Engineered amylase variants

Patent metadata

Field	Value
Publication number	US-11767519-B2
Application number	US-202117460147-A
Country	US
Kind code	B2
Filing date	Aug 27, 2021
Priority date	Aug 28, 2020
Publication date	Sep 26, 2023
Grant date	Sep 26, 2023

Abstract

The present invention provides engineered amylase polypeptides and compositions thereof. The engineered amylase polypeptides have been optimized to provide improved thermostability, protease stability, and stability under a range of pH conditions, including acidic (pH<7) conditions. The invention also relates to the use of the compositions comprising the engineered amylase polypeptides for therapeutic and/or nutritional purposes. The present invention also provides polynucleotides encoding the engineered amylase polypeptides, as well as methods for making the engineered polynucleotides and amylase polypeptides.

First claim

What is claimed is:

1. A recombinant amylase comprising a polypeptide sequence comprising at least 90% sequence identity to SEQ ID NO: 1728, wherein the polypeptide sequence comprises at least a substitution 272G/K/P/R/S, 394E/G/N/R/S, or 91H, or any combinations thereof, wherein the amino acid positions are relative to SEQ ID NO: 2.

2. The recombinant amylase of claim 1 , wherein the polypeptide sequence comprises SEQ ID NO: 156, 174, 276, 308, 422, 456, 546, 632, 664, 780, 810, 1104, 1122, 1224, 1256, 1370, 1404, 1494, 1580, 1612, 1728, or 1758.

3. The recombinant amylase of claim 1 , wherein said recombinant amylase exhibits at least one improved property selected from: i) enhanced catalytic activity; ii) increased tolerance to acidic pH conditions; iii) increased tolerance to at least one protease; iv) increased tolerance to at least one bile salt; v) increased thermotolerance; or a combination of any of i), ii), iii), iv), and v), as compared to a reference amylase having the sequence of SEQ ID NO: 2 or 18.

4. A composition comprising at least one recombinant amylase of claim 1 .

5. A pharmaceutical composition comprising the recombinant amylase of claim 1 , and a pharmaceutically acceptable carrier or excipient.

6. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 91% sequence identity to SEQ ID NO: 1728.

7. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 95% sequence identity to SEQ ID NO: 1728.

8. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 99% sequence identity to SEQ ID NO: 1728.

9. The recombinant amylase of claim 1 , wherein the polypeptide sequence comprises at least the substitution 272G, 272K, 272P, 272R, or 272S.

10. The recombinant amylase of claim 1 , wherein the polypeptide sequence comprises at least the substitution 394E, 394G, 394N, 394R, or 394S.

11. The recombinant amylase of claim 1 , wherein the polypeptide sequence comprises at least the substitution 91H.

12. The recombinant amylase of claim 1 , wherein the polypeptide sequence comprises at least the substitution 272G, 394G, or 91H, or any combinations thereof.

13. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 92% sequence identity to SEQ ID NO: 1728.

14. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 93% sequence identity to SEQ ID NO: 1728.

15. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 94% sequence identity to SEQ ID NO: 1728.

16. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 96% sequence identity to SEQ ID NO: 1728.

17. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 97% sequence identity to SEQ ID NO: 1728.

18. The recombinant amylase of claim 1 , comprising a polypeptide sequence comprising at least 98% sequence identity to SEQ ID NO: 1728.

Assignees

• Codexis Inc
• Nestle Sa

Inventors

• Chng Chinping
• Dellas Nikki
• Duan Da
• Garcia Ravi David
• Maniar Harvinder Chagger

Classifications

• C12N9/2417

  from microbiological source · CPC title

• C12N9/2414 Primary

  Alpha-amylase (3.2.1.1.) · CPC title

• A61K38/47

  acting on glycosyl compounds (3.2), e.g. cellulases, lactases · CPC title

• A61P1/18

  for pancreatic disorders, e.g. pancreatic enzymes · CPC title

• C12Y302/01001

  Alpha-amylase (3.2.1.1) · CPC title