IP Library › Granted Patent US 12,415,996
Granted Patent B2
US 12,415,996 · App. 17/251,014 · Granted Sep 16, 2025

Subtilisin variants

Inventors: Viktor Yuryevich Alekseyev (Palo Alto, CA); Lilia Maria Babe (Palo Alto, CA); H. Billur Engin (Palo Alto, CA); David A. Estell (Palo Alto, CA); Frits Goedegebuur (Leiden, NL); Thijs Kaper (Palo Alto, CA); Harm Mulder (Leiden, NL); Sina Pricelius (Leiden, NL); Sander Van Stigt Thans (Leiden, NL)
Assignee: Danisco US Inc.
C12N9/54C11D3/386C11D2111/12C12Y304/21062
View Patent ↗
Loading inventors, assignments & file history…
Monitor This Case
Get email alerts when status or documents change.
Order Certified Copies
Most orders are placed with the USPTO same day — all within 24 business hours.
Order via The Patent Place →
Pre-filled with this patent's details
Quick Facts
Patent No.
US 12,415,996
App. No.
17/251,014
Granted
Sep 16, 2025
Kind
B2
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.

Claims (14)

1. A subtilisin variant comprising an amino acid sequence having a glutamate (E) residue at position 39 and further comprising an amino acid substitution X211E/Q/R wherein the subtilisin variant has at least 85% sequence identity to SEQ ID NO: 1; and

wherein the amino acid positions of the variant are numbered by correspondence with the amino acid sequence of SEQ ID NO: 1.

2. The subtilisin variant according to claim 1 , wherein said variant

(i) is isolated;

(ii) has proteolytic activity; or

(iii) comprises a combination of (i) to (ii).

3. The subtilisin variant 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: 1.

4. The subtilisin variant of claim 1 , wherein said variant has one or more improved property when compared to a parent or reference subtilisin; wherein the improved property is selected from improved cleaning performance in detergent, improved stability; and combinations thereof.

5. The subtilisin variant of claim 4 , wherein the improved property is

(i) improved cleaning performance in detergent, wherein said variant has a blood-milk-ink (BMI), crème brûlée and/or egg stain cleaning performance index (PI)≥1.1 compared to the subtilisin having the amino acid sequence of SEQ ID NO: 1; and/or

(ii) improved stability, wherein said variant has a stability PI≥1.1 compared to the subtilisin having the amino acid sequence of SEQ ID NO: 1.

6. An enzyme composition comprising one or more subtilisin variant according to claim 1 .

7. The enzyme composition according to claim 6 , wherein said enzyme composition is an enzyme granule.

8. The composition according to claim 6 , further comprising (i) one or more other enzymes selected from 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, tannases, transglutaminases, xylan acetyl-esterases, xylanases, and xylosidases; (ii) one or more surfactants; (iii) one or more ions selected from 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 subtilisin variant; (vii) one or more bleaching agents; and/or (viii) combinations thereof.

Assignments (3)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Apr 14, 2025
From: GENENCOR INTERNATIONAL B.V.
To: DANISCO US INC.
Reel/Frame 070828/0925 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Mar 14, 2025
From: BABE, LILIA MARIA; ENGIN, H. BILLUR; KAPER, THIJS; ESTELL, DAVID A.
To: DANISCO US INC.
Reel/Frame 070509/0531 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Feb 19, 2025
From: ALEKSEYEV, VIKTOR YURYEVICH; GOEDEGEBUUR, FRITS; MULDER, HARM; PRICELIUS, SINA; VAN STIGT THANS, SANDER
To: GENENCOR INTERNATIONAL B.V.
Reel/Frame 070254/0129 →
Continuity (2)
Provisional Application 62686809 · Jun 19, 2018
Related Publication 20210214703A1 · Jul 15, 2021
References Cited (20)
US 11499146B2 · Babe · 2022 [cited by examiner]
US 20190185789A1 · Souter · 2019 [cited by examiner]
US 20190330610A1 · Babe · 2019 [cited by examiner]
US 20220098524A1 · Souter · 2022 [cited by examiner]
WO WO2016205755A1 · 2016 [cited by examiner]
WO 2017192692A1 · 2017 [cited by applicant]
WO WO2018118950A1 · 2018 [cited by examiner]
WO WO2019125984A1 · 2019 [cited by examiner]
WO WO2019125985A1 · 2019 [cited by examiner]
WO WO2019125986A1 · 2019 [cited by examiner]
Martinez. Increasing activity and thermal resistance of Bacillus gibsonii alkaline protease (BgAP) by directed evolution. Biotechnol. Bioeng., 110: 711-720. (2013). [cited by examiner]
Fransceus (J Ind Microbiol Biotechnol. May 2017;44(4-5):687-695. [cited by examiner]
Sanavia. Computational and Structural Biotechnology Journal, vol. 18, 2020, pp. 1968-1979. [cited by examiner]
Studer. Residue mutations and their impact on protein structure and function: detecting beneficial and pathogenic changes. Biochem. J. (2013) 449, 581-594. [cited by examiner]
A0A1H9VK28_9BACI. UniProtKB/TrEMBL Database. Nov. 22, 2017. [cited by examiner]
Deng, Aihua et al., Secretory Expression, Functional Characterization, and Molecular Genetic Analysis of Novel Halo-Solvent-Tolerant Protease from Bacillus gibsonii, Journal of Microbiology and Biotechnology, Feb. 28, 2… [cited by applicant]
Jakob, Felix et al., Surface charge engineering of a Bacillus gibsonii subtilisin protease, Applied Microbiology and Biotechnology, Nov. 21, 2012, pp. 6793-6802, vol. 97, No. 15. [cited by applicant]
Martinez, Ronny et al., Increasing Activity and Thermal Resistance of Bacillus gibsonii Alkaline Protease (BgAP) by Directed Evolution, Biotechnology and Bioengineering, Nov. 1, 2012, pp. 711-720, vol. 110, No. 3. [cited by applicant]
Vojcic, Ljubica et al., Advances in protease engineering for laundry detergents, New Biotechnology, Dec. 1, 2015, pp. 629-634, vol. 32, No. 6. [cited by applicant]
PCT/US2019/033873—International Search Report and Written Opinion—mailed Sep. 17, 2019. [cited by applicant]