Biocatalysts and methods for the synthesis of substituted lactams

Patent metadata

Field	Value
Publication number	US-2017191043-A1
Application number	US-201715449677-A
Country	US
Kind code	A1
Filing date	Mar 3, 2017
Priority date	Sep 8, 2011
Publication date	Jul 6, 2017
Grant date	—

Abstract

The present disclosure relates to transaminase polypeptides capable of aminating a dicarbonyl substrate, and polynucleotides, vectors, host cells, and methods of making and using the transaminase polypeptides.

First claim

What is claimed is:

1 . An engineered polypeptide comprising an amino acid sequence having at least 80% sequence identity to SEQ ID NO:4, and a substitution at position X136 selected from X136F, X136K, X136L, X136M, and X136Y.

2 . The engineered polypeptide of claim 1 , wherein the amino acid sequence further comprises one or more residue differences as compared to SEQ ID NO:4 selected from: X42G; X54P; X54R; X69A; X69G; X69T; X122M; X124F; X124L; X124N; X124R; X126A; X126T; X150S; X152C; X152G; X152I; X152L; X152S; X156A; X156S; X156T; X157L; X165N, X192A, X193G, X192H, X192K, X192N, X193Q, X192R, and X192S.

3 . The engineered polypeptide of claim 1 , wherein the amino acid sequence further comprises a combination of residue differences selected from:
(a) X124W and X327L; (b) X209M and X300G; (c) X122F, X223V and X284A; (d) XI92A, X215H and X311T; (e) X62N, X124F, X126A and X136L; (f) X124W, X126A, X136L, X192A and X284A; and (g) X124W, X126A, X136L, X152R/X152L/X152I, and X192A.

4 . The engineered polypeptide of claim 1 , in which the amino acid sequence further comprises at least one or more residue differences as compared to SEQ ID NO:4 selected from: X54A; X54L; X56D; X56E; X61G; X61W; X62A; X62L; X62N; X62S; X64C; X68I; X122F, X122W; X122Y; X124K; X124M; X124S; X124W; X126C; X126I; X139E; X140K; X143V; X150L; X155I; X155L; X159G; X160L; X176C; X199L; X199V; X209F; X209M; X209Q; X209V; X223S; X223T; X223V; X227I; X282L, X282V; X284A; X284P; X284S; X284T; and X284V.

5 . An engineered polynucleotide encoding the polypeptide of claim 1 .

6 . A vector comprising the polynucleotide of claim 5 .

7 . A host cell comprising the vector of claim 6 .

8 . A method of converting compound 2 to compound 1,

comprising contacting compound 2 with the engineered polypeptide of claim 1 , under suitable reaction conditions to produce compound 1.

9 . A method of converting compound 2 to compound 1d and compound 1a

comprising contacting compound 2 with the engineered polypeptide of claim 1 , under suitable reaction conditions to produce compound 1d and 1a, under suitable reaction conditions wherein said engineered polypeptide comprises at least 1.5 fold the activity of SEQ ID NO:4.

Assignees

• Codexis Inc

Inventors

• Cabirol Fabien Louis
• Chen Haibin
• Gohel Anupam
• Salim Paulina Elena
• Smith Derek J
• Janey Jacob
• Kosjek Birgit
• Tang Weng Lin
• Hsieh Helen
• Pham Son Q

Classifications

• C12N9/1096 Primary

  transferring nitrogenous groups (2.6) · CPC title

• C12P17/12

  containing a six-membered hetero ring · CPC title

• C12Y206/01

  Transaminases (2.6.1) · CPC title

• C07D211/76

  attached in position 2 or 6 · CPC title

• C12P13/005

  Amino acids other than alpha- or beta amino acids, e.g. gamma amino acids · CPC title