTTGAGTATGCAAAGGAACATGGCTTATTTGTTGCTTTCTCTGCAGAGGA

TGCGACAAGAACACCAATAGAGGACTTGATTAAAGTGCATAAAGCCGCTGAAGAGGCTGGAGCAGATAGGG

TTCATATAGCAGACACAACTGGCTGTGCTACCCCCCAAAGTATGGAGTTTATATGTAAAACATTGAAGGAG

AACTTAAAAAAGGCACATATTGGAGTGCATTGTCACAACGACTTTGGATTTGCAGTTATAAATTCAATATA

TGGTTTAATTGGAGGAGCTAAGGCAGTTTCAACAACAGTTAATGGTATTGGAGAGAGGGCAGGGAATGCAG

CTTTAGAAGAGCTAATTATGGCTTTAACTGTCTTGTATGATGTTGATTTGGGATTAAACTTGGAGGTTCTT

CCAGAGTTATGCAGAATGGTTGAGGAATACTCTGGAATAAAGATGCCAAAGAACAAACCAATAGTTGGAGA

GCTTGTATTTGCTCATGAAAGTGGAATTCACGTTGATGCTGTCATAGAGAATCCATTAACCTATGAACCCT

TCCTTCCAGAGAAAATAGGGCTTAAGAGAAATATTTTGTTAGGGAAGCATTCTGGATGCAGAGCCGTTGCC

TATAAGCTAAAACTTATGGGAATTGATTACGATAGAGAGATGTTGTGCGAGATTGTTAAAAAGGTTAAAGA

GATTAGAGAGGAAGGTAAATTTATAACTGATGAAGTCTTTAAGGAGATTGTTGAAGAAGTTTTAAGGAAGA

GAAATAAAAATTAA

[Serial No. 111] Metiianococcus jannaschii AksA, MJ0503

MTKVLVMFMDFLFENSWKAVCPYNPKLDLKDIYIYDTTLRDGEQTPGVCFTKEQKLEIARKLDELGLKQIE

AGFPIVSEREADIVKTIANEGLNADILALCRALKKDIDKAIECDVDGIITFIATSPLHLKYKFNNKSLDEI

LEMGVEAVEYAKEHGLFVAFSAEDATRTPIEDLIKVHKAAEEAGADRVHIADTTGCATPQSMEFICKTLKE

NLKKAHIGVHCHNDFGFAVINSIYGLIGGAKAVSTTVNGIGERAGNAALEELIMALTVLYDVDLGLNLEVL s 151 201127961

PELCRMVEEYSGIKMPKNKPIVGELVFAHESGIHVDAVIENPLTYEPFLPEKIGLKRNILLGKHSGCRAVA

YKLKLMGIDYDREMLCEIVKKVKEIREEGKPITDEVFKEIVEEVLRKRNKN

[SEQ ID NO: 112] Methanococcus jannasch: ii AksA, MJ0503 Codon Pair optimized gene,

ATGACCAAAGTTCTGGTAATGTTCATGGACTTCCTGTTCGAAAACTCCTGGAAAGCGGTTTGCCCGTACAA

CCCGAAACTGGATCTGAAAGACATCTACATCTACGACACCACTCTGCGTGACGGTGAACAGACTCCGGGCG

TTTGCTTCACCAAAGAGCAGAAGCTGGAAATCGCTCGTAAGCTGGACGAACTGGGTCTGAAGCAGATCGAA

GCTGGCTTCCCGATCGTTTCTGAACGTGAAGCTGACATCGTTAAAACTATCGCTAACGAAGGTCTGAACGC

TGACATCCTGGCACTGTGCCGTGCGCTGAAGAAAGACATCGACAAAGCAATCGAATGCGACGTTGACGGTA

TCATCACTTTCATCGCAACTTCTCCGCTGCACCTGAAATACAAATTCAACAACAAATCTCTGGATGAAATC

CTGGAAATGGGCGTTGAAGCGGTAGAATACGCTAAAGAGCACGGTCTGTTCGTTGCATTCTCTGCAGAAGA

TGCAACTCGTACTCCGATCGAAGATCTGATCAAAGTTCACAAAGCAGCTGAAGAAGCGGGTGCTGACCGCG

TTCACATCGCTGACACCACTGGCTGCGCAACTCCGCAGTCTATGGAATTCATCTGCAAAACTCTGAAAGAA

AACCTGAAGAAAGCACACATCGGCGTACACTGCCACAACGACTTCGGTTTCGCTGTTATCAACTCCATCTA

CGGTCTGATCGGTGGTGCGAAAGCGGTATCTACTACCGTTAACGGTATCGGTGAACGTGCTGGTAACGCTG

CACTGGAAGAGCTGATCATGGCGCTGACCGTACTGTACGACGTTGACCTGGGTCTGAACCTGGAAGTTCTG

CCGGAACTGTGCCGTATGGTTGAAGAATACTCCGGTATCAAGATGCCGAAAAACAAGCCAATCGTTGGTGA

ACTGGTATTCGCTCACGAATCCGGTATCCACGTTGACGCTGTTATCGAAAACCCGCTGACTTACGAACCGT

TCCTGCCGGAAAAAATCGGTCTGAAACGTAACATCCTGCTGGGTAAGCACTCTGGTTGCCGTGCTGTTGCT

TACAAGCTGAAACTGATGGGTATCGACTACGACCGTGAAATGCTGTGCGAAATCGTTAAGAAAGTTAAAGA

AATCCGTGAAGAAGGTAAATTCATCACTGACGAAGTTTTCAAAGAGATCGTTGAAGAAGTTCTGCGTAAGC

GTAACAAAAAC

[Serial No. II3] Methanocaldococcus jannaschii DSM 2661 (Phantom) Wild-type Gene

TTGACATTGGTAGAGAAGATACTATCAAAAAAAGTTGGTTATGAAGTTTGTGCAGGAGATAGCATAGAGGT

TGAAGTTGATTTGGCAATGACACACGATGGAACAACACCTTTAGCATACAAAGCTTTAAAGGAAATGAGTG

ATAGTGTTTGGAATCCAGATAAAATAGTCGTTGCCTTTGACCACAATGTTCCACCAAACACAGTTAAAGCT

GCTGAAATGCAAAAATTAGCTTTGGAGTTTGTTAAAAGATTTGGCATTAAAAATTTCCATAAAGGTGGAGA

AGGCATCTGTCATCAAATCTTAGCTGAAAATTATGTTTTGCCAAACATGTTTGTAGCTGGTGGAGACAGCC

ATACATGCACACATGGAGCTTTTGGAGCTTTTGCTACTGGCTTTGGAGCTACTGATATGGCTTACATCTAT

GCAACAGGAGAAACATGGATTAAAGTGCCAAAAACAATTAGGGTAGATATAGTTGGAAAAAATGAAAATGT

TTCTGCCAAAGATATTGTTTTAAGGGTTTGTAAGGAAATTGGGAGAAGAGGAGCAACATACATGGCTATTG

AGTATGGTGGAGAGGTTGTTAAAAACATGGACATGGATGGAAGGCTAACTTTATGCAACATGGCAATAGAG

ATGGGAGGAAAAACAGGAGTGATAGAGGCTGATGAAATTACTTATGATTATTTAAAGAAAGAGAGAGGACT

TTCTGATGAGGATATAGCTAAATTAAAAAAAGAGAGAATAACAGTAAATAGAGATGAAGCAAACTACTATA 152 201127961 aggagatagaaattgacataacagatatggaagaacaagttgctgttccacaccacccagataacgtaaag

CCAATTAGTGATGTTGAAGGGACTGAGATAAATCAAGTTTTTATTGGGAGTTGCACAAATGGAAGGTTGAG

TGATTTAAGAGAAGCAGCTAAATATTTAAAAGGTAGGGAGGTTCATAAAGATGTTAAGCTAATTGTTATCC

CGGCATCT^AAAAAGGTATTTTTGCAAGCGTTAAAAGAGGGTATTATAGATATCTTTGTTAAAGCTGGGGCG

ATGATTTGCACTCCGGGATGCGGACCTTGCTTAGGAGCTCATCAAGGGGTTTTGGCTGAGGGAGAAATTTG

TTTATCAACAACAAACAGAAACTTTAAAGGAAGGATGGGGCATATAAATAGCTATATTTACTTGGCATCTC

CAAAGATTGCCGCAATAAGTGCAGTTAAGGGATATATAACCAACAAATTGGATTAA

[Serial No. 114] Methanococcus j’annaschii AksD, MJ1003

MTLVEKILSKKVGYEVCAGDSIEVEVDLAMTHDGTTPLAYKALKEMSDSVWNPDKIWAFDHNVPPNTVKA

AEMQKIiALEFVKRFGIKNFHKGGEGICHQILAENYVLPNMFVAGGDSHTCTHGAFGAFATGFGATDMAYIY

ATGETWIKVPKTIRVDIVGKNENVSAKDIVLRVCKEIGRRGATYMAIEYGGEWKNMDMDGRLTLCNMAIE

MGGKTGVIEADEITYDYLKKERGLSDEDIAKLKKERITVNRDEANYYKEIEIDITDMEEQVAVPHHPDNVK

PISDVEGTEINQVFIGSCTNGRLSDLREAAKYLKGREVHKDVKLIVIPASKKVFLQALKEGIIDIFVKAGA

MICTPGCGPCLGAHQGVLAEGEICLSTTNRNFKGRMGHINSYIYLASPKIAAISAVKGYITNKLD

[SEQ ID NO: 115] (Methanococcus jannaschii) AksD, MJ1003 codon pair optimal 4匕 gene

ATGACTCTGGTTGAGAAGATCCTCTCCAAGAAAGTTGGTTACGAAGTTTGCGCAGGCGACTCCATCGAAGT

TGAAGTTGACCTGGCGATGACTCACGACGGTACTACTCCGCTGGCTTACAAAGCGCTGAAAGAGATGTCTG

ACTCCGTATGGAACCCGGACAAGATCGTTGTTGCATTCGACCACAACGTACCGCCGAACACCGTTAAAGCA

GCTGAAATGCAGAAGCTGGCGCTGGAATTCGTTAAGCGCTTCGGTATCAAAAACTTCCACAAAGGTGGTGA

AGGTATCTGCCACCAGATCCTGGCTGAAAACTACGTTCTGCCGAACATGTTCGTTGCTGGCGGCGACTCTC

ACACCTGTACTCACGGTGCATTCGGTGCATTCGCAACTGGCTTCGGTGCAACTGACATGGCTTACATCTAC

GCAACTGGCGAAACCTGGATCAAAGTTCCGAAAACTATCCGCGTTGATATCGTTGGTAAAAACGAAAACGT

ATCTGCGAAAGACATCGTTCTGCGCGTTTGCAAAGAAATCGGTCGTCGCGGTGCAACTTACATGGCTATCG

AATACGGTGGTGAAGTTGTTAAAAACATGGACATGGACGGTCGTCTGACTCTGTGCAACATGGCTATCGAA

ATGGGTGGTAAAACTGGCGTTATCGAAGCTGACGAAATCACTTACGACTACCTGAAGAAAGAGCGTGGTCT

GTCTGACGAAGATATCGCTAAACTGAAGAAAGAGCGTATCACCGTTAACCGTGACG7VAGCTAACTACTACA

AAGAAATCGAAATCGACATCACTGACATGGAAGAACAGGTTGCTGTACCGCACCACCCGGATAACGTTAAG

CCAATCTCTGACGTTGAAGGTACTGAAATCAACCAGGTATTCATCGGTTCCTGCACCAACGGTCGTCTGTC

TGATCTGCGTGAAGCTGCGAAATACCTGAAAGGTCGTGAAGTTCACAAAGACGTTAAGCTGATCGTTATCC cggcttccaagaaagtattcctgcaggcgctgaaagaaggtatcatcgacatcttcgttaaagcgggtgcg

ATGATCTGTACTCCGGGTTGCGGTCCGTGCCTGGGTGCACACCAGGGCGTACTGGCAGAAGGTGAAATCTG

CCTGTCTACTACCAACCGTAACTTCAAAGGTCGTATGGGTCACATCAACTCTTACATCTACCTGGCTTCTC

CGAAAATCGCTGCTATCTCTGCTGTTAAAGGTTACATCACTAACAAGCTGGAT

S 153 201127961 [Serial No. 116] Methanocaldococcus jamiaschii DSM 2661 (Module) Wild-type Gene

ATGATTATTAAGGGAAGAGCTCACAAATTTGGGGATGATGTAGATACAGACGCAATAATTCCAGGACCTTA

CTTAAGGACTACAGACCCTTACGAGTTAGCTTCACACTGCATGGCAGGGATAGATGAAAACTTCCCGAAAA

AGGTTAAGGAGGGGGATGTGATAGTTGCTGGAGAGAATTTTGGTTGTGGTTCAAGTAGGGAGCAGGCTGTA

ATAGCAATAAAATACTGTGGTATTAAGGCTGTGATAGCAAAAAGCTTTGCAAGAATATTCTATAGAAATGC

AATAAACGTTGGATTAATACCAATAATAGCAAATACAGATGAAATTAAAGACGGAGACATAGTAGAGATTG

ATTTAGATAAAGAAGAGATTGTAATAACCAATAAAAACAAAACAATAAAGTGTGAAACACCAAAAGGTTTA

GAAAGAGAAATATTGGCTGCTGGTGGCTTAGTCAATTATTTAAAAAAGAGAAAACTAATACAATCAAAAAA

AGGTGTAAAAACATGA

[Serial No. 117] Methanococcus jamiaschii AksE, MJ1271

MI IKGRAHKFGDDVDTDAII PGPYIiRTTDPYELASHCMAGIDENFPKKVKEGDVIVAGENFGCGSSREQAV IAIKYCGIKAVIAKSFARIFYRNAINVGLIPIIANTDEIKDGDIVEIDLDKEEIVITNKNKTIKCETPKGL ERE ILAAGGLVNYLKKRKLIQSKKGVKT

[Serial No. 118] Methanococcus jannaschii AksE, MJ1271 codon pair optimal 匕 gene

ATGATCATCAAAGGTCGTGCGCACAAGTTCGGTGACGACGTTGACACTGACGCTATCATCCCAGGTCCGTA

CCTCCGTACTACTGACCCGTACGAACTGGCATCTCACTGCATGGCGGGTATCGACGAAAACTTCCCGAAGA

AAGTTAAAGAAGGTGACGTTATCGTTGCTGGCGAAAACTTCGGTTGCGGTTCTTCCCGTGAGCAGGCTGTT

ATCGCTATCAAATACTGCGGTATCAAAGCGGTTATCGCTAAATCTTTCGCACGTATCTTCTACCGTAACGC

AATCAACGTAGGTCTGATCCCGATCATCGCTAACACCGACGAAATCAAAGACGGTGACATCGTTGAAATCG

ACCTGGATAAAGAAGAAATCGTTATCACTAACAAAAACAAAACTATCAAGTGCGAAACTCCGAAAGGTCTG

GAACGTGAAATCCTGGCAGCTGGCGGTCTGGTTAACTACCTGAAGAAACGTAAGCTGATTCAGTCCAAGAA

AGGCGTAAAAACT

[Serial No. 119] * Methanocaldococcus jannaschii DSM 2661 (morpho) wild-type gene

ATGATGAAGGTGTGTGTTATAGAAGGGGATGGAATAGGAAAAGAAGTGATTCCAGAGGCCATAAAAATATT

AAATGAGTTGGGAGAGTTTGAAATAATAAAAGGAGAGGCAGGATTAGAATGTTTAAAAAAATATGGTAATG

CACTTCCAGAGGATACAATAGAAAAAGCTAAAGAGGCAGATATTATTTTGTTTGGGGCTATAACCTCACCA

AAGCCAGGGGAAGTTCAAAATTATAAAAGCCCTATAATAACGTTGAGGAAGATGTTTCATTTATATGCAAA 154 201127961

TGTAAGACCAATAAACAACTTTGGAATTGGACAATTAATTGGGAAAATTGCAGATTATGAATTCTTAAATG

CTAAGAATATTGATATAGTTATTATAAGAGAGAATACGGAAGATTTATATGTTGGTAGAGAGAGATTAGAA

AATGATACAGCAATAGCTGAGAGGGTTATAACAAGAAAGGGTAGCGAGAGAATAATAAGATTTGCATTTGA

ATATGCTATAAAAAATAATAGGAAAAAGGTATCTTGCATCCATAAAGCTAATGTTTTAAGAATAACTGATG

GTTTATTCTTAGAGGTTTTTAATGAAATAAAAAAACATTATAATATAGAGGCAGATGATTATTTAGTTGAT

TCAACAGCTATGAACTTAATAAAACATCCTGAAAAATTTGATGTTATTGTTACAACAAACATGTTTGGGGA

TATTTTATCAGATGAGGCATCTGCATTAATTGGAGGACTTGGTTTAGCTCCTTCAGCAAATATAGGAGATG

ATAAAGCATTATTTGAGCCAGTTCATGGTTCAGCTCCAGATATAGCTGGGAAAGGTATAGCAAATCCAATG

GCATCTATATTAAGTATTGCTATGCTTTTTGATTATATTGGAGAGAAAGAAAAGGGAGATTTGATTAGAGA

GGCAGTGAAATACTGCTTAATAAACAAAAAAGTTACTCCTGACTTGGGAGGGGATTTAAAGACAAAAGATG

TTGGAGACGAAATTCTAAATTACATTAGAAAGAAGTTAAAGGGATATTGA

[Serial No. 120] Aiethanococcus jannaschii AksF, MJ1596

MMKVCVIEGDGIGKEVIPEAIKILNELGEFEIIKGEAGLECLKKYGNALPEDTIEKAKEADIILFGAITSP

KPGEVQNYKSPIITLRKMFHLYANVRPINNFGIGQLIGKIADYEFLNAKNIDIVIIRENTEDLYVGRERLE

NDTAIAERVITRKGSERIIRFAFEYAIKNNRKKVSCIHKANVLRITDGLFLEVFNEIKKHYNIEADDYL'VD

STAMNLIKHPEKFDVIVTTNMFGDILSDEASALIGGLGLAPSANIGDDKALFEPVHGSAPDIAGKGIANPM

ASILSIAMLFDYIGEKEKGDLIREAVKYCLINKKVTPDLGGDLKTKDVGDEILNYIRKKLKGY

[Serial No. 121] Methanococcus j’annaschii AksF, MJ1596 Mimazi Optimized Gene

ATGATGAAAGTTTGCGTTATCGAAGGTGACGGTATCGGTAAAGAAGTTATCCCGGAAGCTATCAAGATCCT

GAACGAACTGGGTGAATTCGAAATCATCAAAGGTGAAGCGGGTCTGGAATGCCTGAAGAAATACGGTAACG

CACTGCCAGAAGATACCATCGAAAAAGCGAAAGAAGCTGACATCATCCTGTTCGGTGCAATCACTTCTCCG

AAGCCGGGTGAAGTTCAGAACTACAAATCTCCGATCATCACTCTGCGTAAGATGTTCCACCTGTACGCTAA

CGTACGTCCGATCAACAACTTCGGTATCGGTCAGCTGATCGGTAAGATCGCTGACTACGAGTTCCTGAACG

CTAAAAACATCGACATCGTTATCATCCGTGAAAACACTGT^AGATCTGTACGTTGGTCGTGAACGTCTGGAA

AACGACACTGCTATCGCTGAGCGCGTTATCACTCGTAAAGGTTCTGAACGTATCATCCGCTTCGCATTCGA

ATACGCAATCAAAAACAACCGTAAGAAAGTTTCCTGCATCCACAAAGCTAACGTACTGCGTATCACTGACG

GTCTGTTCCTGGAAGTATTCAACGAAATCAAGAAACACTACAACATCGAAGCTGACGACTACCTGGTTGAC

TCCACTGCAATGAACCTGATCAAGCACCCGGAAAAATTCGACGTTATCGTTACCACTAACATGTTCGGTGA

CATCCTGTCTGACGAAGCGTCTGCACTGATCGGTGGTCTGGGTCTGGCACCGTCTGCTAACATCGGTGACG

ACAAAGCGCTGTTCGAACCGGTTCACGGTTCTGCACCGGATATCGCTGGTAAAGGTATCGCTAACCCGATG

GCTTCTATCCTGTCTATCGCGATGCTGTTCGACTACATCGGTGAAAAAGAGAAAGGCGACCTGATCCGTGA

AGCGGTAAAATACTGCCTGATCAACAAGAAAGTTACTCCGGATCTGGGTGGTGACCTGAAAACCAAAGACG

TTGGTGACGAAATCCTGAACTACATCCGTAAGAAACTGAAAGGTTAC 155 201127961 [Serial No. 122] Methanococcus vannieim SB AksF, Mevan—0040 Wild type AAAAAG&quot;! AAATTT1 AAAAAT7 AACAGC7 GAAATAi ATAAAAl

ATGGGCTATA

CCTGAAACAT

GGGTACGAGT

AAAAATTCTG

AATAAGCCAT

ATACGGCCCA

TGTCTTTACG

ATAATTTCAA

AATAATCGAA

TTATTTTTGG

AATGATTACT

GTAATGGTTA

GGGGGTCTTG

GTTCATGGTT

TTAAGTGCTT

AATGCAGTTA

AAAACTTCCG

TGCCAAAAAT TAAGAGTTTT GTTTTAAGAG ATTCAATTCT ATCGTAGTCC CTTATAATTT TAAAAAGGGA AAAAGGGAAG lGTGTC 'TCGA TAATAGATGC CAACAAACCT GAATGTCGCC CAGCCCCAGA CAATGATGCT AAAAAACGAT AAGTTGTAAA

CTGTGTCATA AAACGAAGTT ATGCGGAGAA TTTTGGTTCA AATACTTACG TAAAGACTTG GTATTATGAC CGAAAGAATA TTGCATACAC 'AGCT !AATG TTTTGGAGAT TTCTGCAAAT TATTGCTGGA TGACCATTTA AAATAATGGT TAAAGTTATA

ACTGGTGATG CACGACTTTG TCAATACCTG GTAACTACTC TTAAGGCAGG GATTTTGTCA 'AAACG •TTTG AAAGCGAATG AAACTTTATG TACCTTATTA ATTTTATCTG ATTGGGGATA AAAGGAATAT AAAATGAATA TATTTGACAC GAATTTATTC

GAATTGGAAA

AATATATTGA

AAAGCACGAT

CAAAACCAAC

AATTAGACCT

TAATACGGGA

AAGTAGCAAT

CATTTGAATA

TATTAAGAGT

AAAACTTTGG

AAAATCCATA

ATGAGGCCGC

ATTTAGGATT

CTAATCCGAT

AAAAGGCGGA

CCGATCTTGG

GGGATGAAAT

AGAAGTCGTG

AGCCCATGCT

TCAAACAGCT

TGAATTAAAA

TTATGCAAAC

AAATACTGAG

TGCCGAAAGA

TGCAAGGTTA

AACTGACGGA

TATATCGAGC

TATGTTTGAT

AGGACTTATT

ATTTGAGCCT

TGCGACAATT

AATTATAAGA

TGGAAGCCTG

CTAA

[Serial No. l23] Methanococcus vannielii SB AksF, Mevan 0040

MGYMPKICVITGDGIGKEWPETLRVLNEVHDFEYIEAHAGYECFKRCGESIPESTIQTAKNSDSILFGSV

TTPKPTELKNKPYRSPILTLRQELDLYANIRPTYNFKDLDFVIIRENTECLYVKREYYDEINEVAIAERII

SKKGSERIIKFAFEYARLNNRKKVSCIHKANVLRVTDGLFLEIFEKIAKLYENFGISSNDYLIDATAMYLI

KNPYMFDVMVTTNLFGDILSDEAAGLIGGLGMSPSANIGDNLGLFEPVHGSAPDIAGKGISNPIATILSAS MMLDHLKMNKKAEIIRNAVKKTINNGYLTPDLGGSLKTSEWNKVIEFIRDEI [SEQ ID NO: 124] Codon pair optimization gene, Mevan_0040

ATGGGTTACATGCCGAAAATCTGCGTTATCACTGGCGACGGTATCGGTAAAGAAGTTGTTCCGGAAACTCT

GCGCGTACTGAACGAAGTTCACGACTTCGAATACATCGAAGCACACGCGGGTTACGAGTGCTTCAAGCGCT

GCGGTGAATCCATCCCGGAATCCACTATTCAGACTGCGAAAAACTCTGACTCCATCCTGTTCGGTTCTGTT

ACCACTCCGAAACCAACTGAACTGAAAAACAAGCCGTACCGCTCTCCGATTCTGACTCTGCGTCAGGAACT

GGATCTGTACGCTAACATCCGTCCGACTTACAACTTCAAAGACCTGGACTTCGTTATCATCCGTGAAAACA

CTGAATGCCTGTACGTTAAGCGTGAATACTACGACGAAATCAACGAAGTTGCTATCGCTGAACGTATCATC

TCCAAGAAAGGTTCTGAACGTATCATCAAATTCGCTTTCGAATACGCACGTCTGAACAACCGTAAGAAAGT

TTCCTGCATCCACAAAGCTAACGTACTGCGCGTAACTGACGGTCTGTTCCTGGAAATCTTCGAGAAGATCG

CGAAACTGTACGAAAACTTCGGTATCTCTTCTAACGACTACCTGATCGACGCAACTGCAATGTACCTGATC

AAAAACCCGTACATGTTCGACGTAATGGTTACCACTAACCTGTTCGGTGACATCCTGTCTGACGAAGCTGC

TGGTCTGATCGGTGGTCTGGGTATGTCTCCGTCTGCTAACATCGGTGACAACCTGGGTCTGTTCGAACCGG 156 201127961

TTCACGGTTCTGCACCGGATATCGCTGGTAAAGGTATCTCCAACCCGATCGCGACTATCCTGTCTGCGTCT

ATGATGCTGGATCACCTGAAAATGAACAAGAAAGCAGAAATCATCCGTAACGCTGTTAAGAAAACTATCAA

CAACGGTTACCTGACTCCGGACCTGGGTGGTTCTCTGAAAACTTCTGAAGTTGTTAACAAAGTTATCGAAT

TCATCCGCGACGAGATT

[SEQ ID 125] Martens Yue alkoxy ί request bacteria (Methaiiococrcus maripaludis) S2 AksF, MMP0880 wild-type gene atgagaaacactcccaaaatttgtgttatcaatggtgacggtattggaaacgaagtagttcctgaaacggt gcgagttttaaatgaacttggtgacttcgaattcattcatgcccatgcaggttacgaatgttttaaaagat gtggcgatgcgataccagaaaacacaattgaaattgcaaaagaatctgattgtattttatttggatcagtt accactccaaaaccgactgaattaaaaaataaatcatatagaagtccaatattaactttaagaaaagaact tgacctttatgcaaatattaggccaacttataactttgataatcttgattttgttataattcgagaaaata ctgaaggactctatgtaaaaaaagaatattacgacgaaaaaaacgaagttgcaattgctgagcgaataatt tcaaaatttggaagttctagaattgtaaaatttgcttttgattatgcggttcaaaataacagaaaaaaagt atcctgcatacataaagcaaacgtattacgggttactgacggattatttttagaagttttcgaagaaatgt ctaaacattacgaaaaattaggaataaagtctgatgactacctaattgacgcgacagoaatgtatttgatt agaaacccgcaaatgtttgatgtattggttacaacaaatctttttggagatattttatctgatgaagctgc aggacttattggcggacttggaatgtctccttcagcaaacattggtgataaaaacggattatttgagccag ttcatggatctgcaccagacattgctggaaaaggaatttcaaacccgattgcaacaatattgagtgctgca atgatgcttgaccatttaaaaatgaataaagaagccg Aatacattagaaaagcggttaaaaaaacggttga atgtaaatatttaactcctgatcttgggggaaacttaaaaacttttgaagttacggaaaaaatcattgaat ccataaggtctcagatgattcagtga [serial number 126] 甲甲甲菌菌 fAiethanococcus maripaludis) S2 AksF, MMP0880

MRNTPKICVINGDGIGNEWPETVRVLNELGDFEFIHAHAGYECFKRCGDAIPENTIEIAKESDCILFGSV

TTPKPTELKNKSYRSPILTLRKELDLYANIRPTYNFDNLDFVIIRENTEGLYVKKEYYDEKNEVAIAERII

SKFGSSRIVKFAFDYAVQNNRKKVSCIHKANVLRVTDGLFLEVFEEMSKHYEKLGIKSDDYLIDATAMYLI

RNPQMFDVLVTTNLFGDILSDEAAGLIGGLGMSPSANIGDKNGLFEPVHGSAPDIAGKGISNPIATILSAA

MMLDHLKMNKEAEYIRKAVKKTVECKYLTPDLGGNLKTFEVTEKIIESIRSQMIQ

[SEQ ID NO: 127] Codon pair optimization gene, MMP0880

ATGCGTAACACTCCGAAAATCTGCGTTATCAACGGTGACGGTATCGGTAACGAAGTTGTTCCGGAAACCGT

TCGCGTACTGAACGAACTGGGTGACTTCGAATTCATCCACGCGCACGCTGGTTACGAATGCTTCAAGCGCT

GCGGTGACGCTATCCCGGAAAACACCATCGAAATCGCTAAAGAGTCTGACTGCATCCTGTTCGGTTCTGTA

ACTACTCCGAAACCAACTGAACTGAAAAACAAGTCTTACCGCTCTCCGATTCTGACTCTGCGTAAAGAGCT

GGATCTGTACGCTAACATCCGTCCGACTTACAACTTCGACAACCTGGATTTCGTTATCATCCGTGAAAACA

CTGAAGGTCTGTACGTTAAGAAAGAATACTACGACGAGAAAAACGAAGTTGCTATCGCTGAACGTATCATC

TCCAAGTTCGGTTCTTCTCGCATCGTTAAATTCGCATTCGACTACGCAGTACAGAACAACCGTAAGAAAGT

TTCCTGCATCCACAAAGCGAACGTTCTGCGCGTAACTGACGGTCTGTTCCTGGAAGTTTTCGAAGAAATGT

CCAAGCACTACGAAAAACTGGGTATCAAATCTGACGACTACCTGATCGACGCAACTGCGATGTACCTGATC 157 201127961

CGTAACCCGCAGATGTTCGACGTTCTGGTTACTACCAACCTGTTCGGTGACATCCTGTCTGACGAAGCAGC

TGGTCTGATTGGTGGTCTGGGTATGTCTCCGTCTGCTAACATCGGTGACAAAAACGGTCTGTTCGAACCGG

TTCACGGTTCTGCACCGGATATCGCTGGTAAAGGTATCTCCAACCCGATCGCGACTATCCTGTCTGCTGCA

ATGATGCTGGATCACCTGAAAATGAACAAAGAAGCTGAATACATCCGTAAAGCGGTTAAGAAAACCGTTGA

ATGCAAATACCTGACTCCGGACCTGGGTGGTAACCTGAAAACTTTCGAAGTTACTGAAAAGATCATCGAAT CCATCCGTTCTCAGATGATTCAA [Serial No. 128] Methanococcus maripaludisj S2 AksE, MMP0381 wild type

ATGAAAATAA CTGGTAAGGT GCACTTATTT GGGGATGACA TCGATACTGA TGCGATAATT CCCGGAGCTT ATTTAAAAAC GACTGATGAA TATGAGCTTG CATCGCACTG TATGGCAGGA ATTGACGAAA ATTTTCCAGA AAGGGTCGAA GATGGTGACT TTTTAGTTGC AGGTGAAAAT TTTGGATGCG GAAGTTCAAG GGAACAGGCC CCAATTGCCA TAAAATACTG CGGAATCAAG GCAATAATTG TTGAGAGTTT TGCAAGGATA TTTTACAGAA ATTGCATAAA TTTAGGAGTA TTTCCAATTG AATGCAAGGG AATATCAAAA CACGTCAAAG ATGGGGATGT AATAGAATTA GATCTTGAAG AAAAAAAAGT TATCTTAAAA GACACGGTTC TTGACTGCAA TCTTCCGACA GGGACTGCAA AAGATATAAT GGATGAAGGC GGGCTTATAA ATTACGCAAA GAAACAAAAA AATTAA

[Serial No. 129] Helicobacter punctatus maripaJudis) 52 AksE, MMP0381

MKITGKVHLFGDDIDTDAIIPGAYLKTTDEYELASHCMAGIDENFPERVEDGDFLVAGENFGCGSSREQAP

IAIKYCGIKAIIVESFARIFYRNCINLGVFPIECKGISKHVKDGDVIELDLEEKKVILKDTVLDCNLPTGT

AKDIMDEGGLINYAKKQKN

[SEQ ID NO: 130] Codon pair optimization gene, MMP0381

ATGAAGATCACCGGTAAAGTTCACCTGTTCGGTGACGACATCGACACTGACGCTATCATTCCGGGTGCTTA

CCTGAAAACCACTGACGAATACGAACTGGCTTCTCACTGCATGGCGGGTATCGACGAAAACTTCCCGGAAC

GCGTTGAAGATGGCGACTTCCTGGTTGCTGGCGAAAACTTCGGTTGCGGTTCTTCCCGTGAACAGGCACCG

ATTGCTATCAAATACTGCGGTATCAAAGCAATCATCGTTGAATCCTTCGCACGTATCTTCTACCGTAACTG

CATCAACCTGGGCGTATTCCCGATCGAATGCAAAGGTATCTCCAAGCACGTTAAAGACGGTGACGTTATCG

AACTGGATCTGGAAGAGAAGAAAGTTATCCTGAAAGACACCGTACTGGACTGCAACCTCCCGACTGGTACT

GCGAAAGATATCATGGACGAAGGTGGTCTGATCAACTACGCTAAGAAGCAGAAAAAC

[Serial No. 131] Methanococcus maripaludisj S2 AksD# MMP1480 wild type

ATGACACTTG CTGAGAAAAT CATTTCAAAA AATGTTGGAA AAAATGTTTA CGCCAAAGAC AGCGTCGAAA TAAGCGTAGA TATTGCAATG ACACATGACG GGACCACCCC GCTTACGGTA AAAGCCTTTG AGCAGATTTC AGATAAAGTA TGGGATAATG AAAAGATAGT CATTATTTTT 158 201127961

GACCACAATA

TTCATAAAAA

GTTCTACCTG

ACATGTACTC

TACGTCTATG

ACTGGAGAAA

GGAAGACGTG

TCAATGGATG

ATTATCGAAG

GAAATTCTTG

AAAACAATTG

GATAACGTAA

TCATGCACAA

AAAGTTAATG

GCCCTAAATG

GGATGCGGAC

GCTACAACTA

TCTTCTCCAA

TCCCTGCAAA

AGCAGGGGAT

AAAAGGGCCA

ATGGGGCCTT

CAACAGGAAA

ATGAAAATAT

GGGCTACGTA

AAAGAATGGT

CTGATGATAC

AATTGAAAAA

AATTTGACAT

AAGGAGTTTC

ACGGAAGATT

AAAACACAAG

AAGGATTAAT

CATGTCTTGG

ACCGGAACTT

AAATAGCTGC

CACGTCAAAA

TAAAAATTAC

TGTAAAACCA

TGGAGCATTT

AACCTGGCTT

TTCTGGAAAA

CATGTCTTTA

TCTGTCAAAC

TACATATAGA

AAATAAAATA

AACCGGTATG

AGAAGTTGAA

AAACGACTTA

ATTAA.TTGTA

TGATATCTTT

AGCCCATCAG

TAAAGGAAGA

AAAATCTGCG

GCTGCAAATA

TACCTCGATG

AACATGATAA

GCTACAGGTT

AGAGTTCCTG

GACATTATTT

GAATACGGTG

ATGGCTATTG

TATCTTGAAA

ACAGTTGATG

GAAGAACAGG

GGAACAGAAT

AGAATTGCTG

ATCCCTGCAT

GTAGATTCCG

GGGGTTTTAG

ATGGGAAACA

GTTAAAGGCT

TGCAGGTTAT

GCGAAGGAAT

TTGCGGGAGC

TTGGTGCTAC

AAACTATCCG

TAAAAACTTG

GAAATGCAGT

AAATGGGCGG

ATGCAGGAGT

AATCCGAAGA

TTGCATGCCC

TAAACCAGGT

CAAAATATTT

CAAAGTCAAT

GAGCATTAAT

GTGATGGAGA

CGAACGCACA

ACATTACAAA

AACGAGAGAA

ATGCCATCAA

TGACAGCCAC

AGACATGGGT

CGTAAATGTA

TAAGGAAGTT

CCACAATCTT

AAAAGCAGGA

TTCGCGCGAA

AGACTACTAC

TCACCACCCT

ATTCATCGGT

GAAAGGAAAA

ATTTAAAGAA

ATGTACCCCT

AGTATGCCTT

AGTTTACCTC

TGAATAA

[Serial No. 132] (Methanococcus maripaludis) S2 AksD, MMP1480

MTLAEKIISKNVGKNVYAKDSVEISVDIAMTHDGTTPLTVKAFEQISDKVWDNEKIVIIFDHNIPANTSKA ANMQVITREFIKKQGIKNYYLDGEGICHQVLPEKGHVKPNMI work AGADSHTCTHGAFGAFATGFGATDMGYV YATGKTWLRVPETIRAmVTGENENISGKDIILKTCKEVGRRGATYMSLEYGGNAVHNLSMDERMVLSNMAI EMGGKAGIIEADDTTYRYLENAGVSREEILELKKNKITVDESEEDYYKTIEFDITGMEEQVACPHHPDNVK ανβΕνΕαΤΕΙ ^ ΝΟνΡίσβσΓΝαίΙΙιΝϋΙιΙΙΙΑΑΚΥΙ ^ ΚΚνΝΕΝΤΙ ^ ΐνίΡΑβΚβΙΡΚΕΑΒΝΕαΒΙϋΙΡνϋβΟΑ LICTPGCGPCLGAHQGVLGDGEVCLATTNRNFKGRMGNTNAQVYLSSPKIAAKSAVKGYITNE

[SEQ ID NO: I33] Codon pair optimization gene, MMP1480

ATGACTCTGGCTGAGAAGATCATCTCCAAAAACGTTGGTAAAAACGTTTACGCGAAAGACTCCGTTGAAAT

CTCCGTTGACATCGCGATGACTCACGACGGTACTACTCCGCTGACCGTTAAAGCGTTCGAACAGATCTCTG

ACAAAGTATGGGATAACGAGAAGATCGTTATCATCTTCGACCACAACATCCCGGCTAACACCTCTAAAGCT

GCTAACATGCAAGTTATCACTCGTGAATTCATCAAGAAGCAGGGTATCAAAAACTACTACCTGGATGGTGA

AGGTATCTGCCACCAGGTACTGCCGGAAAAAGGTCACGTTAAGCCGAACATGATCATCGCTGGCGCAGACT

CTCACACTTGCACTCACGGTGCATTCGGTGCATTCGCTACCGGTTTCGGTGCAACTGACATGGGTTACGTT

TACGCAACTGGTAAAACCTGGCTGCGCGTACCGGAAACCATTCGCGTTAACGTAACTGGCGAAAACGAAAA

CATCTCCGGTAAAGACATCATCCTGAAAACTTGCAAAGAAGTTGGTCGTCGCGGTGCAACTTACATGTCTC

TGGAATACGGTGGTAACGCTGTTCACAACCTGTCTATGGACGAACGTATGGTTCTGTCTAACATGGCTATC

GAAATGGGTGGTAAAGCTGGTATCATCGAAGCTGACGACACCACTTACCGCTACCTGGAAAACGCTGGCGT

TTCCCGTGAAGAAATCCTGGAACTGAAGAAAAACAAGATCACCGTTGACGAATCTGAAGAAGATTACTACA

AAACTATTGAATTCGACATCACCGGTATGGAAGAACAGGTTGCTTGCCCACACCACCCGGACAACGTTAAA

GGCGTTTCTGAAGTTGAAGGTACTGAACTGAACCAGGTATTCATCGGTTCCTGCACCAACGGTCGTCTGAA 159 201127961

CGATCTGCGTATTGCTGCGAAATACCTGAAAGGTAAGAAAGTTAACGAAAACACCCGTCTGATCGTTATCC

CGGCATCTAAATCTATCTTCAAAGAAGCGCTGAACGAAGGTCTGATCGACATCTTCGTTGACTCCGGTGCA

CTGATCTGCACTCCGGGTTGCGGTCCGTGCCTGGGTGCACACCAGGGCGTTCTGGGTGACGGTGAAGTTTG

CCTGGCAACCACTAACCGTAACTTCAAAGGTCGTATGGGTAACACCAACGCTCAGGTTTACCTGTCCTCTC

CGAAGATCGCTGCGAAGTCTGCGGTAAAAGGTTACATCACTAATGAG

[SEQ ID 134] Ehrlich Yue burning meningitidis (A ^ etiianococcus aeo2icus) Nankai 3, aksE Maeo-0652, a wild-type atgataataaaaggaaatattcatttatttggtgatgatattgataccgatgccataattcccggggcct accttaaaacaacagacccaaaggagttggcatctcattgcatggctggaattgatgaaaaattttcaac aaaggtaaaagacggcgatataattgttgcaggtgaaaattttggctgtggaagtagtagggaacaggca ccaatatccataaaacacaccggaataaaggcagtagttgctgaaagttttgcacggatattttatagaa attgtattaatataggattaatacctataacttgcgaaggaataaatgaacaaatccaaaacctaaaaga tggcgacacaatagaaattgatttgcaaaatgaaacaataaaaataaattctatgatgttaaattgtgga gctcccaaagggatagaaaaagaaattttagatgctggtggattagtacaatatacaaaaaataagttaa [SEQ ID 135] Ehrlich Yue burning meningitidis outer tei / ia / iococcus aeoZ /cus) Nankai 3, aksE, Maeo_0652

MIIKGNIHLFGDDIDTDAIIPGAYLKTTDPKELASHCMAGIDEKFSTKVKDGDIIVAGENFGCGSSREQAP ISIKHTGIKAWAESFARIFYRNCINIGLIPITCEGINEQIQNLKDGDTIEIDLQNETIKINSMMLNCGAP KGIEKEILDAGGLVQYTKNKLKK

[Serial No. 136] Codon pair optimization, Maeo_0652

ATGATCATCAAAGGTAACATCCACCTGTTCGGTGACGACATCGACACTGACGCTATCATCCCAGGTGCTTA

CCTGAAAACCACTGACCCGAAAGAGCTGGCATCTCACTGCATGGCGGGTATCGACGAAAAATTCTCTACCA

AAGTTAAAGACGGTGACATCATCGTTGCTGGCGAAAACTTCGGTTGCGGTTCTTCCCGTGAACAGGCACCG

ATCTCCATCAAGCACACCGGTATCAAAGCGGTTGTTGCTGAATCCTTCGCTCGCATTTTCTACCGTAACTG

CATCAACATCGGTCTGATCCCGATCACCTGTGAAGGTATCAACGAACAGATTCAGAACCTGAAAGACGGTG

ACACCATCGAAATCGATCTGCAGAACGAAACCATCAAGATCAACTCCATGATGCTGAACTGCGGTGCACCG

AAAGGTATCGAAAAAGAAATCCTGGATGCTGGCGGTCTGGTACAGTACACCAAGAACAAGCTGAAGAAA

[SEQ ID 137] Ehrlich for methane bacteria (Methanococcus aeoJLicusJ Nankai 3, AksD Maeo-0311, a wild-type atgacattggcagaggaaatattatcaaaaaaagtaggaaaaaaagtaaaagcaggagatgttgtagaaat agatatagatttagcaatgactcatgatggaacaacaccattatctgcaaaggcatttaaacagataaccg ataaggtatgggacaataaaaaaatagtcatagtatttgaccataatgtcccagcaaatacattaaaagcc gcaaatatgcaaaaaattacacgagaatttataaaagaacaaaatataataaatcattatttggatggtga a99cgtatgccatcaagtgctacctgaaaacggacatatacaaccaaacatggttatagctggtggagata gtcacacatgcacctatggggcatttggggcatttgcaacaggatttggggctaccgacatggggaatata tatgcaacaggaaaaacttggttaaaagttccaaaaaccataagaataaatgttaatggagaaaatgataa aattactggaaaagatattattttaaaaatttgtaaagaagttggacgaagtggagctacttacatggcac 160 201127961 ttgaatacggcggggaagcaataaaaaaattaagtatggacgaaagaatggttttaagcaatatggctatc gaaatgggcggaaaagttgggcttatcgaagccgatgaaaccacatataattaccttagaaatgtaggaat tagtgaagaaaaaatattagaattgaaaaaaaatcaaataaccattgatgagaacaatatagataatgata attattataaaattataaatatagatattacagacatggaggagcaagtggcatgccctcaccatcccgat aatgttaaaaatatttcggaagta aaaggagctcccataaatcaggtgttcataggttcatgcacaaatgg taggttgaacgatttaagaatagcctcaaaatatttaaaaggaaaaaaggttcataatgatgttagattaa tagtaatacctgcttcaaaatcaatatttaaacaggcattaaaagaaggattaattgatatttttgtagat gctggagctttaatttgcacccccggatgcggtccttgtttgggggcccaccaaggagttttaggagatgg tgaagtttgtttagccaccacaaatagaaatttcaaaggaagaatgggaaatacgacagcggaaatatatt tatcctcccccgctattgccgcaaaaagtgcaattaaaggatatatcacaaatgaataa [SEQ ID 138] Ehrlich A burning lactis aeo / Zcus) Nankai 3, AksD, Maeo_0311

MTLAEEILSKKVGKKVKAGDWEI ΟΙϋΙ ^ ΑΜΤΗΟσΤΤΡΙ ^ ΑΚΑΓΚΏΙΤϋΚνΝΟΝΚΚΙ VIVFDHNVPANTLKA ANMQKITREFIKEQNIINHYLDGEGVCHQVLPENGHIQPNMVIAGGDSHTCTYGAFGAFATGFGATDMGNI YATGKTWLKVPKTIRINVNGENDKITGKDIILKICKEVGRSGATYMALEYGGEAIKKLSMDERMVLSNMAI EMGGKVGLIEADETTYNYLRNVGISEEKILELKKNQITIDENNIDNDNYYKIINIDITDMEEQVACPHHPD NVKNISEVKGAPINQVFIGSCTNGRLNDLRIASKYLKGKKVHNDVRLIVIPASKSIFKQAIjKEGLIDIFVD AGALICTPGCGPCLGAHQGVLGDGEVCLATTNRNFKGRMGNTTAEIYLSSPAIAAKSAIKGYITNE

[SEQ ID NO: 139] Codon pair optimization gene, Maeo_0311

ATGACTCTGGCTGAAGAAATCCTGTCCAAGAAAGTTGGTAAGAAAGTTAAAGCGGGTGACGTTGTTGAAAT

CGATATCGACCTGGCGATGACTCACGACGGTACTACTCCGCTGTCTGCGAAAGCATTCAAGCAGATCACTG

ACAAAGTATGGGATAACAAGAAAATCGTTATCGTTTTCGACCACAACGTTCCGGCTAACACCCTGAAAGCT

GCTAACATGCAGAAGATCACTCGCGAATTCATCAAAGAGCAGAACATCATCAACCACTACCTGGACGGTGA

AGGTGTTTGCCACCAGGTACTGCCGGAAAACGGTCACATTCAGCCGAACATGGTTATCGCTGGCGGCGATT

CTCACACCTGTACTTACGGCGCATTCGGTGCGTTCGCTACTGGCTTCGGTGCAACTGACATGGGTAACATC

TACGCAACTGGTAAAACCTGGCTGAAAGTTCCGAAAACTATTCGTATCAACGTTAACGGTGAAAACGACAA

GATCACCGGTAAAGACATCATCCTGAAAATCTGCAAAGAAGTTGGTCGTTCTGGTGCAACTTACATGGCGC

TGGAATACGGTGGTGAAGCAATCAAGAAACTGTCTATGGACGAACGTATGGTTCTGTCTAACATGGCTATC

GAAATGGGTGGTAAAGTTGGTCTGATCGAAGCTGACGAAACCACTTACAACTATCTGCGTAACGTTGGTAT

TTCTGAAGAGAAGATCCTGGAACTGAAGAAAAACCAGATCACTATCGACGAAAACAACATCGACAACGACA

ACTACTACAAAATCATCAACATCGACATCACTGACATGGAAGAACAGGTTGCTTGCCCGCACCACCCGGAT

AACGTTAAAAACATCTCTGAAGTTAAAGGCGCACCAATCAACCAGGTATTCATCGGTTCCTGCACCAACGG

TCGCCTGAACGATCTGCGCATTGCTTCTAAATACCTGAAAGGTAAGAAAGTTCACAACGACGTACGTCTGA

TCGTTATCCCGGCTTCCAAGTCTATCTTCAAGCAGGCGCTGAAAGAAGGTCTGATCGACATCTTCGTTGAC

GCTGGCGCGCTGATCTGCACTCCGGGTTGCGGTCCGTGCCTGGGTGCACACCAGGGCGTACTGGGTGACGG

TGAAGTTTGCCTGGCAACTACCAACCGTAACTTCAAAGGTCGTATGGGTAACACCACTGCTGAAATCTACC

TGTCCTCTCCGGCAATCGCTGCTAAATCTGCTATCAAAGGTTACATCACTAACGAG 161 201127961 [SEQ ID NO: 140] Wild type gene, nifv

ATGGCTAGCGTGATCATCGACGACACTACCCTGCGTGACGGTGAACAGAGTGCCGGGGTCGCCTTCAATGC

CGACGAGAAGATCGCTATCGCCCGCGCGCTCGCCGAACTGGGCGTGCCGGAGTTGGAGATCGGCATTCCCA

GCATGGGCGAGGAAGAGCGCGAGGTGATGCACGCCATCGCCGGTCTCGGCCTGTCGTCTCGCCTGCTGGCC

TGGTGCCGGCTATGCGACGTCGATCTCGCGGCGGCGCGCTCCACCGGGGTGACCATGGTCGACCTTTCGCT

GCCGGTCTCCGACCTGATGCTGCACCACAAGCTCAATCGCGATCGCGACTGGGCCTTGCGCGAAGTGGCCA

GGCTGGTCGGCGAAGCGCGCATGGCCGGGCTCGAGGTGTGCCTGGGCTGCGAGGACGCCTCGCGGGCGGAT

CTGGAGTTCGTCGTGCAGGTGGGCGAAGTGGCGCAGGCCGCCGGCGCCCGTCGGCTGCGCTTCGCCGACAC

CGTCGGGGTCATGGAGCCCTTCGGCATGCTCGACCGCTTCCGTTTCCTCAGCCGGCGCCTGGACATGGAGC

TGGAAGTGCACGCCCACGATGATTTCGGGCTGGCCACGGCCAACACCCTGGCCGCGGTGATGGGCGGGGCG

ACTCATATCAACACCACGGTCAACGGGCTCGGCGAGCGTGCCGGCAACGCCGCGCTGGAAGAGTGCGTGCT

GGCGCTCAAGAACCTCCACGGTATCGACACCGGTATCGATACCCGCGGCATCCCGGCCATCTCCGCGCTGG

TCGAGCGGGCCTCGGGGCGCCAGGTGGCCTGGCAGAAGAGCGTGGTCGGCGCCGGGGTGTTCACTCACGAG

GCCGGTATCCACGTCGACGGACTGCTCAAGCATCGGCGCAACTACGAGGGGCTGAATCCCGACGAACTCGG

TCGCAGCCACAGTCTGGTGCTGGGCAAGCATTCCGGGGCGCACATGGTGCGCAACACGTACCGCGATCTGG

GTATCGAGCTGGCGGACTGGCAGAGCCAAGCGCTGCTCGGCCGCATCCGTGCCTTCTCCACCAGGACCAAG

CGCAGCCCGCAGCCTGCCGAGCTGCAGGATTTCTATCGGCAGTTGTGCGAGCAAGGCAATCCCGAACTGGC

CGCAGGAGGAATGGCATGA

[Serial No. 141] Aged solid bacteria (A. vinelandiij nifV*

MASVIIDDTTLRDGEQSAGVAFNADEKIAIARALAELGVPELEIGIPSIViG EEEREVMHAIAGLGLSSRLLAWCRLCDVDLAAARSTGVTMVDLSLPVSDL MLHHKLNRDRDWALREVARLVGEARMAGLEVCLGCEDASRADLEFWQVG EVAQAAGARRLRFADTVGVMEPFGMLDRFRFLSRRLDMELEVHAHDDFGL ΑΤΑΝΤΙιΑΑνί ^ ΑΤΗΙΝΤΤνΝαίιαΕΗΑαΝΑΑΙίΕΕίΓνίιΑΙ ^ ΚΝΒΗΘΙΟΤαΐϋ TRGIPAISALVERASGRQVAWQKSWGAGVFTHEAGIHVDGLLKHRRNYE GLNPDELGRSHSLVLGKHSGAHMVRNTYRDLGIELADWQSQALLGRIRAF STRTKRSPQPAELQDFYRQLCEQGNPELAAGGMA

[SEQ ID NO: 142] Diamine-pimelate decarboxylase gene of wild type leaf beetle (T*_man price na)

ATGGACATCCTGAGGAAAGTGGCAGAGATTCATGGGACACCCACCTACGTATACTTCGAGG

AAACACTGCGAAAAAGGTCACGTCTTGTAAAAGAGGTCTTCGAGGGAGTGAATCTCCTTCCA

ACGTTTGCCGTGAAAGCGAACAACAATCCTGTTCTTTTGAAGATTCTAAGAGAAGAGGGTTT

CGGCATGGACGTGGTGACAAAGGGGGAACTCCTCGCGGCTAMCTGGCGGGAGTTCCTTC

CCATACCGTTGTATGGAACGGCAACGGAAAGAGCAGGGATCAAATGGAACACTTTTTGAGA

GAAGATGTGAGAATCGTCAACGTGGATTCGTTCGAGGAGATGGAGATCTGGAGAGAATTGA

ACCCGGAAGGCGTGGAGTATTTCATCAGGGTGAATCCGGAGGTCGATGCGAAGACACACC

CTCACATCTCCACCGGCTTGAAMAGCACAAGTTCGGAATACCACTGGAAGATCTGGATTCG

TTCATGGAAAGATTCAGATCAATGAACATAAGAGGTCTCCATGTTCACATAGGATCGCAGAT

AACCCGGGTTGAACCCTTTGTGGAAGCCTTCAGTAAAGTTGTTCGGGCTTCTGAAAGGTATG 162 201127961

GATTCGAAGAGATCAACATCGGCGGCGGCTGGGGAATAAACTACAGCGGAGAGGAACTCG

ACCTGTCCAGTTACAGAGAAAAGGTTGTTCCTGATTTGAAGAGATTCAAAAGAGTCATCGTC

GAAATAGGAAGGTACATCGTAGCACCTTCTGGGTATCTGCTCCTCAGAGTGGTGCTCGTCA

AAAGAAGACATAACAAGGCGTTCGTTGTAGTCGATGGTGGGATGAATGTCCTCATAAGACC

GGCACTTTATTCCGCATATCACAGGATCTTTGTGCTCGGAAAACAGGGTAAAGAGATGAGG

G C AG ATGTG GTTG GTCCACTGTGCG AAAG CGGTG ACGTG ATCGCGTACG ACCGGG AACTT

CCAGAGGTCGAACCGGGTGACATCATCGCTGTGGAAAACGCGGGAGCTTACGGTTACACTA

TGTCGAACAACTACAACTCGACCACACGTCCAGCTGAAGTGCTCGTCAGAGAAAACGGAAG

AATTTCTCTGATAAGAAGAAGGGAAACGGAGATGGATATTTTCAAAGACGTGGTGATGTGA

[Serial No. 143] Amino acid sequence of the diaminopimelate decarboxylase Q9X1K5 of the leaf aphid (Γ·mar/i/ma)

MDILRKVAEIHGTPTYVYFEETLRKRSRLVKEVFEGVNLLPTFAVKANNNPVLLKILREEGFGMDV

VTKGELLAAKLAGVPSHTVVWNGNGKSRDQMEHFLREDVRIVNVDSFEEMEIWRELNPEGVEY

FIRVNPEVDAKTHPHISTGLKKHKFGIPLEDLDSFMERFRSMNIRGLHVHIGSQITRVEPFVEAFS

KVVRASERYGFEEINIGGGWGINYSGEELDLSSYREKVVPDLKRFKRVIVEIGRYIVAPSGYLLLR

VVLVKRRHNKAFVVVDGGMNVLIRPALYSAYHRIFVLGKQGKEMRADVVGPLCESGDVIAYDRE

LPEVEPGDIIAVENAGAYGYTMSNNYNSTTRPAEVLVRENGRISLIRRRETEMDIFKDVVM

[SEQ ID NO: 144] Codon-optimized leaf beetle (7: manWma) diaminopimelate decarboxylase gene

ATGGACATCCTGAGAAAGGTCGCGGAGATTCACGGTACTCCGACGTACGTCTACTTCGAAG

AGACTTTGCGTAAACGCAGCCGCTTGGTGAAAGAGGTCTTTGAGGGCGTTAATCTGCTGCC

GACGTTCGCGGTGAAGGCGAATAACAATCCGGTCCTGCTGAAGATCCTGCGCGAGGAGGG

TTTTGGTATGGACGTGGTCACCAAGGGCGAGCTGCTGGCGGCAAAACTGGCGGGTGTCCC

GAGCCATACCGTCGTTTGGAATGGTAATGGCAAATCGCGTGACCAGATGGAGCATTTTCTG

CGTGAGGACGTTCGTATCGTTAATGTGGACTCTTTTGAAGAGATGGAAATCTGGCGTGAACT

GAATCCGGAGGGTGTCGAGTATTTCATCCGTGTCAAGCCAGAAGTGGACGCTAAAACGCAT

CCGCACATCAGCACGGGCCTGAAGAAACACAAGTTCGGTATCCCGCTGGAAGATCTGGACA

GCTTCATGGAACGTTTCCGTAGCATGAACATTCGCGGCCTGCACGTTCACATCGGTTCCCA

GATTACCCGCGTCGAACCGTTCGTTGAGGCTTTTAGCAAGGTGGTTCGTGCGAGCGAGCGT

TATGGTTTCGAAGAAATCAACATCGGTGGTGGTTGGGGCATTAACTACTCCGGTGAAGAGCT

GGATCTGAGCTCTTATCGTGAAAAGGTGGTCCCGGACCTGAAACGCTTCAAGCGTGTGATT

GTTGAGATTGGCCGCTACATCGTGGCGCCGTCTGGTTACTTGCTGCTGCGTGTTGTGCTGG

TGAAACGTCGCCATAACAAAGCCTTTGTTGTGGTGGATGGTGGCATGAATGTGTTGATTCGT

CCGGCACTGTACAGCGCCTACCACCGCATTTTCGTCTTGGGCAAGCAAGGCAAAGAGATGC

GTGCGGACGTCGTCGGCCCTCTGTGCGAGAGCGGTGATGTTATTGCTTACGATCGTGAGCT

GCCTGAAGTTGAACCGGGTGACATCATTGCCGTTGAGAACGCAGGCGCGTACGGTTATACC

ATGAGCAATAACTATAACAGCACCACCCGTCCAGCCGAGGTGCTGGTTCGCGAGAACGGTC

GTATTAGCCTGATTCGTCGCCGTGAAACGGAAATGGATATCTTTAAGGATGTGGTTATGTAA

[SEQ ID NO: 145] Diamine-pimelate decarboxylase gene of wild-type glutamate glutamate (C. gf/utem/cum)

ATGGCTACAGTTGAAAATTTCAATGAACTTCCCGCACACGTATGGCCACGCAATGCCGTGCG

CCAAGAAGACGGCGTTGTCACCGTCGCTGGTGTGCCTCTGCCTGACCTCGCTGAAGAATAC

GGAACCCCACTGTTCGTAGTCGACGAGGACGATTTCCGTTCCCGCTGTCGCGACATGGCTA

CCGCATTCGGTGGACCAGGCAATGTGCACTACGCATCTAAAGCGTTCCTGACCAAGACCAT

TGCACGTTGGGTTGATGAAGAGGGGCTGGCACTGGACATTGCATCCATCAACGAACTGGGC

ATTGCCCTGGCCGCTGGTTTCCCCGCCAGCCGTATCACCGCGCACGGCAACAACAAAGGC

GTAGAGTTCCTGCGCGCGTTGGTTCAAAACGGTGTGGGACACGTGGTGCTGGACTCCGCA

CAGGAACTAGAACTGHGGAHACGHGCCGCTGGTGAAGGCAAGATTCAGGACGTGUGA

TCCGCGTAAAGCCAGGCATCGAAGCACACACCCACGAGTTCATCGCCACTAGCCACGAAGA

CCAGAAGTTCGGATTCTCCCTGGCATCCGGTTCCGCATTCGAAGCAGCAAAAGCCGCCAAC

AACGCAGAAAACCTGAACCTGGTTGGCCTGCACTGCCACGTTGGTTCCCAGGTGTTCGACG

CCGAAGGCTTCAAGCTGGCAGCAGAACGCGTGTTGGGCCTGTACTCACAGATCCACAGCG

AACTGGGCGTTGCCCTTCCTGAACTGGATCTCGGTGGCGGATACGGCATTGCCTATACCGC

AGCTGAAGAACCACTCAACGTCGCAGAAGTTGCCTCCGACCTGCTCACCGCAGTCGGAAAA

ATGGCAGCGGAACTAGGCATCGACGCACCAACCGTGCTTGTTGAGCCCGGCCGCGCTATC

GCAGGCCCCTCCACCGTGACCATCTACGAAGTCGGCACCACCAAAGACGTCCACGTAGAC

GACGACAAAACCCGCCGTTACATCGCCGTGGACGGAGGCATGTCCGACAACATCCGCCCA

GCACTCTACGGCTCCGAATACGACGCCCGCGTAGTATCCCGCTTCGCCGAAGGAGACCCA 163 201127961

GTAAGCACCCGCATCGTGGGCTCCCACTGCGAATCCGGCGATATCCTGATCAACGATGAAA

TCTACCCATCTGACATCACCAGCGGCGACTTCCTTGCACTCGCAGCCACCGGCGCATACTG

CTACGCCATGAGCTCCCGCTACAACGCCTTCACACGGCCCGCCGTCGTGTCCGTCCGCGC

TGGCAGCTCCCGCCTCATGCTGCGCCGCGAAACGCTCGACGACATCCTCTCTAGAGAGGC

ATAA

[Serial No. 146] Amino acid sequence of diaminopimelate decarboxylase P09890 of C. glutamicum

MATVENFNELPAHVWPRNAVRQEDGVVTVAGVPLPDLAEEYGTPLFVVDEDDFRSRCRDMAT

AFGGPGNVHYASKAFLTKTIARWVDEEGLALDIASINELGIALAAGFPASRITAHGNNKGVEFLRA

LVQNGVGHVVLDSAQELELLDYVAAGEGKIQDVLIRVKPGIEAHTHEFIATSHEDQKFGFSLASG

SAFEAAKAANNAENLNLVGLHCHVGSQVFDAEGFKLAAERVLGLYSQIHSELGVALPELDLGGG

YGIAYTAAEEPLNVAEVASDLLTAVGKMAAELGIDAPTVLVEPGRAIAGPSTVTIYEVGTTKDVHV

DDDKTRRYIAVDGGMSDNIRPALYGSEYDARVVSRFAEGDPVSTRIVGSHCESGDILINDEIYPS

DITSGDFLALAATGAYCYAMSSRYNAFTRPAVVSVRAGSSRLMLRRETLDDILSLEA

[SEQ ID NO: 147] Codon-optimized glutamic acid decarboxylase gene of Corynebacterium glutamicum (a g/utem/cum)

ATGGCCACGGTCGAAAATTTTAATGAGCTGCCGGCGCACGTCTGGCCTCGTAACGCGGTCC

GCCAAGAGGACGGTGTGGTTACCGTCGCCGGTGTTCCGCTGCCGGACCTGGCAGAAGAAT

ATGGTACTCCGCTGTTCGTGGTTGACGAGGATGACTTTCGTAGCCGTTGTCGTGATATGGC

GACCGCTTTTGGTGGCCCTGGTAACGTGCATTATGCCTCCAAGGCGTTTCTGACCAAAACG

ATTGCACGTTGGGTCGATGAAGAGGGCCTGGCACTGGACATTGCATCGATTAACGAACTGG

GCATTGCTCTGGCAGCGGGTTTTCCGGCGAGCCGTATTACCGCCCATGGCAACAACAAAGG

CGTGGAATTCTTGCGTGCGCTGGTGCAGAATGGTGTTGGCCACGTTGTTCTGGACAGCGCG

CAGGAGCTGGAGCTGCTGGACTACGTCGCGGCAGGCGAAGGTAAGATCCAAGACGTGCTG

ATCCGCGTCAAACCGGGTATCGAAGCGCATACGCACGAATTCATCGCGACCAGCCACGAG

GATCAGAAATTCGGTTTCAGCCTGGCCTCGGGTAGCGCATTTGAGGCGGCGAAAGCGGCA

AACAATGCGGAGAATCTGAATTTGGTTGGTCTGCATTGCCATGTCGGTAGCCAGGTTTTCGA

CGCCGAGGGCTTCAAACTGGCGGCAGAGCGTGTTCTGGGTTTGTACAGCCAAATTCACTCT

GAGCTGGGCGTGGCGTTGCCGGAACTGGATCTGGGTGGTGGCTATGGCATCGCATATACT

GCGGCCGAAGAGCCGCTGAATGTTGCCGAGGTCGCAAGCGACCTGCTGACGGCAGTGGG

CAAGATGGCGGCTGAACTGGGTATTGATGCGCCGACCGTCCTGGTGGAACCGGGTCGTGC

CATCGCCGGTCCATCCACGGTTACCATCTATGAGGTGGGTACCACCAAGGATGTTCACGTC

GACGATGATAAGACTCGCCGCTACATTGCAGTGGATGGTGGCATGAGCGACAACATCCGTC

CAGCGCTGTATGGTAGCGAGTACGATGCGCGTGTTGTGAGCCGCTTTGCTGAAGGCGACC

CGGTCAGCACGCGTATTGTCGGCAGCCACTGTGAGAGCGGCGACATCCTGATTAACGATGA

GATTTACCCGTCCGACATCACGAGCGGTGATTTTCTGGCTCTGGCCGCCACTGGCGCGTAC

TGCTACGCGATGAGCAGCCGCTACAATGCGTTCACGCGTCCTGCTGTTGTGTCTGTCCGTG

CGGGTAGCAGCCGCCTGATGCTGCGCCGTGAAACCTTGGATGACATCTTGTCCCTGGAGG

CATAA

[SEQ ID NO: 148] Wild type B. subtilis (β. suijW (8) diaminopimelate decarboxylase gene

ATGACATTGTTCTTACACGGCACAAGCAGACAAAATCAACATGGTCATTTAGAAATCGGAGG

TGTGGATGCTCTCTATTTAGCGGAGAAATATGGTACACCTCTTTACGTATATGATGTGGCTTT

AATACGTGAGCGTGCTAAAAGCTTTAAGCAGGCGTTTATTTCTGCAGGGCTGAAAGCACAG

GTGGCATATGCGAGCAAAGCATTCTCATCAGTCGCAATGATTCAGCTCGCTGAGGAAGAGG

GACTTTCTTTAGATGTCGTATCCGGAGGAGAGCTATATACGGCTGTTGCAGCAGGCTTTCCG

GCAGAACGCATCCACTTTCATGGAAACAATAAGAGCAGGGAAGAACTGCGGATGGCGCTTG

AGCACCGCATCGGCTGCATTGTGGTGGATAATTTCTATGAAATCGCGCTTCTTGAAGACCTA

TGTAAAGAAACGGGTCACTCCATCGATGTTCTTCTTCGGATCACGCCCGGAGTAGAAGCGC

ATACGCATGACTACATTACAACGGGCCAGGAAGATTCAAAGTTTGGTTTCGATCTTCATAAC

GGACAAACTGAACGGGCCATTGAACAAGTATTACAATCGGAACACATTCAGCTGCTGGGTG

TCCATTGCCATATCGGCTCGCAAATCTTTGATACGGCCGGTTTTGTGTTAGCAGCGGAAAAA

ATCTTCAAAAAACTAGACGAATGGAGAGATTCATATTCATTTGTATCCAAGGTGCTGAATCTT

GGAGGAGGTTTCGGCATTCGTTATACGGAAGATGATGAACCGCTTCATGCCACTGAATACG

TTGAAAAAATTATCGAAGCTGTGAAAGAAAATGCTTCCCGTTACGGTTTTGACATTCCGGAAA

TTTGGATCGAACCGGGCCGTTCTCTCGTGGGAGACGCAGGCACAACTCTTTATACGGTTGG

CTCTCAAAAAGAAGTGCCGGGTGTCCGCCAATATGTGGCTGTAGACGGAGGCATGAACGAC

AATATTCGTCCTGCGCTTTACCAAGCTAAATATGAAGCTGCGGCAGCCAACAGGATCGGAG 164 201127961

AAGCGCATGACAAAACGGTATCAATTGCCGGAAAGTGCTGTGAAAGCGGAGATATGCTGAT

TTGGGATATTGACCTGCCGGAAGTAAAAGAAGGCGATCTTCTTGCCGTTTTTTGTACAGGCG

CTTATGGATACAGCATGGCCAACAATTATAACCGTATTCCGAGACCCGCCGTTGTATTTGTC

GAAAACGGTGAGGCTCATTTAGTCGTGAAGCGAGAAACATACGAAGATATTGTAAAACTTGA

TCTGCCATTTAAAACGGGTGTAAAGCAATAA

[SEQ ID NO: 149] Amino acid sequence of diaminopimelate decarboxylase P23630 of Bacillus subtilis (a Si76i///S)

MTLFLHGTSRQNQHGHLEIGGVDALYLAEKYGTPLYVYDVALIRERAKSFKQAFISAGLKAQVAY

ASKAFSSVAMIQLAEEEGLSLDVVSGGELYTAVAAGFPAERIHFHGNNKSREELRMALEHRIGCI

VVDNFYEIALLEDLCKETGHSIDVLLRITPGVEAHTHDYITTGQEDSKFGFDLHNGQTERAIEQVL

QSEHIQLLGVHCHIGSQIFDTAGFVLAAEKIFKKLDEWRDSYSFVSKVLNLGGGFGIRYTEDDEPL

HATEYVEKIIEAVKENASRYGFDIPEIWIEPGRSLVGDAGTTLYTVGSQKEVPGVRQYVAVDGGM

NDNIRPALYQAKYEAAAANRIGEAHDKTVSIAGKCCESGDMLIWDIDLPEVKEGDLLAVFCTGAY

GYSMANNYNRIPRPAVVFVENGEAHLVVKRETYEDIVKLDLPFKTGVKQ

[SEQ ID NO: 150] Codon-optimized Bacillus subtilis (a SUW///S) diaminopimelate decarboxylase gene

ATGACCTTATTCCTGCACGGTACCTCTCGCCAGAACCAACACGGCCACTTGGAAATCGGTG

GTGTTGACGCACTGTATCTGGCGGAGAAGTACGGTACCCCGTTGTATGTCTACGACGTGGC

CCTGATCCGTGAGCGCGCAAAGAGCTTCAAACAGGCTTTCATTAGCGCTGGTCTGAAGGCG

CAAGTTGCGTATGCGAGCAAAGCGTTTAGCAGCGTTGCCATGATCCAACTGGCGGAAGAAG

AGGGTCTGAGCCTGGACGTCGTGTCTGGCGGTGAGCTGTACACCGCGGTTGCTGCGGGCT

TCCCTGCAGAACGCATTCACTTCCATGGCAACAATAAGAGCCGTGAAGAGCTGCGTATGGC

GCTGGAGCATCGTATTGGTTGCATCGTTGTGGATAACTTTTACGAGATTGCACTGCTGGAAG

ATCTGTGCAAAGAAACGGGTCACAGCATCGATGTGCTGCTGCGCATTACTCCGGGCGTCGA

GGCCCACACCCACGACTACATTACGACGGGCCAGGAAGATAGCAAGTTCGGTTTCGACCTG

CATAATGGTCAAACGGAGCGTGCCATCGAACAGGTGCTGCAATCGGAGCATATTCAACTGT

TGGGTGTGCACTGTCACATCGGCAGCCAGATTTTCGACACCGCAGGCTTTGTCCTGGCTGC

AGAGAAGATTTTCAAGAAACTGGATGAATGGCGCGATTCCTACAGCTTTGTGTCCAAGGTGC

TGAATCTGGGCGGTGGTTTTGGCATCCGCTATACCGAAGATGACGAACCGCTGCACGCAAC

GGAGTACGTTGAGAAAATCATTGAGGCGGTGAAAGAGAACGCGAGCCGCTATGGTTTCGAT

ATTCCGGAGATTTGGATCGAGCCAGGCCGCAGCCTGGTGGGTGACGCCGGCACGACGCTG

TATACTGTCGGTTCTCAGAAAGAAGTTCCAGGCGTCCGTCAGTATGTTGCTGTGGACGGTG

GTATGAACGACAATATCCGTCCGGCGCTGTATCAGGCGAAATACGAGGCAGCTGCAGCGAA

CCGTATCGGCGAAGCCCACGACAAAACCGTCAGCATCGCGGGCAAATGCTGTGAAAGCGG

TGATATGCTGATTTGGGACATCGATTTGCCGGAGGTCAAAGAGGGCGACTTGCTGGCTGTT

TTCTGTACCGGTGCGTATGGTTACAGCATGGCCAATAACTACAATCGTATTCCGCGTCCGGC

CGTTGTGTTTGTTGAGAATGGTGAAGCACATTTGGTTGTGAAGCGTGAAACCTACGAGGACA

TCGTCAAACTGGATCTGCCGTTTAAGACCGGTGTCAAGCAATAA

[Serial No. 151] diaminopimelate decarboxylase gene of P. pui/ofa

ATGAACGCTTTCAACTACCGCGACGGCCAGCTGTTCGCGGAAGGGGTGGCCCTGTCGGCC

GTCGCCGAACGTTTCGGCACCCCCACCTACGTGTATTCGCGCGCCCACATCGAGGCCCAG

TACCGCAGCTACACCGACGCCCTGCAAGGCGCCGAGCACCTGGTGTGCTTCGCGGTCAAG

GCCAACTCCAACCTCGGCGTGCTGAACGTGCTGGCACGCCTGGGCGCAGGCTTCGACATT

GTCTCCGGCGGTGAGCTGGAGCGCGTGCTGGCTGCTGGCGGGCGCGCCGACCGCGTGGT

GTTCTCCGGCGTCGGCAAAACCCGCGAAGACATGCGCCGCGCCCTGGAAGTGGGCGTGCA

CTGCTTCAACGTCGAATCCACCGACGAGCTGGAGCGCCTGCAGGTCGTGGCCGCCGAAAT

GGGCAAGGTCGCCCCGGTGTCGCTGCGGGTAAACCCGGATGTAGACGCCGGCACCCACC

CGTACATCTCCACGGGCCTTAAAGAAAACAAGTTCGGTATCGCCATCGCCGACGCCGAGGC

CATCTACGTGCGTGCCGCGCAGCTTCCGAACCTGGAAGTGGTCGGCGTCGACTGCCACAT

CGGCTCACAGOTGACCACCGTGGAGCCGTTCCTCGATGCCCTCGACCGCCTGCTGGACCT

GGTCGATCGCCTCGCCGACTGCGGCATCCACCTGCGCCATCTGGACCTGGGTGGCGGCGT

TGGCGTGCGCTACCGCGACGAGGAGCCACCGCTGGTGGCCGACTACATCAAGGCTATTCG

CGAACGCGTAGGCAAGCGCGACCTGGCCCTGGTGTTCGAGCCGGGCCGCTACATCGTGGC

CAACGCCGGCGTGTTGCTGACCCGCGTGGAATACCTCAAGCACACCGAACACAAAGACTTC

GCCATCATCGATGCGGCAATGAACGACCTGATCCGCCCGGCCCTTTACCAGGCCTGGATG

GGTGTCAGCGCGGTCATCCCACGCGAAGGCGAAGGGCGTGCCTACGACCTGGTCGGCCC

AATCTGCGAGACCGGCGACTTCCTCGGCAAGGACCGCGTGTTGAACCTGGCCGAAGGCGA 165 201127961

CCTGCTGGCCGTGCAGTCCGCGGGCGCCTATGGTTTTGTCATGAGTTCCAACTACAACACC

CGTGGCCGTTGCGCTGAAATCCTGGTCGACGGCGACCAGGCGTTCGAAGTACGCCGCCGC

GAGACCATCGCCGAACTGTACGCTGGCGAAAGCCTGCTGCCGGAGTAA

[SEQ ID NO: 152] Amino acid sequence of diaminopimelate decarboxylase of Pseudomonas putida (P. ρϋί/da)

MNAFNYRDGQLFAEGVALSAVAERFGTPTYVYSRAHIEAQYRSYTDALQGAEHLVCFAVKANS

NLGVLNVLARLGAGFDIVSGGELERVLAAGGRADRVVFSGVGKTREDMRRALEVGVHCFNVES

TDELERLQVVAAEMGKVAPVSLRVNPDVDAGTHPYISTGLKENKFGIAIADAEAIYVRAAQLPNLE

VVGVDCHIGSQLTTVEPFLDALDRLLDLVDRLADCG Old LRHLDLGGGVGVRYRDEEPPLVADYIK

AIRERVGKRDLALVFEPGRYIVANAGVLLTRVEYLKHTEHKDFAIIDAAMNDLIRPALYQAWMGVS

AVIPREGEGRAYDLVGPICETGDFLGKDRVLNLAEGDLLAVQSAGAYGFVMSSNYNTRGRCAEI

LVDGDQAFEVRRRETIAELYAGESLLPE 166

Claims (1)

201127961 VII. Patent application scope: 1. A method for preparing a compound containing the following amine group (1) H2N——CH2——A——R Preparation of 〇II HO-C--C by ot-keto acid having the following formula —A—R II (2); wherein A represents a hydrocarbyl group, the hydrocarbyl group may contain one or more substituents, and/or contain one or more heteroatoms; R represents a monofunctional group; and wherein the preparation comprises At least one biocatalyst catalyzed reaction step is employed. 2. The method of claim 1, wherein the biocatalyst comprises an enzyme having catalytic activity for catalyzing an amine transfer reaction and/or a reductive amination reaction, preferably the enzyme has at least one selected from the group consisting of Catalytic activity of the group consisting of aminotransferase (EC 2.6.1) and amino acid dehydrogenase (EC 1.4.1). 3. The method of claim 2, wherein the aminotransferase or amino acid dehydrogenase is selected from the group consisting of β-aminoisobutyric acid: α-ketoglutarate aminotransferase, β - alanine aminotransferase, aspartate aminotransferase, 4-amino-butyrate aminotransferase (EC 2.6.1.19), L-lysine 6-aminotransferase (EC 2.6. 1.36), 2-aminoadipate aminotransferase (EC 2.6.1.39), 5-aminopentanoic acid aminotransferase (EC 2.6.1.48), 2-amino group S. 1 201127961 Amino acid amide The transferase is taken as 2·6丄67), the lysine: pyruvate 6-aminotransferase (EC2.6.L71), and the group consisting of the amine acid deaminase (eci 4 i i8). 4. The method of claim 2, wherein the towel is selected from the group consisting of enzymes having catalytic activity for catalytic amine transfer reaction and/or reductive amination reaction from the following organisms, The biological system is selected from the group consisting of Vibrio (10) coffee; Pseudomonas (/3 (10) noodles); Bacillus (type mountain); Hawthorn (10) rc_//5); Iron Fern (Suppressed); (Cer plus (10)); mammal; Streptomyces coli (6); thermophilic bacteria (77 lerm (6); yeast (Sacc / i Qing my (8)); short bacillus (valence; coryneform bacteria (C (?r}; « ekCienMm); Proteus (Rhuyi M5); Agrobacterium; Bacillus licheniformis (GeMflc Mountain (4); Acinetobacter (Adneiokcier); Ralstonia (and this icm (4); Salmonella (Salmonella); Rhodobacter a group consisting of anti-Staphylococcus, especially selected from Bacillus subtilis (such as, for example, /4), weihenstephanensis, Rhodobacter sp/iaerm'iiei, Staphylococcus aureus ( Siflp/iy/ococcw·? awrewi), Veterans Pneumoconiosis (Le ί·ο«β/Ζα /7newmc&gt;/? /ii7a), European Phytophthora sphaeroides (price? (7) such as claw (10) like (10) from), gonorrhea, Pseudomonas syringae CRyewi/omoncLy syringae) 'Rhodopseudomonas ρα/ ι^ίη··?), Vibrio fluvialis (V^r^/Zwvi'a/i·?), and organisms of the group consisting of the genus Pseudomonas 201127961. 5. If the scope of the patent is 2nd The method of any one of the four, wherein the aminotransferase comprises, for example, SEQ ID NO: 2, SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO: 12, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19, SEQ ID NO: 21. No. 23, SEQ ID NO: 25, SEQ ID NO: 27, SEQ ID NO: 29, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 69 amino acid sequence, or any homologue of such sequences. A method according to any one of the preceding claims, wherein the biocatalyst comprises an enzyme having a decarboxylation reaction catalyzing the decarboxylation of an α-keto acid or an amino acid, preferably a decarboxylase (EC 4.1.1), especially From glutamate decarboxylase (EC 4.1.L15), diaminopimelate decarboxylase (EC 4.1.1.20), aspartate de-slowing enzyme (ec 4.1.1. 11), branched 〇c-keto acid decarboxylase, α-protein I acid isovalerate decarboxylase, α__glutaric acid decarboxylase, pyruvate decarboxylase (EC 4.1.1.1), and oxalic acid Decarboxylase of the group consisting of decarboxylase (ec 4.1.1.3). 7. The method of claim 6, wherein the enzyme having catalytic activity for catalyzing a decarboxylation reaction is selected from the group consisting of Cucurbita (CwcMrWiaceae); yeast; Candida; Hanillary yeast; Kluyveromyces; Rhizopus (10)); Neurospora; Monocytogenes (10); Escherichia coli (Ac/ieWc/na); Mycobacterium; Clostridium; Lactobacillus; Pseudomonas (Tmm also, and lactic acid ball 3 201127961 bacteria (LaciococCMi) in the organisms of the group of organisms. 8 · If applying for the special wire _6 phantom method, its towel rarely catalyzes the catalytic reaction to the enemy acid An active enzyme comprising an amino acid such as SEQ ID NO: 31, SEQ ID NO: 34, SEQ ID NO: 37, SEQ ID NO: 4, SEQ ID NO: 43, SEQ ID NO: 46, SEQ ID NO: 143, SEQ ID NO: 146, SEQ ID NO: 149 A method of homologue of any of the homologues of the genus of the genus of the ketone acid, wherein the keto acid is a biocatalyst having an α-keto acid decapric acid activity. In the presence of biocatalytic conversion to the aldehyde oxime-CAR of the following formula ° Ο) wherein A and R are as defined in the scope of the patent application, after which the aldehyde is biocatalyzed to a compound of the formula, at least one of the amine groups, and at least one catalyzes the monoacid The base compound is subjected to a biocatalyst in which an amino group transfer and/or a reductive amination reaction is carried out. 10. A method according to any one of the preceding claims, wherein the α_-acid is biocatalyzed to be converted to the following formula: α-amino acid 〇II Η0 C-CH-A-R NH2 (4) wherein The ruler is as defined in the scope of the patent application, in the presence of at least one amine donor' and at least one biocatalyst capable of catalyzing the transamination and/or reductive amination of the alpha-keto acid, after which the ... The amino acid is biocatalyzed to a compound of formula (1) in the presence of a biocatalyst which catalyzes the deacylation of the amine group 201127961 acid. 11. The method of the patent scope is as follows, where R stands for _ official base, far from the group of -CN, -COOH and -xian 2. A sulfonate wherein A represents one or more substituents, 12. A process according to any one of the preceding claims, a hydrocarbyl group of 2 to 10 carbon atoms which may contain and/or contain one or more heteroatoms . The method of any one of the preceding claims, wherein the compound of the formula (1) is a compound other than 6-aminohexanoic acid. U. The method of claim n, wherein A represents a (C group 'X is an integer, selected from the group of 2, 3, 5, 6, 7, and 8: soil • as claimed The method of claim 14, wherein the compound of the formula (1) is selected from the group consisting of diamine, dihexyl H, ω-aminobutyric acid, ω-amino-pentanoic acid, ω-amino-heptanoic acid and a group of ω amino octanoic acid, especially selected from the group consisting of hydrazine &gt; amino heptanoic acid and diamine heptane. 16. A method of applying the method of 专 围 丨 to 12 items, wherein the preparation is The compound of the formula (1) is 6-aminohexanoic acid, and the OC-keto acid from which 6-aminohexanoic acid is prepared is (X-ketopimelic acid. 17. A method for preparing a polymer comprising the foregoing A compound of the formula (1) or a mixture comprising at least one of the compounds prepared by the method of any one of the patent applications, optionally in the presence of one or more other monomers, is subjected to a polymerization reaction. An acid comprising a sequence of any of SEQ ID NO: 112, 115, 118, 121, 124, 127, 130, 133, 136, 139, and a functional analog thereof, which is fine in E. coli host There are similar, identical or better performances of 201127961. 201127961 IV. Designation of representative drawings: (1) The representative representative figure of this case is: ( ) (None) (2) Simple description of the symbol of the representative figure: If there is a chemical formula in this case, please reveal the chemical formula that best shows the characteristics of the invention: