IP Library › Granted Patent US 10,093,887
Granted Patent B2
US 10,093,887 · App. 14/337,024 · Granted Oct 9, 2018

Compositions and methods comprising serine protease variants

Inventors: Wolfgang Aehle (Zwingenberg, DE); Luis G. Cascao-Pereira (Redwood City, CA); David A. Estell (San Francisco, CA); Frits Goedegebuur (Vlaardingen, NL); James T. Kellis, Jr. (San Carlos, CA); Ayrookaran J. Poulose (Belmont, CA); Brian F. Schmidt (Half Moon Bay, CA)
Assignee: DANISCO US INC.
C11D3/386C11D3/38609C11D3/38618C11D3/38636C11D3/38645C11D3/38654C12N9/50C12N9/54C12Y304/21062
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Quick Facts
Patent No.
US 10,093,887
App. No.
14/337,024
Granted
Oct 9, 2018
Kind
B2
Abstract

The present invention provides methods for protein engineering and serine protease variants produced there from. Specifically, the present invention provides serine protease variants having one or more substitutions as compared to a reference serine protease. In addition, the present invention provides compositions comprising these serine protease variants. In some embodiments, the present invention provides cleaning compositions comprising at least one of these serine protease variants.

Claims (11)

1. An isolated subtilisin variant of a Bacillus subtilisin, wherein said subtilisin variant is a mature form having proteolytic activity and at least 70% sequence identity to SEQ ID NO:2, and, wherein said variant comprises a combination of substitutions selected from: S87N/G118V/S128L/P129Q/S130A/S24W/S101L/Q109K, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/S101L/Q109R/N185R/A215R, S87N/G118V/S128L/P129Q/S130A/172V/Q109R/S164I, S87N/G118V/S128L/P129Q/S130A/S101Y/Q109R/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/S101Y/Q109L/N185R/A215R, S87R/G118R/S128L/P129Q/S130A/S101K/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/S24W/S101L/Q109L, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/S101Y/Q109R/N185R/A215R, S87N/G118V/S128L/P129Q/S130A/172V/S101L/Q109R/S164I, S87N/G118V/S128L/P129Q/S130A/S101Y/Q109R/S188D/T213E/N248R, S87R/G118R/S128L/P129Q/S130A/172V/S164I/S188D/N248R, S 87N/G118V/S 128L/P129Q/S130A/S24W/S101Y/Q109K, S87N/G118V/S128L/P129Q/S130A/N43S/P52V/S 101R/Q109R, S 87N/G118 V/S128L/P129Q/S130A/172V/S101Y/Q109R/S164I, S87N/G118V/S128L/P129Q/S130A/S101L/Q109L/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/172V/Q109R/S164I/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N76D/S101L, S 87N/G118V/S128L/P129Q/S130A/S24W/S101Y/Q109L, S87N/G118V/S128L/P129Q/S130A/N76D/S164I, S87R/G118R/S128L/P129Q/S130A/N76D/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/S101Y/Q109L/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N76D/S101M, S87N/G118V/S128L/P129Q/S130A/N18K/N76D/G97P/N185R/A215R, S87N/G118V/S128L/P129Q/S130A/I72V/S101L/S164I, S87R/G118R/S128L/P129Q/S130A/S101L/S188D/N248R, S87N/G118 V/S128L/P129Q/S130A/S101Y/Q109L/S188D/T213E/N248R, S87R/G118R/S128L/P129Q/S130A/Q109L/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/S101L/N185R/A215R, S 87N/G118V/S128L/P129Q/S130A/I72V/S101Y/S164I, S87N/G118V/S128L/P129Q/S130A/S101Y/T213E/N248R, S 87N/G118V/S128L/P129Q/S130A/I72V/S101L/Q109L/S164I, S87R/G118R/S128L/P129Q/S130A/S101V/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/S101Y/N185R/A215R, S87N/G118V/S128L/P129Q/S130A/I72V/S101V/S164I, S87N/G118V/S128L/P129Q/S130A/S101L/T213E/N248R, S87N/G118V/S128L/P129Q/S130A/I72V/S101Y/Q109L/S164I, S87R/G118R/S128L/P129Q/S130A/S101H/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/Q109R/N185R/A215R, S87N/G118V/S128L/P129Q/S130A/I72V/S101H/S164I, S87N/G118V/S128L/P129Q/S130A/S101L/Q109R/S188D/N248R, S87N/G118V/S128L/P129Q/S130A/N18K/G97P/S101L/Q109L/N185R/A215R, S87R/G118R/S128L/P129Q/S130A/S101M/S188D/N248R, N76D/S87R/G118R/S128L/P129Q/S130A/S188D, N76D/S87R/G118R/S128L/P129Q/S130A/S188D/N248K, S87N/G118V/G127S/S128L/P129Q/S130A, S87N/S101H/G118V/S128L/P129Q/S130A, S87N/S101K/G118V/S128L/P129Q/S130A, S87N/S101V/G118V/S128L/P129Q/S130A, S87N/S101Y/G118V/S128L/P129Q/S130A, S87N/S101L/G118V/S128L/P129Q/S130A, and I72V/S87N/G118V/S128L/P1290/S130A/S164I, wherein the positions are numbered by correspondence with the amino acid sequence of B. amyloliquefaciens subtilisin BPN′ set forth as SEQ ID NO:1.

2. A cleaning composition comprising the subtilisin variant set forth in claim 1 .

3. The cleaning composition of claim 2 , wherein said composition comprises a liquid, gel, tablet, powder and/or granule detergent.

4. The cleaning composition of claim 2 , further comprising at least one additional enzyme.

5. The cleaning composition of claim 4 , wherein said at least one additional enzyme is selected from hemicellulases, cellulases, peroxidases, proteases, metalloproteases, xylanases, lipases, phospholipases, esterases, perhydrolasess, cutinases, pectinases, pectate lyases, mannanases, keratinases, reductases, oxidases, phenoloxidases, lipoxygenases, ligninases, pullulanases, tannases, pentosanases, malanases, ß-glucanases, arabinosidases, hyaluronidase, chondroitinase, laccase, and amylases, or mixtures thereof.

6. The cleaning composition of claim 2 , wherein said cleaning composition is selected from laundry detergents and dish detergents.

7. The cleaning composition of claim 2 , comprising a laundry detergent.

8. The cleaning composition of claim 7 , wherein said laundry detergent is a heavy duty detergent.

9. The cleaning composition of claim 2 , further comprising at least one bleaching agent.

10. The cleaning composition of claim 2 , wherein said cleaning composition is a phosphate-free detergent.

11. The cleaning composition of claim 2 , wherein said cleaning composition is a cold water detergent.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Oct 9, 2014
From: AEHLE, WOLFGANG; CASCAO-PEREIRA, LUIS G.; ESTELL, DAVID A.; SCHMIDT, BRIAN F.; GOEDEGEBUUR, FRITS; KELLIS, JAMES T., JR.; POULOSE, AYROOKARAN J.
To: DANISCO US INC.
Reel/Frame 033926/0436 →
Continuity (6)
Continuation 12616097 · Nov 10, 2009
Provisional Application 61113545 · Nov 11, 2008
Provisional Application 61113552 · Nov 11, 2008
Provisional Application 61113548 · Nov 11, 2008
Provisional Application 61218802 · Jun 19, 2009
Related Publication 20150031589A1 · Jan 29, 2015
Cited By (3)
US 12,286,604 US 12,286,605 US 12,509,672