Novel enone reductases, methods for producing same, and methods for selectively reducing a carbon-carbon double bond of an alpha, beta-unsaturated ketone using the reductases

Abstract

The object of the present invention is to provide novel enone reductases useful for producing ketones. Accordingly, novel enone reductases derived from the genus Kluyveromyces are provided by the present invention. In addition, the present invention also provides genes encoding the enzymes and vectors containing the genes, as well as transformants. Furthermore, the present invention provides enone reductases derived from yeast. Methods for selectively reducing the carbon-carbon double bonds of α,β-unsaturated ketones using these enone reductases are provided. Ketones useful as a raw material for pharmaceuticals can be produced based on an enzymatic reaction.

What is claimed is:  
     
         1 . A method for selectively reducing the carbon-carbon double bonds of an α,β-unsaturated ketone, the method comprising reacting the α,β-unsaturated ketone with an enzymatically active material selected from the group consisting of: 
 (A) a polypeptide having enone-reducing activity, wherein the polypeptide is encoded by a polynucleotide selected from the group consisting of: 
 (i) a polynucleotide comprising the nucleotide sequence of any one of SEQ ID NO:3, SEQ ID NO:5, and SEQ ID NO:7;  
 (ii) a polynucleotide encoding a polypeptide comprising the amino acid sequence of any one of SEQ ID NO:4, SEQ ID NO:6, and SEQ ID NO:8;  
 (iii) a polynucleotide encoding an amino acid sequence comprising the sequence of any one of SEQ ID NO:4, SEQ ID NO:6, and SEQ ID NO:8, in which one or more amino acids are substituted, deleted, inserted and/or added;  
 (iv) a polynucleotide hybridizing under stringent conditions with a polynucleotide consisting of the nucleotide sequence of any one of SEQ ID NO:3, SEQ ID NO:5, and SEQ ID NO:7; and  
 (v) a polynucleotide encoding an amino acid sequence exhibiting 60% or higher percent identity to the amino acid sequence of any one of SEQ ID NO:4, SEQ ID NO:6, and SEQ ID NO:8;  
 
 (B) a microorganism producing the polypeptide of (A); and  
 (C) a processed product of the microorganism of (B).  
 
     
     
         2 . The method of  claim 1 , wherein the microorganism producing the polypeptide is a transformant harboring an expression vector comprising a polynucleotide selected from the group consisting of: 
 (i) a polynucleotide comprising the nucleotide sequence of any one of SEQ ID NO:3, SEQ ID NO:5, and SEQ ID NO:7;    (ii) a polynucleotide encoding a polypeptide comprising the amino acid sequence of any one of SEQ ID NO:4, SEQ ID NO:6, and SEQ ID NO:8;    (iii) a polynucleotide encoding an amino acid sequence comprising the sequence of any one of SEQ ID NO:4, SEQ ID NO:6, and SEQ ID NO:8, in which one or more amino acids are substituted, deleted, inserted and/or added;    (iv) a polynucleotide hybridizing under stringent conditions with a polynucleotide consisting of the nucleotide sequence of any one of SEQ ID NO:3, SEQ ID NO:5, and SEQ ID NO:7; and    (v) a polynucleotide encoding an amino acid sequence exhibiting 60% or higher percent identity to the amino acid sequence of any one of SEQ ID NO:4, SEQ ID NO:6, and SEQ ID NO:8.    
     
     
         3 . The method of  claim 1 , wherein the polypeptide is encoded by a polynucleotide selected from the group consisting of: 
 (A) a polynucleotide comprising the nucleotide sequence of SEQ ID NO:3;    (B) a polynucleotide encoding a polypeptide comprising the amino acid sequence of SEQ ID NO:4;    (C) a polynucleotide encoding an amino acid sequence comprising the sequence of SEQ ID NO:4, in which one or more amino acids are substituted, deleted, inserted and/or added;    (D) a polynucleotide hybridizing under stringent conditions with a polynucleotide consisting of the nucleotide sequence of SEQ ID NO:3; and    (E) a polynucleotide encoding an amino acid sequence exhibiting 60% or higher percent identity to the amino acid sequence of SEQ ID NO:4.    
     
     
         4 . The method of  claim 3 , wherein the polypeptide is encoded by a polynucleotide comprising the nucleotide sequence of SEQ ID NO:3, or a polynucleotide encoding a polypeptide comprising the amino acid sequence of SEQ ID NO:4.  
     
     
         5 . The method of  claim 1 , wherein the microorganism producing the polypeptide is a transformant harboring an expression vector comprising a polynucleotide selected from the group consisting of: 
 (A) a polynucleotide comprising the nucleotide sequence of SEQ ID NO:3;    (B) a polynucleotide encoding a polypeptide comprising the amino acid sequence of SEQ ID NO:4;    (C) a polynucleotide encoding the amino acid sequence comprising the sequence of SEQ ID NO:4, in which one or more amino acids are substituted, deleted, inserted and/or added;    (D) a polynucleotide hybridizing under stringent conditions with a polynucleotide consisting of the nucleotide sequence of SEQ ID NO:3; and    (E) a polynucleotide encoding an amino acid sequence exhibiting 60% or higher percent identity to the amino acid sequence of SEQ ID NO:4.    
     
     
         6 . The method of  claim 5 , wherein the polynucleotide comprises the nucleotide sequence of SEQ ID NO:3, or a nucleotide sequence encoding the amino acid sequence of SEQ ID NO:4.