Compositions and methods comprising serine protease variants
원문보기
IPC분류정보
국가/구분
United States(US) Patent
등록
국제특허분류(IPC7판)
C12N-009/54
C11D-003/386
출원번호
US-0115801
(2012-05-04)
등록번호
US-9856466
(2018-01-02)
국제출원번호
PCT/US2012/036608
(2012-05-04)
§371/§102 date
20131105
(20131105)
국제공개번호
WO2012/151534
(2012-11-08)
발명자
/ 주소
Amin, Neelam S.
Augustyn, Katherine
Basler, Joshua R.
Cascao-Pereira, Luis G.
Collier, Katherine D.
Concar, Edward M.
Estell, David A.
Kellis, Jr., James T.
Magennis, Euan John
Pisarchik, Alexander
Poulose, Ayrookaran J.
Souter, Philip Frank
Ward, Glenn Steven
Yao, Jian
출원인 / 주소
DANISCO US INC.
인용정보
피인용 횟수 :
0인용 특허 :
72
초록▼
The present invention provides serine protease variants, more specifically subtilisin variants produced there from. Specifically, the present invention provides serine protease variants, more specifically subtilisin variants having one or more substitutions as compared to a reference serine protease
The present invention provides serine protease variants, more specifically subtilisin variants produced there from. Specifically, the present invention provides serine protease variants, more specifically subtilisin 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, more specifically subtilisin variants. In some embodiments, the present invention provides cleaning compositions comprising at least one of these serine protease variants, more specifically subtilisin variants.
대표청구항▼
1. An isolated subtilisin variant, wherein said subtilisin variant is a mature form having proteolytic activity and comprises an amino acid sequence comprising a combination of amino acid substitutions at positions selected from T022R+S101G+Q245R, wherein said subtilisin variant has at least 80% ami
1. An isolated subtilisin variant, wherein said subtilisin variant is a mature form having proteolytic activity and comprises an amino acid sequence comprising a combination of amino acid substitutions at positions selected from T022R+S101G+Q245R, wherein said subtilisin variant has at least 80% amino acid identity with a Bacillus lentus subtilisin GG36 protease comprising the amino acid sequence shown in SEQ ID NO:2, and wherein amino acid positions of the subtilisin variant are numbered according to the numbering of corresponding amino acid positions in the amino acid sequence of Bacillus amyloliquefaciens subtilisin BPN′ shown in SEQ ID NO:1. 2. The isolated subtilisin variant of claim 1, wherein the subtilisin variant is a subtilisin variant of Bacillus lentus subtilisin GG36 protease, wherein said Bacillus lentus subtilisin GG36 protease comprises the amino acid sequence shown in SEQ ID NO:2. 3. The isolated subtilisin variant of claim 1, wherein the total net charge of the variant is 0, +1, +2, +3, +4, +5, −1, −2, −3, −4, or −5 relative to the total net charge of the Bacillus lentus subtilisin GG36 protease. 4. The isolated subtilisin variant of claim 3, wherein the total net charge is obtained by one or more substitutions selected from: A001E, V004E, R010H/A, Q012E, A015D, N018D, R019H/S, V026D, K027E, N043D, R045C/T/S/P, S049D, V051E, G061E, N062D/E, N076D, N077D, S078D, S087D, A098E, S101D, S106E, G115E, G118D, N123D, S128D, P129E, S130D/E, S132D, A158E, G159D/E, S160D, S166D, R170T, N183D, N184D, N185E, R186H, S188D/E, A194E, A200D, Y209E, A215D, N204D, S212D, L217D/E, N218D, A230E, K235F, K237E, N238D, S240D, N243D, Q245D, R247L, N248D/E, K251C, N263D, N269D/E, A272D, A273E, or R275H/S, and wherein amino acid positions of the protease variant are numbered according to the numbering of corresponding amino acid positions in the amino acid sequence of Bacillus amyloliquefaciens subtilisin BPN′ shown in SEQ ID NO:1. 5. The isolated subtilisin variant of claim 3, wherein the total net charge is obtained by one or more substitutions selected from: A001R, V004R, Q012R, P014R, H017R, N018R/K, G020K/R, T022R/K, S024R/K, G025R, D032I, D041K/L/N, N043R/K, G046R, A048R, F050R/K, P055R, S056R/K, T057R, Q059K, G061R, N076K, S078R, P086R, S087R, E089G/P/I, G097R, S099R, Q109R, G115R, G118R, S132K, S144R, G159R/K, D181C/S/T, L196K, N204K, Q206R, K235R, Q236R/K, K237R, N238R, S240R, W241R, S242R/K, V244R, Q245R/K, N248R, H249R, N252R/K, S256R, G258R, T260K, N269R/K, or E271F/H/T/L/W/R/S/I/A/G/V, and wherein amino acid positions of the protease variant are numbered according to the numbering of corresponding amino acid positions in the amino acid sequence of Bacillus amyloliquefaciens subtilisin BPN′ shown in SEQ ID NO:1. 6. The isolated subtilisin variant of claim 1, wherein the subtilisin variant has one or more of the following characteristics: a) a Test Method 2 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 10, from 1.1 to about 8, or even from 1.1 to about 5; b) a Test Method 3 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 10, from 1.1 to about 8, or even from 1.1 to about 5; c) a Test Method 4 performance index of at least 1.0, at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.0 to about 10, from 1.0 to about 8, or even from 1.0 to about 5; and/or d) a Test Method 6 performance index of at least 1.0, at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.0 to about 10, from 1.0 to about 8, or even from 1.0 to about 5; and/or e) Test Method 7 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 15, from 1.1 to about 10, or even from 1.1 to about 7. 7. A composition comprising at least one subtilisin variant of claim 1, wherein said composition is not a fabric and home care product. 8. The composition of claim 7, wherein said composition is a cleaning composition. 9. The composition of claim 8, wherein said cleaning composition is a granular, powder, solid, bar, liquid, tablet, gel, or paste composition. 10. The composition of claim 7, further comprising at least one bleaching agent. 11. The composition of claim 7, wherein said cleaning composition is phosphate-free. 12. The composition of claim 7, wherein said cleaning composition contains phosphate. 13. The composition of claim 7, further comprising at least one additional enzyme. 14. The composition of claim 13, wherein said at least one additional enzyme is selected from hemicellulases, cellulases, peroxidases, proteases, metalloproteases, xylanases, lipases, phospholipases, esterases, perhydrolases, cutinases, pectinases, pectate lyases, mannanases, keratinases, reductases, oxidases, phenoloxidases, lipoxygenases, ligninases, pullulanases, tannases, pentosanases, malanases, β-glucanases, arabinosidases, hyaluronidases, chondroitinases, laccases, and amylases, or any combination thereof. 15. The composition of claim 7, wherein said subtilisin variant is not a cold water protease variant, wherein said cold water protease variant has one or more of the following characteristics: a) a Test Method 2 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 10, from 1.1 to about 8, or even from 1.1 to about 5; b) a Test Method 3 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 10, from 1.1 to about 8, or even from 1.1 to about 5; c) a Test Method 4 performance index of at least 1.0, at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.0 to about 10, from 1.0 to about 8, or even from 1.0 to about 5; and/or d) a Test Method 6 performance index of at least 1.0, at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.0 to about 10, from 1.0 to about 8, or even from 1.0 to about 5; and/or e) Test Method 7 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 15, from 1.1 to about 10, or even from 1.1 to about 7. 16. A method of cleaning, comprising contacting a surface or an item with a cleaning composition comprising at least one subtilisin variant of claim 1. 17. A method of cleaning comprising contacting a surface or an item with a cleaning composition set forth in claim 7. 18. The method of claim 16, wherein said subtilisin variant is not a cold water protease variant, wherein said cold water protease variant has one or more of the following characteristics: a) a Test Method 2 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 10, from 1.1 to about 8, or even from 1.1 to about 5; b) a Test Method 3 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 10, from 1.1 to about 8, or even from 1.1 to about 5; c) a Test Method 4 performance index of at least 1.0, at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.0 to about 10, from 1.0 to about 8, or even from 1.0 to about 5; and/or d) a Test Method 6 performance index of at least 1.0, at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.0 to about 10, from 1.0 to about 8, or even from 1.0 to about 5; and/or e) Test Method 7 performance index of at least 1.1, at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 1.6, at least 1.7, at least 1.8, at least 1.9, at least 2, from 1.1 to about 15, from 1.1 to about 10, or even from 1.1 to about 7. 19. The isolated subtilisin variant of claim 1, wherein said variant is selected from T022R-S101G-S103A-V104I-A232V-Q245R;T022R-S024F-S101G-S103A-V104I-A232V-Q245R-E271H;T022R-S101G-S103A-V104I-A232V-Q245R-E271H;T022R-S024F-S101G-S103A-V104I-A232V-Q245R-T260M;T022R-S024F-S101G-S103A-V104I-A232V-Q245R;T022R-S101G-S103A-V104I-A232V-P239G-Q245R;T022R-S101G-S103A-V104I-A232V-Q245R-T260K-L267N;T022R-S101G-S103A-V104I-S128A-A232V-Q245R;T022R-S024F-S101G-S103A-V104I-V121F-A232V-Q245R;T022R-S078N-S101G-S103A-V104I-N116L-A232V-Q245R;T022R-S101G-S103A-V104I-A232V-P239G-Q245R-E271I;T022R-S024G-G097S-S101G-S103A-V104I-Q109G-N116L-S128A-A232V-Q245R;T022R-S024G-S078N-S101G-S103A-V104I-N116L-S128A-A232V-Q245R;T022R-G097S-S101G-S103A-V104I-S128A-A232V-Q245R;T022R-S024G-S078N-G097A-S101G-S103A-V104I-N116L-L217Q-A232V-Q245R;T022R-S024G-S078N-G097S-S101G-S103A-V104I-Q109N-N116L-L217Q-A232V-Q245R;T022R-S024G-S101G-S103A-V104I-N116L-L217Q-A232V-Q245R;T022R-S024G-G097A-S101G-S103A-V104I-Q109G-N116L-L217Q-A232V-Q245R;T022R-G097A-S101G-S103A-V104I-Q109G-N116L-L217Q-A232V-Q245R;T022R-S024G-S078N-G097A-S101G-S103A-V104I-N116L-S128A-L217Q-A232V-Q245R;T022R-S024G-G097A-S101G-S103A-V104I-N116L-S128A-L217Q-A232V-Q245R;T022R-G097S-S101G-S103A-V104I-L217Q-A232V-Q245R;T022R-S078N-G097S-S101G-S103A-V104I-N116L-S128A-L217Q-A232V-Q245R;T022R-S101G-S103A-V104I-Q109N-S128A-A232V-Q245R;T022R-S024G-G097S-S101G-S103A-V104I-S128A-L217Q-A232V-Q245R;T022R-S101G-S103A-V104I-A232V-Q245R-E271A;R010A-T022R-S024F-S101G-S103A-V104I-A232V-Q245R-L267N-E271H;T022R-S101G-S103A-V104I-A232V-Q245R-E271F;T022R-S101G-S103A-V104I-A232V-Q245R-E271H;T022R-S078N-G097S-S101G-S103A-V104I-Q109G-S128A-A232V-Q245R; andT022R-S101G-S103A-V104I-M222S-A232V-Q245R-E271H.
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이 특허에 인용된 특허 (72)
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