Protease variants and uses thereof

Patent metadata

Field	Value
Publication number	US-2026055390-A1
Application number	US-202519349067-A
Country	US
Kind code	A1
Filing date	Oct 3, 2025
Priority date	Dec 21, 2016
Publication date	Feb 26, 2026
Grant date	—

Abstract

Disclosed herein is one or more subtilisin variant, nucleic acid encoding same, and compositions and methods related to the production and use thereof, including one or more subtilisin variant that has improved stability and/or soil removal compared to one or more reference subtilisin.

First claim

1 . A subtilisin variant comprising an amino acid sequence comprising an amino acid substitution of 129E at position 129,
wherein the amino acid positions of the variant are numbered by correspondence with the amino acid sequence of SEQ ID NO: 17, wherein said variant comprises an amino acid sequence with 85% sequence identity to the amino acid sequence of SEQ ID NO:11, and wherein ad variant has an improved property when compared to a parent or reference subtilisin of SEQ ID NO: 11, wherein the improved property is improved cleaning performance in detergent.

2 - 4 . (canceled)

5 . The subtilisin variant of claim 1 , wherein said variant comprises an amino acid sequence with
86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or less than 100% amino acid sequence identity to the amino acid sequence of SEQ ID NO:11.

6 - 7 . (canceled)

8 . The subtilisin variant of claim 1 , wherein said variant has a blood/milk/ink (BMI) cleaning PI>1.

9 . The subtilisin variant of claim 1 , wherein said
cleaning performance in detergent is measured in accordance with the cleaning performance in laundry (HDL) and ADW detergents assay of Example 1.

10 . A composition comprising one or more subtilisin variant according to claim 1 .

11 . The composition according to claim 10 , wherein said composition is selected from the group consisting of an enzyme composition and a detergent composition.

12 . The composition according to claim 11 , wherein said detergent composition is selected from the group consisting of a laundry detergent, a fabric softening detergent, a dishwashing detergent, and a hard-surface cleaning detergent.

13 . The composition according to claim 10 , further comprising (i) one or more other enzymes selected from the group consisting of acyl transferases, amylases, alpha-amylases, beta-amylases, alpha-galactosidases, arabinases, arabinosidases, aryl esterases, beta-galactosidases, beta-glucanases, carrageenases, catalases, chondroitinases, cutinases, endo-beta-mannanases, exo-beta-mannanases, esterases, exo-mannanases, galactanases, glucoamylases, hemicellulases, hyaluronidases, keratinases, laccases, lactases, ligninases, lipases, lipolytic enzymes, lipoxygenases, mannanases, metalloproteases, nucleases, oxidases, oxidoreductases, pectate lyases, pectin acetyl esterases, pectinases, pentosanases, perhydrolases, peroxidases, phenoloxidases, phosphatases, phospholipases, phytases, polyesterases, polygalacturonases, additional proteases, pullulanases, reductases, rhamnogalacturonases, cellulases, beta-glucanases, tannases, transglutaminases, xylan acetyl-esterases, xylanases, and xylosidases; (ii) one or more surfactants; (iii) one or more ions selected from the group consisting of calcium and zinc; (iv) one or more adjunct material; (v) one or more stabilizers; (vi) from about 0.001% to about 1.0 weight % of the variant; (vii) one or more bleaching agents; and/or (viii) combinations thereof.

14 . The composition of claim 10 , wherein said composition is phosphate-free or contains phosphate and/or is boron free or contains boron.

15 . The composition of claim 10 , wherein said composition is a granular, powder, solid, bar, liquid, tablet, gel, paste or unit dose composition.

16 . A method of cleaning, comprising contacting a surface or an item in need of cleaning with an effective amount of a variant of claim 1 or the composition of claim 10 ; and optionally further comprising the step of rinsing said surface or item after contacting said surface or item with said variant or composition.

17 . (canceled)

18 . The method of claim 16 , wherein said item is fabric.

19 . A polynucleotide comprising a nucleic acid sequence encoding a variant of claim 1 , wherein said polynucleotide is, optionally, isolated.

20 . The polynucleotide of claim 19 , wherein the nucleic acid sequence is operably linked to a promoter.

21 . An expression vector or cassette comprising the polynucleotide of claim 19 .

22 . A recombinant host cell comprising the polynucleotide of claim 19 or the vector or cassette of claim 21 .

23 . A composition comprising a variant of claim 1 wherein said composition is a disinfectant, medical instrument cleaning, animal feed, contact lens cleaning, wound cleaning, or textile, leather or feather processing composition.

24 - 30 . (canceled)

31 . A method of cleaning comprising contacting a surface or an item in need of cleaning with an effective amount of a subtilisin variant or composition containing a subtilisin variant, wherein said variant comprises an amino acid sequence comprising an amino acid substitution of 129E at position 129,
wherein the amino acid positions of the variant are numbered by correspondence with the amino acid sequence of SEQ ID NO: 17, wherein said variant comprises an amino acid sequence with 85% sequence identity to the amino acid sequence of SEQ ID NO: 11, and wherein said variant has an improved property when compared to a parent or reference subtilisin of SEQ ID NO: 11, wherein the improved property is improved cleaning performance in detergent.

32 . The method of claim 31 , wherein the method cleans a blood/milk/ink (BMI) stain.

33 . (canceled)

34 . A subtilisin variant comprising an amino acid sequence comprising a first amino acid substitution of 129E at position 129 and a second amino acid substitution of 39E at position 39, wherein said variant comprises an amino acid sequence with 85% sequence identity to the amino acid sequence of SEQ ID NO:11, and wherein said variant has an improved property when compared to a parent or reference subtilisin of SEQ ID NO:11 comprising a 39E substitution, wherein the improved property is improved cleaning performance.

35 . The subtilisin of claim 34 , wherein improved cleaning performance is improved ADW cleaning performance.

36 . The subtilisin of claim 34 , wherein said variant has a crème brûlée cleaning PI>1.

Assignees

• Danisco Us Inc

Inventors

• Babe Lilia Maria
• Dankmeyer Lydia
• Estell David A
• Goedegebuur Frits
• Kaper Thijs
• Mulder Harm
• Pricelius Sina

Classifications

• C12Y304/21062

  Subtilisin (3.4.21.62) · CPC title

• C11D3/386

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

• C12N9/64

  derived from animal tissue · CPC title

• C12N9/54 Primary

  bacteria being Bacillus · CPC title