Gene suitable for alcohol dehydrogenase, vector and transformant

1. A purified DNA which is any one of the following DNAs (A), (B), (C), and (D):

(A) a DNA comprising the nucleotide sequence shown under SEQ ID NO:2 in the sequence listing;

(B) a DNA hybridizable under a stringent condition with a DNA that comprises the nucleotide sequence complementary to the nucleotide sequence shown under SEQ ID NO:2 in the sequence listing, and coding for a polypeptide that has activity to react with acetophenone to reduce into (S)-1-phenylethanol, wherein said DNA hybridizable under a stringent condition is obtainable by hybridization with a filter in the presence of 0.7 to 1.0 M NaCl, at 65° C., followed by washing the filter with a 0.2-fold concentration of SSC solution at 65° C.;

(C) a DNA having sequence identity of 90% or higher to the nucleotide sequence shown under SEQ ID NO:2 in the sequence listing, and coding for a polypeptide that has activity to react with acetophenone to reduce into (S)-1-phenylethanol; and

(D) a DNA comprising a nucleotide sequence that results from deletion, insertion, substitution and/or addition of 100 or less nucleotides in the nucleotide sequence shown under SEQ ID NO:2 in the sequence listing, and coding for a polypeptide that has activity to react with acetophenone to reduce into (S)-1-phenylethanol.

2. A purified DNA coding for a polypeptide which is derived from Candida maltosa and has the following physicochemical properties (1) to (6):

(1) Action:

The polypeptide oxidizes an alcohol with NAD + as a coenzyme to produce a ketone or an aldehyde, or reduces a ketone or an aldehyde with NADH as coenzyme to produce an alcohol;

(2) Substrate specificity:

A substrate in oxidation reaction is an aliphatic alcohol which may have an aromatic substituent, and the polypeptide preferentially oxidizes (S)-2-butanol in comparison with (R)-2-butanol; and a substrate in reduction reaction is a ketone or an aldehyde, and the polypeptide reacts with acetophenone to reduce into (S)-1-phenylethanol;

(3) Molecular weight:

The molecular weight of the polypeptide is approximately 39,000 as determined by reducing SDS-polyacrylamide gel electrophoresis;

(4) Stability in pH:

Stable pH range of the polypeptide is 5.5 to 7.5;

(5) Optimum temperature:

Optimum temperature of the polypeptide for action is 45° C. to 70° C.;

(6) Inhibitor:

Enzyme activity of the polypeptide is inhibited by ethylenediaminetetraacetic acid, o-phenanthroline, mercury chloride, copper sulfate or zinc sulfate, whereas is not inhibited by 2-mercaptoethanol or dithiothreitol.

3. A vector comprising the DNA according to claim 1 .

4. The vector according to claim 3 ,

wherein the polypeptide with ability to reproduce a reduced coenzyme is glucose dehydrogenase or formate dehydrogenase.

5. A transformant

which is producible by transformation of a host cell with the vector according to claim 3 .

6. The transformant according to claim 5 ,

wherein the host cell is Escherichia coli .

7. A method for producing an alcohol comprising

reacting the transformant according to claim 5 or 6 , and/or a processed product of the transformant with a carbonyl group-containing compound.

8. The method for producing an alcohol according to claim 7 ,

wherein the carbonyl group-containing compound is an asymmetric ketone, and a product therefrom is an optically active alcohol.

9. The method for producing an alcohol according to claim 8 , wherein the asymmetric ketone is a 1-phenylethanone derivative represented by the following formula (1):

wherein R 1 and R 2 are each independently a hydrogen atom, a halogen atom, a hydroxy group, a substituted or unsubstituted alkoxy group, a substituted or unsubstituted alkyl group, an amino group or a nitro group, and R 1 and R 2 may be the same as or different from one another; R 3 is a hydrogen atom, a halogen atom, a hydroxy group or a substituted or unsubstituted alkyl group; and

the product optically active alcohol therefrom is an optically active 1-phenylethanol derivative represented by the following formula (2):

wherein R 1 , R 2 and R 3 are the same as defined above.

10. The method for producing an alcohol according to claim 9 ,

wherein R 3 is a hydrogen atom.

11. The method for producing an alcohol according to claim 8 ,

wherein the asymmetric ketone is a methyl ketone compound represented by the following formula (3):

wherein R 4 is a substituted or unsubstituted alkyl group, a substituted or unsubstituted alkenyl group or a substituted or unsubstituted alkynyl group, and

the product optically active alcohol therefrom is an optically active 1-substituted-1-ethanol derivative represented by the formula (4):

wherein R 4 is the same as defined above.

12. The method for producing an alcohol according to claim 11 ,

wherein R 4 is a C 1-7 alkyl group which may be substituted by a halogen atom, a hydroxy group, an alkoxy group, an alkoxycarbonyl group, an amino group or a nitro group.

13. The method for producing an alcohol according to claim 11 ,

wherein R 4 is a vinyl group.