JP2002142793A - Method for producing optically active amines - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for producing an optically active amine of (S) isomer being an industrially useful compound as an intermediate raw material for medicines, agrochemicals, etc. SOLUTION: This method for producing an optically active amine of (S) isomer is characterized in that a ketone compound represented by general formula (1) (R and X are each independently an aryl group, a heteroaryl group or an alkyl group which may be substituted; R is combined with X to form a carbon ring or a hetero ring; R is a group having a higher order of priority by an order rule) as an amino acceptor is treated in the presence of an amino donor with transaminase not to have reduction in activity in the presence of gabaculine and an amino group transfer is stereoselectively carried out to give an optically active amine compound of general formula (2) (R and X are as shown for the general formula (1)).

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a method for producing optically active amines, such as (S) -1-phenylpropylamine, which are industrially useful compounds as intermediate raw materials for medicines, agricultural chemicals and the like.

[0002]

2. Description of the Related Art Amine is usually produced by an organic chemical synthesis method. In recent years, a method of producing a corresponding optically active amine from a ketone by using a microorganism has attracted attention. As a method for producing amines by an enzymatic method, a method based on a dehydrogenase method (Japanese Patent Publication No. 4-11194) and a transaminase method (Patent Registration No. 2846)
No. 074, WO 97/15682, WO 98
/ 48030 and JP-A-1-174398)
The transaminase method includes a method of optically resolving a racemic amine and a method of producing optically active amines from ketones.

[0003]

However, the method for producing optically active amines from ketones by the dehydrogenase method requires the addition of expensive coenzyme NADH or NADPH or a regeneration system, which increases the cost. This is not preferable for industrial production. In addition, when the optical resolution method is used, the target non-steric compound becomes the growth energy of the microorganism, and the yield is up to 50%. This is not an advantageous method.

Further, a method for producing optically active amines from ketones is disclosed in WO 97/15682, WO 9/15682.
No. 8/48030 and Japanese Patent No. 2846074 also disclose, but WO97 / 15682 and WO98 / 48030 disclose an (R) -form selective enzyme and an (R) -form using the same. Only a method for producing an amine is described. On the other hand, Japanese Patent No. 2846074 describes a method for producing an (S) -form optically active amine from a ketone using an ω-amino acid transaminase. However, the substrate concentration is as low as 4.2 mM, and it is necessary to reduce the substrate concentration to half or less even in comparison with the optical resolution method using a racemic amine. Further, according to the description in the examples of the publication, in the case of a compound such as acetophenone in which both sides of the carbonyl group are not carbon chains, the yield is sharply reduced as compared with 1-phenyl-3-aminobutane and the like. There are also problems.

An object of the present invention is to provide a method for industrially efficiently producing optically active amines of the (S) form from ketones using transaminase. The method for synthesizing amines by the enzymatic method is considered to be an extremely useful means for synthesizing amines used as intermediates for pharmaceuticals and agrochemicals, because they have few synthetic by-products, are safe and simple.

[0006]

Means for Solving the Problems In view of the above-mentioned circumstances, the present inventors have conducted intensive studies on a method of producing optically active amines in the (S) form by reacting a transaminase with a ketone, and as a result, the activity was reduced in the presence of gabacrine. It has been found that the use of a transaminase having (S) isomer selectivity that is not found achieves the object of the present invention, and has completed the present invention.

That is, the gist of the present invention is the following general formula (1):

[0008]

Embedded image

(Wherein R and X are each independently
It represents any of an aryl group, a heteroaryl group and an alkyl group which may be substituted. Further, R and X may be combined to form a carbocyclic or heterocyclic ring. However, R is a group having a higher priority order than X according to the order rule. ) Is used as an amino acceptor, and in the presence of an amino donor, a transaminase that does not show a decrease in activity in the presence of gabacrine is acted to perform stereoselective transamination. Equation (2)

[0010]

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(Wherein R and X are each represented by the general formula (1)
Is synonymous with )), Wherein the optically active amine compound represented by the formula (S) is obtained. Hereinafter, the configuration of the present invention will be described in detail.

[0012]

BEST MODE FOR CARRYING OUT THE INVENTION The production method of the present invention comprises the following general formula (1)

[0013]

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(Wherein R and X are each independently
It represents any of an aryl group, a heteroaryl group and an alkyl group which may be substituted. However, R is a group having a higher priority order than X according to the order rule. ) Is converted to amines of the (S) form. The aryl group which may be substituted, which is a substituent in the above formula, includes a halogen atom such as a chlorine atom, a hydroxyl group, an alkyl group having 1 to 6 carbon atoms, an alkoxy group, and a haloalkyl group. Examples include a phenyl group or a naphthyl group which may be substituted with a substituent. Of these, an unsubstituted aryl group or an alkylaryl group is preferable, and a phenyl group is particularly preferable.

Examples of the optionally substituted heteroaryl group include a halogen atom such as a chlorine atom, a hydroxyl group, and a group having 1 carbon atom.
And a pyridyl group, a pyrazyl group, a thienyl group, a furyl group, and an azolyl group which may be substituted with a substituent inert to a reaction such as an alkyl group, an alkoxy group, a haloalkyl group, or an aryl group. . Examples of the alkyl group which may be substituted include a halogen atom such as a chlorine atom,
Hydroxyl group, alkoxy group, haloalkyl group, aryl group,
Or an alkyl group which may be substituted with a substituent inert to a reaction such as a heteroaryl group, and the like.Specific examples include a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, Examples include a t-butyl group and a benzyl group. Of these, an unsubstituted alkyl group or an arylalkyl group is preferable, and particularly preferable is
It is an alkyl group having 1 to 4 carbon atoms.

Further, R and X may form a carbocyclic or heterocyclic ring together. Further, the carbon ring or the hetero ring may have a substituent which is inert to the reaction such as a halogen atom such as a chlorine atom, a hydroxyl group, a (halo) alkyl group, an alkoxy group, an aryl group, or a heteroaryl group as a side chain. . Preferably, R is an optionally substituted aryl group, and X is an optionally substituted alkyl group, or

[0017]

Embedded image

(Wherein, R ^{1 to} R ⁴ are each independently a group selected from the group consisting of a hydrogen atom, a halogen atom, a hydroxyl group, a (halo) alkyl group, an alkoxy group, an aryl group and a heteroaryl group; m and n are each a natural number, provided that m> n), and specific examples of the compound represented by the above structural formula include 1
-Indanone, 2-indanone, 1-tetralone, 2-
Examples include tetralone, 5-methoxy-2-tetralone, 6-methoxy-2-tetralone, 7-methoxy-2-tetralone, 8-methoxy-2-tetralone, and the like.

Preferred specific examples of the ketone having a substituent include acetophenone, propiophenone, benzylacetone, 4-methoxyphenylacetone, 1-tetralone, 2-tetralone, and 7-methoxy-2-tetralone. . In the production method of the present invention, as the amino donor used, normal propylamine,
Primary amines such as isopropylamine, normal butylamine, secondary butylamine, 2,2-dimethylpropylamine, benzylamine, 1-ethylpropylamine, 2-aminomethoxybutane, 1-phenylethylamine, and 2-phenylethylamine Normal butylamine, 2-methylbutylamine, 2,2-
Dimethylpropylamine or benzylamine is preferred.

The transaminase used in the present invention has (S) -selective transaminase activity,
No substantial decrease in activity in the presence of gabacrine is observed,
That is, when the enzyme solution is made to coexist with gabacrine at a concentration of 1 mM or more, preferably 2 mM or more, and when the reaction is performed after preincubation for 10 minutes or more, preferably 30 minutes or more, when the activity is not coexisted with gabacrine, As far as the transaminase has a decrease in activity of 20% or less, preferably 10% or less, it is not particularly limited, and examples thereof include a transaminase derived from Bacillus.

The optically active amines of the present invention can be obtained by converting the above ketones into a culture solution of a microorganism having the above transaminase,
Alternatively, it can be produced by bringing an amino donor into contact with an aqueous solution containing cells of the microorganism and / or a processed product of the cells. Specific examples of the microorganism having the transaminase include Bacillus sp.
899 strains. In addition, Bacillus SP M
The CI3899 strain was added to the FERM P-179 on August 22, 2000 by
Deposited with 96 accession numbers.

The results of the identification of the strain are shown below. Results of identification of MCI3899 strain 1. Morphological properties (1) Cell shape rod-like (2) Cell polymorphism No (3) Motility Yes (4) Presence or absence of spores No (Normal agar medium, soil extract medium) Cultural properties (1) Gravy agar plate culture After cultivation at 30 ° C for 2 to 3 days, light brown, round, semi-lens-like, opaque colonies are formed, and the colony surface is smooth and glossy. No film formation on liquid surface (3) Broth gelatin stab culture No liquefaction (4) Litmus / milk Decolorization and other changes Physiological properties (1) Gram stain positive (2) Nitrate reduction positive (3) MR test negative (4) VP test negative (5) Hydrogen sulfide generation negative (6) Starch hydrolysis negative (7) Citric acid Use Christensen medium positive (9) Esculin degradation positive (10) Pigment formation Not generated (11) Urease positive (12) Oxidase negative (13) Catalase positive (15) Attitude to oxygen Does not grow under anaerobic conditions (16) OF test No change (17) Generation of acid from saccharide L-arabinose-D-xylose-D-glucose-D-mannose-D-fructose -D-galactose-maltose-sucrose-lactose-trehalose-D-sorbitol-D-mannitol-inositol-glycerin + starch-(+: generates acid,-: does not generate) 5. Chemotaxonomic properties Major isoprenoid quinone Menaquinone MK7 6.16S Phylogenetic position based on rRNA base sequence See Figure 1 7. Taxonomic considerations The strain MCI3899 is composed of 1) gram-positive bacilli,
2) Shows aerobic, 3) Shows motility, 4) Main isoprenoid quinone has characteristics such as menaquinone MK7. Based on these characteristics, this strain is known as Bergey's Manual Systematic Bacteriology [Bergey's M.
anual of Systematic Bacteriology] Volume 2, 110
It was suggested to be included in the genus Bacillus described on pages 4 to 1139 (1986). But,
International Journal of Systemic Bacteriology [International Journal of Sy
Stematic Bacteriology], Vol. 46, pp. 939-946 (1996), phylogenetic taxonomic analysis based on the base sequence of 16S rRNA has newly divided six genera from the genus Bacillus. Therefore, to clarify the phylogenetic positional relationship of this strain, 16S r
The RNA base sequence was determined.

From the phylogenetic tree based on the 16S rRNA nucleotide sequence (FIG. 1), it was found that this strain was included in the genus Bacillus. In addition, "Bacillus macroides"("Bacillu
smacroides ") and more than 99% homology, suggesting that it is the closest relative.
Macroides "has not been officially recognized as a species name of the genus Bacillus at present, and the strain MCI3899 could not be determined to be the same, for example, sporulation was not confirmed.

Therefore, from the above results, the strain MCI38
99 strains were identified as Bacillus sp. The above microorganisms are not only wild strains, but also
Any strain may be used, such as a mutant strain obtained by ordinary mutation treatment such as G treatment, or a recombinant strain derived by a genetic technique such as cell fusion or gene recombination. still,
The host of the genetically modified strain may be the same genus as the parent strain or a different genus such as Escherichia coli as long as it is a transformable microorganism. In the production method of the present invention, one or more of the above microorganisms are used in the form of cells and / or their processed products. Specifically, cells in a wet state obtained by culturing the microorganism and culturing the solution as it is or by centrifugation; cells obtained by treating the cells with acetone, those obtained by freeze-drying, the cells A treated bacterial cell, such as one obtained by physically or enzymatically crushing, can be used. It is also possible to extract and use the enzyme fraction of the (S) -selective transaminase as a crude product or a purified product from the cells or the treated cells. Furthermore, the cells obtained in this manner, the treated cells, the enzyme fraction, and the like are subjected to ordinary immobilization techniques, that is, polyacrylamide gel,
It is also possible to use those immobilized on a carrier represented by carrageenan gel or the like. Therefore, in this specification, the term "microbial cells and / or processed products thereof" is used as a concept including all of the above-mentioned microbial cells, processed microbial cells, enzyme fractions and their immobilized products.

The above-mentioned microorganism is usually used after being cultured, and this culture can be carried out according to a conventional method. The medium used for the culture contains a carbon source, a nitrogen source, inorganic ions and the like that can be utilized by the microorganism. Examples of the carbon source include carbohydrates such as glucose, fructose, sucrose, and saccharose; glycerin, mannitol,
Polyalcohols such as xylitol and ribitol; organic acids such as citric acid and fumaric acid and the like are appropriately used. Nitrogen sources include inorganic nitrogen sources such as ammonium sulfate and sodium nitrate; NZ amine, tryptoose, yeast extract,
Organic nitrogen sources such as polypeptone, urea, meat extract, corn steep liquor, soybean extract and the like can be used as appropriate. As inorganic ions, magnesium, potassium,
What contains metal ions, such as iron, manganese, and molybdenum, phosphate ions, etc. should just be used, but it is convenient to add inorganic salts which combined them.

In the medium, ethylamine, normal propylamine, isopropylamine, normal butylamine, secondary butylamine, 1,2-dimethylbutylamine, 1,2
1,3,3-tetramethylbutylamine, benzylamine, 1-
Ethylpropylamine, 2-aminomethoxybutane, 1-
It is preferable to add one or more amine compounds selected from primary amines such as phenylethylamine to induce enzyme activity. The inducer need not be the same compound as the amino donor used in the reaction, and is preferably normal propylamine, isopropylamine,
1-phenylethylamine. As the additive concentration of the inducer, 0.01 to 3 g / L can be used, but 0.1 to 1 g / L.
Is preferred.

The reaction system of the present invention includes a method in which ketones and an amino donor are directly added to the above-mentioned culture medium (medium) in which microorganisms are cultured, and the reaction is carried out. A method in which the body is added to an aqueous medium containing a ketone and an amino donor as it is or as a processed product of the cells is given. In addition, any reaction mode such as a standing reaction, a shaking reaction or a stirring reaction may be used, but when the substrate is insoluble in an aqueous solution, a shaking or stirring reaction is preferable.

As the reaction temperature, a temperature at which the transaminase exhibits activity, for example, 0 ° C. to 80 ° C. can be used, and preferably 20 to 50 ° C. The pH may be in any range as long as the transaminase reacts.
The reaction is carried out on the neutral to slightly basic side, that is, in the range of 7 to 10, preferably 7.5 to 8.

As the aqueous medium, a buffer having an appropriate buffer capacity can be used. As a buffer to be used, tris (hydroxymethyl) aminomethane (Tris), N-
Tris (hydroxymethyl) -methyl-2-aminoethanesulfonic acid (TES), and 2- (N-morpholino)
PKa in neutral pH range such as ethanesulfonic acid (MES)
A buffer using an amine compound having the following, an organic acid buffer such as oxalic acid and maleic acid, and a buffer such as EDTA, phosphoric acid and carbonic acid are not limited thereto. Preferably, the buffer to be added is phosphate and Tri.
s buffer. The buffer concentration is, for example, 1 mM to 1 M, preferably 0.01 to 0.5 M.

The reaction substrate concentration is, for example, 0.1 m
M-2M can be used, but preferably 10 mM-100
mM. Further, the concentrations of the respective reaction substrates of the ketones and the amino donor need not be equal. For example, when one substrate is inhibited by an enzyme, the substrate may be added sequentially in the presence of a large excess of the other substrate, or the reaction equilibrium may be shifted to the product side. Alternatively, one substrate may be used in a large excess with respect to the other substrate. The reaction substrate may or may not be dissolved in the reaction system.

An organic solvent, a surfactant and / or a fatty acid may be added to the reaction system as a reaction promoting substance. Examples of the organic solvent to be added include aliphatic hydrocarbons such as heptane and isooctane, aromatic hydrocarbons such as toluene, alcohols, esters such as ethyl acetate, and DMS.
Examples include aprotic polar solvents such as O and DMF, nitriles such as acetonitrile, glycerol, and the like, preferably aliphatic or aromatic hydrocarbon solvents, and particularly preferably isooctane. As the amount of the organic solvent to be added, 0.1 to 60% v / v can be used, but preferably 1 to 20% v / v.

As the surfactant, Tween 80, Tri
nonionic surfactants such as tonX-100, SDS
Examples of the surfactant include those commonly used in enzyme reactions and bacterial cell reactions, such as anionic surfactants such as
Ween80. As the amount of surfactant to be added,
0.01-5% w / v can be used, but preferably 0.1% w / v.
05-0.5% w / v.

The fatty acid may be a saturated fatty acid or an unsaturated fatty acid, and examples thereof include linoleic acid, linolenic acid, oleic acid, lauric acid, palmitic acid, caproic acid and esters thereof, preferably oleic acid and palmitic acid. Is an acid. As the amount of fatty acid to be added, 0.01 to 5% v / v can be used, but preferably 0.1 to 1% w / v.

Among the above-mentioned organic solvents, surfactants and fatty acids, preferred are organic solvents or surfactants, and particularly preferred are organic solvents. The above-mentioned organic solvent, surfactant and / or fatty acid may be added at any time, such as at the start of the reaction or during the reaction, but is preferably at the start of the reaction.
As a method for separating the objective optically active amine after the reaction, extraction using an organic solvent, distillation, and column chromatography are combined, and depending on the type of the compound, the separation is performed by a usual method combining crystallization and the like. be able to. For example, specifically, using ethyl acetate, toluene, or the like as an extraction solvent, the reaction solution is first acidified to remove ketones by extraction, and then alkalinized to extract amines containing the target compound. To separate. The objective optically active amines can be isolated by further distilling the extracted fraction.

[0035]

EXAMPLES The present invention will be described below in detail with reference to examples, but the present invention is not limited to the following examples. (Example 1) Glycerol 2 g / L, polypeptone
4g / L, yeast extract 4g / L, monopotassium phosphate
1 g / L, dipotassium phosphate 3 g / L, sodium chloride 1 g / L, magnesium sulfate 0.2 g / L, manganese sulfate n-hydrate 10 mg / L, iron (II) chloride heptahydrate 10 mg / L, chloride Calcium 10mg / LpH
Dispense 100 mL of medium containing 7.0 and sterilize (121
B. sp. MCI3899 (FERM) in a 500 mL baffled Erlenmeyer flask
P-17996) and inoculated at 30 ° C. with shaking.
Six hours after sowing, 500 μL of a solution of 100 mg of 1-phenylethylamine dissolved in 400 μL of ethanol was added, followed by shaking culture at 30 ° C. for 14 hours, followed by centrifugation (800 hours).
(0 g for 10 minutes). Physiological saline was added to the obtained cells to make the total volume 100 mL, suspended, and then centrifuged again (8000 g for 10 minutes) to wash.

A physiological saline solution was added to the above cells again, and 100 m
The suspension was centrifuged to collect cells, and 0.3 mL of a reaction solution (propiophenone 10 mM, n-butylamine 50 mM, pyrodoxal phosphate 0.1 mM, Tween 80
(Kanto Chemical Co., Ltd.) 1 g / L, 100 mM phosphate buffer pH 7.7)
, And subjected to a shaking reaction at 30 ° C for 24 hours. After completion of the reaction, the reaction solution was subjected to centrifugation (8000 g, 5 minutes) to remove the cells, and then subjected to HPLC. The amount of ethylbenzylamine produced was measured. As a result, it was 9.5 mM and the optical purity was 100% ee. <To measure the amount of ethylbenzylamine produced> Column: ULTRON ESPhCD Mobile phase 1: 1 g / L KH ₂ PO ₄ , 3 g / LK ₂ HPO ₄ (pH7.3), 50% acetonitrile Flow rate: 1 mL / min Detection: UV (210 nm) ) <For optical purity measurement> Column: Daicel CROWNPAK CR (+) Mobile phase 2: aqueous perchloric acid (pH 1.5) Flow rate: 0.8 mL / min Detection: UV 210 nm (Reference Example 1) Using aminobutylamine from normal butylamine The reaction was carried out in the same manner as in Example 1 except that each of the amine compounds shown in Table 1 was used.

The results are shown in Table 1. The activity is shown as a relative activity when normal butylamine is used as the amino donor and the activity is 100%.

[0038]

[Table 1]

As can be seen from the above table, when primary amines are used as amino donors, they show activity, but L-alanine,
And ω- such as γ-aminobutyric acid, L-lysine and ornithine
It turns out that amino acids hardly act as amino donors. (Reference Example 2) Bacillus SP M as in Example 1.
500 mL of CI3899 (FERM P-17996)
Was inoculated into 10 Erlenmeyer flasks with baffles (1000 mL of culture solution), cultured for 19 hours, and washed with saline to collect cells. The cells were suspended in 60 mL of a disruption buffer (10 mM phosphoric acid pH 7.7, 20 uM pyridoxal phosphate, 1 mM DTT) and then disrupted by sonication. After crushing, centrifuge (10,000 g
(30 minutes) to remove the crushed bacterial cells, ammonium sulfate was added to the supernatant to 25% saturation to remove precipitated proteins, and ammonium sulfate was added to 50% saturation to precipitate. The protein was separated. This was dissolved in the above buffer solution, dialyzed against the same buffer solution, and then applied to a DEAE-Sepharose Fast Flow (Pharmacia) column (φ1.6 × 10 cm) equilibrated with this buffer solution. Was eluted. The activated fractions were collected, ammonium sulfate was added to a final concentration of 1M, and the column was equilibrated with the above-mentioned crushing buffer containing 1M ammonium sulfate.
9 × 15 cm) and eluted with a linear concentration gradient of 1M to 0M ammonium sulfate. This active fraction was collected, concentrated and desalted by ultrafiltration, and then MCI-GEL Prot-
Apply to DEAE (7.5mm IDx100mm) column, 0.15M ~ 0.4M Na
Eluted with a linear Cl gradient. The active fractions were collected and concentrated by ultrafiltration to obtain a crude enzyme solution.

Using the crude enzyme solution, the effect of various enzyme inhibitors on transaminase was examined.
Add enzyme inhibitor solution to 100 ul of crude enzyme solution (final concentration
After preincubation at room temperature for 30 minutes, 100 uL of the reaction solution (0.1 M phosphate buffer (pH 7.7), 50 mM sec-butylamine, 0.1 mM pyridoxal phosphate, 15 mM propiophenone, 0.1% Tween80 (Kanto Chemical) 1) at 30 ° C
The reaction was performed in a tube for 4 hours. After completion of the reaction, the reaction solution was diluted 5-fold, and the solution was analyzed by HPLC to determine (S) -ethylbenzylamine produced by the reaction. The results at that time are shown as relative activities to those when no inhibitor was added.

[0041]

[Table 2]

With respect to the ω-amino acid transaminase, Pseudomonas s is well known as the enzyme.
p. Literature on ω-amino acid transaminase from F-1 (Agric. Biol. Chem.), 42, 2363-2367
Page (1978), Agricultural Biological Chemistry (Agric. Biol. Chem.), 43, 1043-1048
(1979), Journal of Biological Chemistry (J. Biol. Chem), 258, 2260-2265 (1983)
Year))). From the above table, it can be seen that the transaminase used in the present invention does not show any decrease in activity against gabacriin, which is known as a selective inhibitor of ω-amino acid transaminase, and is not an ω-amino acid transaminase.

In addition, 15 mM propiophenone and 2
The reaction was carried out using a combination of the following ω-amino acid and amino receptor instead of 50 mM butylamine.
In addition, since the solubility of each substrate in water is high, it is 0.
There was no need to add 1% Tween80 and it was not added. The results are shown in the table.

[0044]

[Table 3]

From these results, it can be seen that the transaminase used in the present invention has no or almost no activity on ω-amino acids, and has different substrate specificity and properties from ω-amino acid transaminases. I understand.

[0046]

According to the present invention, it is possible to efficiently produce (S) optically active amines which are industrially useful as intermediates for medical and agricultural chemicals at low cost and high purity.

[Brief description of the drawings]

FIG. 1 shows the phylogenetic position of the Bacillus sp. MCI3899 strain based on the 16S rRNA nucleotide sequence.

 ──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Makoto Goto 1000 Kamoshita-cho, Aoba-ku, Yokohama-shi, Kanagawa Prefecture Inside Mitsubishi Chemical Research Institute (72) Inventor Reiko Shida 1000-Kamoshita-cho, Aoba-ku, Yokohama-shi, Kanagawa Mitsubishi Chemical Research Co., Ltd. Yokohama Research Laboratory F-term (reference) 4B064 AE01 CA02 CA21 CB30 CD05 CD12 DA01

Claims (12)

[Claims] [Claim 1] The following general formula (1) (Wherein, R and X each independently represent an optionally substituted aryl group, heteroaryl group or alkyl group. Further, R and X are taken together to form a carbocyclic or heterocyclic group. Wherein R is a group having a higher order of priority according to the ordering rule than X.) The amino acid acceptor is used as the amino acceptor, and in the presence of an amino donor and coexistence of gabacrine. By reacting a transaminase that does not show a decrease in the activity of the compound with the effect of stereoselective transamination, the following general formula (2) can be obtained. (Wherein R and X have the same meanings as in the general formula (1), respectively). A method for producing an optically active amine of the (S) form, characterized by obtaining an optically active amine compound represented by the following formula (1). 2. The method according to claim 1, wherein R is an optionally substituted aryl group and X is an optionally substituted alkyl group. 3. A compound represented by the general formula (1): (Wherein, R ^{1 to} R ⁴ are each independently a group selected from the group consisting of a hydrogen atom, a halogen atom, a hydroxyl group, a (halo) alkyl group, an alkoxy group, an aryl group, and a heteroaryl group; And n are natural numbers, where m> n.)
The manufacturing method as described. 4. The method according to claim 1, wherein the amino donor is a primary amine. 5. The method according to claim 1, wherein the transaminase has no ω-amino acid transaminase activity. 6. A transaminase comprising Bacillus (Baci).
The production method according to any one of claims 1 to 5, wherein the method is a transaminase derived from the genus llus). 7. The method according to claim 1, wherein the transaminase activity is induced by adding an amine during the culturing. 8. The concentration of the ketone compound is from 0.1 mM to 1 mM.
The production method according to any one of claims 1 to 7, wherein the concentration is 00 mM. 9. The method according to claim 1, wherein the pH of the reaction system is 7-9. 10. The method according to claim 1, wherein the temperature in the reaction system is 0 ° C. to 80 ° C. 11. The method according to claim 1, wherein the reaction is carried out in the presence of a surfactant and / or an organic solvent. 12. A culture of a microorganism producing a transaminase which has an activity of performing stereoselective transamination and does not show a decrease in the activity in the presence of gabacrine, isolated cells and / or treated cells. The method according to any one of claims 1 to 11, wherein a ketone is contacted with a ketone compound and an amine.