Protease variants and polynucleotides encoding same

Patent metadata

Field	Value
Publication number	US-9926550-B2
Application number	US-201414899168-A
Country	US
Kind code	B2
Filing date	Jul 28, 2014
Priority date	Jul 29, 2013
Publication date	Mar 27, 2018
Grant date	Mar 27, 2018

Abstract

The present invention relates to protease variants and methods for obtaining protease variants. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

First claim

The invention claimed is:

1. A protease variant, comprising a substitution in the hydrophobic cluster corresponding to position 137 of SEQ ID NO: 3, wherein
(a) the variant has a sequence identity to SEQ ID NO: 3 of at least 85% and less than 100%, and
(b) the variant has protease activity.

2. The variant according to claim 1 , which comprises a substitution at two or more positions corresponding to position 137 and any of the positions 121, 124, or 162.

3. The variant according to claim 2 , wherein the amino acid at the position corresponding to position 124 of SEQ ID NO: 3 is Ala.

4. The variant according to claim 1 , wherein the amino acid at the position corresponding to position 137 of SEQ ID NO: 3 is selected from the group consisting of Glu, Cys, Ser, Ala, Met, Thr, Gln, and Gly.

5. The variant according to claim 2 , wherein the amino acid at the position corresponding to position 162 of SEQ ID NO: 3 is selected from the group consisting of Trp and Arg.

6. The variant of claim 2 , wherein the amino acid at the position corresponding to position 121 of SEQ ID NO: 3 is selected from the group consisting of Ser, Cys, Val and Thr.

7. The variant of claim 1 , which further comprises one or more substitutions selected from the group consisting of L34I, Y39D; T40{D,P,L}; Q70N; T74M; L81{F,H,V}; A102T; R130A, G132 {I,E}; S133T S144{Q,R}; D155N; G159S; V162R, S171 {W, K, E, N}; S173{P,V}; G174{S,T}; S175 {A, V, P}; N176G; T177S; G179 {C, V, Q, S, T, E, H, K, M, N, A, Y}; F180Y; T241P; I247M; H256F, S274I; V286Q and T297P.

8. The variant of claim 2 , comprising any of the following substitutions:

I137M S173P

I121V S175P

I137E S175P

I121V S175A

I137E S175A

I137E S144Q

I137E S144R

I137M S144R

I121T S175P

I137M S144Q

V124A S133T

V124A R130A

I137E S173Y G174S S175A F180Y

V162R S173P G174T S175V T177S F180Y

I121V S173P G174T S175V T177S F180Y

V162R S173P G174K S175P N176G T177S F180Y

I121V S173P G174K S175P N176G T177S F180Y

I137E S173P G174K S175P N176G T177S F180Y

I121V I137E S173Y G174S S175A F180Y

L81V I137E S173Y G174S S175A F180Y

I137E S173Y G174S S175A F180Y T241P

Q70N I137E S173Y G174S S175A F180Y

I137E S173Y G174S S175A F180Y S274I

I137E S173Y G174S S175A F180Y T297P

I137E S173P G174T S175V T177S F180Y T241P

I137E S173P G174T S175V T177S F180Y V286Q

I137E S171N S173P G174T S175V T177S F180Y

I137E S173P G174T S175A T177S F180Y

I121V I137E S173P G174K S175P N176G T177S F180Y

Q70N I137E S173P G174K S175P N176G T177S F180Y

I137E S173P G174K S175P N176G T177S F180Y S274I

I137E S173P G174K S175P N176G T177S F180Y V286Q

I137E S173P G174K S175P N176G T177S F180Y T297P

I137E S171N S173P G174K S175P N176GT177S F180Y

I137E S173P G174K S175A N176G T177S F180Y

V162R S173Y G174S S175A F180Y

I121V S173Y G174S S175A F180Y

I121V S144Q

V162R S173P G174T S175V T177S F180Y

L81V I137E S173Y G174S S175A F180Y

I121V I137E S173P G174T S175V T177S F180Y

I137E S173P G174T S175V T177S F180Y T297P

I137E S171N S173P G174T S175V T177S F180Y

V162R S173Y G174S S175A F180Y

I137E S144Q S173Y G174S S175A F180Y

T40L I137E S173Y G174S S175A F180Y

I137E S173Y S175A F180Y

I137E S173P S175P N176G T177S F180Y

I137E S173Y G174S S175A F180Y V286Q

I137E S173P G174T S175V T177S F180Y S274I

T74M I137E S173P

L34I I137E S173P

Y39D I137E S173P

T40P I137E S173P

I137E S173P I247M

I137E S173P H256F

I137E S144Q S173P G174T S175V T177S F180Y

I137E S144Q S173P G174K S175P N176G T177S F180Y

I137E S173Y G174S S175P F180Y

V162R S171N S173P G174K S175P N176G T177S F180Y

I137E S171N S173P G174K S175P N176G T177S F180Y T241P

I137E S173P S175P F180Y

V162R S173P S175P F180Y

I137E S173P S175V T177S F180Y.

9. The variant of claim 1 , which has an improved detergent stability compared to the protease with SEQ ID NO: 3.

10. The variant of claim 1 , wherein the variant is selected from the group consisting of:
a. a polypeptide having at least 85% sequence identity to the mature polypeptide of SEQ ID NO: 2;
b. a polypeptide encoded by a polynucleotide having at least 85% identity

Assignees

• Novozymes As

Inventors

• Rasmussen Frank Winther
• Lenhard Rolf Thomas
• Toscano Miguel Duarte Guilherme Perreira
• Friis Esben Peter
• Larsen Signe Eskildsen
• Knotzel Jurgen Carsten Franz
• Bauer Michael

Classifications

• C12N9/54 Primary

  bacteria being Bacillus · CPC title

• C11D3/386

  Preparations containing enzymes {, e.g. protease or amylase} · CPC title

• C12Y304/21

  Serine endopeptidases (3.4.21) · CPC title

• C11D3/38681

  Chemically modified or immobilised enzymes · CPC title