Ketoreductase polypeptides and uses thereof

1. A ketoreductase polypeptide capable of enantiospecifically reducing (R)-2-methylpentanal to (R)-2-methylpentanol with an enantiomeric excess, wherein the polypeptide amino acid sequence has at least 90% identity to a reference sequence of SEQ ID NO: 4, wherein the residue corresponding to X153 is glutamine.

2. The polypeptide of claim 1 , wherein the residue corresponding to X145 is serine.

3. The polypeptide of claim 1 wherein the amino acid sequence further comprises one or more of the following features:

residue corresponding to X72 is glutamic acid;

residue corresponding to X82 is serine;

residue corresponding to X96 is alanine, arginine, glutamine or lysine;

residue corresponding to X104 is methionine;

residue corresponding to X117 is serine;

residue corresponding to X147 is cysteine, leucine, methionine or valine;

residue corresponding to X157 is threonine;

residue corresponding to X177 is glutamic acid;

residue corresponding to X178 is glycine;

residue corresponding to X202 is arginine, isoleucine or valine;

residue corresponding to X223 is a glycine, methionine, alanine, valine, leucine or isoleucine;

residue corresponding to X236 is arginine; and

wherein the amino acid sequence can optionally have one or more differences at other amino acid residues as compared to the reference sequence.

4. The polypeptide of claim 1 , wherein the polypeptide has an E-value ≧51 for the reduction of (R)-2-methylpentanal in a mixture with the (S)-2-methylpentanal.

5. The polypeptide of claim 1 , wherein the percent enantiomeric excess is at least 98%.

6. The polypeptide of claim 1 , wherein the polypeptide comprises an amino acid sequence of SEQ ID NO: 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, or 44.

7. The polypeptide of claim 1 , wherein >0 but ≦2 g/L of the ketoreductase polypeptide is capable of converting at least about 45% of ≧200 g/L (R)-2-methylpentanal to (R)-2-methylpentanol in less than 24 hours.

8. The polypeptide of claim 7 , wherein the polypeptide comprises an amino acid sequence of SEQ ID NO: 24, 26, 28, or 44.

9. A method for resolving a mixture of (R)-2-methylpentanal and (S)-2-methylpentanal by reduction of the (R)-enantiomer to a (R)-2-methylpentanol product, which comprises contacting the mixture with the ketoreductase polypeptide of claim 1 under reaction conditions suitable for reducing the (R)-2-methylpentanal to (R)-2-methylpentanol.

10. The method of claim 9 , wherein the mixture comprises a cofactor for the ketoreductase polypeptide and optionally a cofactor regenerating system.

11. The method of claim 10 , wherein cofactor regenerating system comprises glucose dehydrogenase and glucose; formate dehydrogenase and formate; or isopropanol and a secondary alcohol dehydrogenase.

12. The method of claim 11 , wherein the secondary alcohol dehydrogenase is the ketoreductase polypeptide.

13. The method of claim 9 , wherein the mixture of (R)-2-methylpentanal and (S)-2-methylpentanal is a racemic mixture.

14. The method of claim 9 , wherein at least about 45% of the (R)-2-methylpentanal is reduced to the (R)-2-methylpentanol product in less than 24 hours when the method is conducted with ≧220 g/L (R)-2-methylpentanal and >0 but ≦2 g/L of the ketoreductase polypeptide.

15. The method of claim 9 , which further comprises recovering the (R)2-methylpentanol.

16. The method of claim 9 , wherein the method comprises contacting the mixture with whole cells that express the ketoreductase polypeptide, or an extract or lysate of such cells.

17. A composition comprising a compound of structural formula (I)

and/or a compound of structural formula (II)

and a ketoreductase of claim 1 .