Biocatalytic process for preparing eslicarbazepine and analogs thereof

Patent metadata

Field	Value
Publication number	US-9365878-B2
Application number	US-201514790311-A
Country	US
Kind code	B2
Filing date	Jul 2, 2015
Priority date	Apr 13, 2011
Publication date	Jun 14, 2016
Grant date	Jun 14, 2016

Abstract

The present disclosure relates to biocatalysts and its uses for the efficient preparation of eslicarbazepine, eslicarbazepine acetate, and analogs thereof.

First claim

What is claimed is:

1. A non-naturally occurring polypeptide capable of converting compound (2c)

to compound (1c)

in enantiomeric excess in presence of NADPH, the ketoreductase polypeptide comprising an amino acid sequence that has at least 80% sequence identity to SEQ ID NO:10 and has at least the following features: X80 is T; X96 is V or R; X145 is L; X153 is T; X190 is P; X196 is L or M; and X226 is V.

2. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence has at least 93% sequence identity to SEQ ID NO: 36.

3. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence further comprises one or more of the following features: X17 is H; X29 is T; X43 is R; X71 is P or G; X87 is L; X95 is Y; X131 is C; X173 is L; and X199 is M.

4. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence further comprises one or more of the following features: X17 is M or H; X29 is T; X40 is R; X43 is R or V; X64 is V; X94 is G or A; X95 is Y or M; X147 is Q or M; X152 is L or A; X157 is C or S; X173 is L; X199 is M; and X200 is P.

5. The non-naturally occurring polypeptide of claim 1 , where the amino acid sequence further comprises one or more of the following features: X25 is T; X76 is A; X144 is V; X150 is L; X194 is R; X233 is G; and X249 is W or F.

6. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence comprises at least the following features: X64 is V; X71 is P or G; X80 is T; X87 is L; X94 is G or A; X96 is V or R; X145 is L; X147 is Q or M; X153 is T; X173 is L; X190 is P; X196 is M or L; X199 is M; and X226 is V.

7. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence further comprises one or more of the following features: X17 is M or H; X29 is T; X40 is R; X43 is R or V; X95 is M or Y; X131 is C; X152 is L or A; and X200 is P.

8. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence further comprises one or more of the following features: X25 is T; X76 is A; X144 is V; X150 is L; X157 is C or S; X194 is R; X233 is G; and X249 is W or F.

9. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence comprises at least the following features: X17 is H or M; X25 is T; X29 is T; X40 is R; X43 is R or V; X64 is V; X71 is G or P; X80 is T; X87 is L; X94 is G; X95 is Y or M; X96 is R or V; X131 is C; X145 is L; X147 is Q or M; X152 is A or L; X153 is T; X157 is S or C; X173 is L; X190 is P; X196 is M or L; X199 is M; X200 is P; and X226 is V.

10. The non-naturally occurring polypeptide of claim 9 , wherein the amino acid sequence further comprises one or more of the following features: X76 is A; X144 is V; X150 is L; X194 is R; X233 is G; and X249 is W or F.

11. The non-naturally occurring polypeptide of claim 1 , wherein the amino acid sequence comprises a sequence selected from SEQ ID NO: 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, and 38.

12. A non-naturally occurring polypeptide comprising an amino acid sequence having at least 80% sequence identity to SEQ ID NO:2 and having one or more residue differences as compared to the reference sequence of SEQ ID NO:2 at residue positions corresponding to X71, X87, and X131, wherein the polypeptide has ketoreductase activity.

13. The non-naturally occurring polypeptide of claim 12 , wherein the amino acid sequence comprises one or more of the following features: X71 is P or G; X87 is L, and X131 is C.

14. The non-naturally occurring polypeptide of claim 1 , wherein the polypeptide is immobilized on a solid support.

15. The non-naturally occurring polypeptide of claim 12 , wherein the polypeptide is immobilized on a solid support.

Assignees

• Codexis Inc

Inventors

• Gohel Anupam
• Smith Derek J
• Wong Brian
• Sukumaran Joly
• Yeo Wan Lin
• Collier Steven J
• Novick Scott

Classifications

• C12P17/10 Primary

  Nitrogen as only ring hetero atom · CPC title

• C12N9/0006

  acting on CH-OH groups as donors (1.1) · CPC title

• C12N11/00

  Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof · CPC title

• C12Y101/00

  Oxidoreductases acting on the CH-OH group of donors (1.1) · CPC title

• C07D223/22

  Dibenz [b, f] azepines; Hydrogenated dibenz [b, f] azepines · CPC title