US2012149624A1PendingUtilityA1
Subtilase Variants
Est. expiryNov 6, 2022(expired)· nominal 20-yr term from priority
Inventors:Henriette DraborgPeter Kamp HansenMads Eskelund BjornvadMads Norregaard-MadsenMikael Mikkelsen
C11D 3/386C12N 9/54
55
PatentIndex Score
0
Cited by
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References
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Claims
Abstract
The present invention relates to novel subtilase variants exhibiting alterations relative to the parent subtilase in one or more properties including: Wash performance, thermal stability, storage stability or catalytic activity. The variants of the invention are suitable for use in e.g. cleaning or detergent compositions, such as laundry detergent compositions and dish wash compositions, including automatic dish wash compositions.
Claims
exact text as granted — not AI-modified1 . A protease variant comprising:
a) an insertion, substitution or deletion of one of the amino acid residues K,H,R,E,D,Q,N,C,V,L,I,P,M,F,W,Y,G,A,S,T in one or more of the positions 62, 68, 97, 98, 99, 106, 131, 170, 245, 252,
in combination with at least one of the following modifications
*0AQSVPWG; A1T,V; Q2L; S3T,A,L; V4L,A; I8V,T; S9G,D,R,K,L,V; R10H,K; V11A; Q12D; A13V; P14S,T,D,A,M,V,K,Q,L,H,R,I; A15M,T; A16P; H17R; N18S,H; R19W,K,L,F,G,I; G20*,R,A; L21F,LP,LW,LA,LG; T22S,A,K,TV,TG,TL,TW,TV,G,L,TY; G23S; S24P; K27R, V28I; V30I; I35T,V;T38S; P40L; N43D; R45H,K; G46D; A48T; S49N; F50S; V51A,I,D; P52V,A; P55S,A; S57P; G61E,D,S,R,GP; N62D,ND,NE,DE,NG,E,S; V68A,S,L,I; T71A; I72V; L75I; N76S,D; N77S; S78T; V81A; A85T; S87C; A88V,T; E89G; K94N; V95C,T; L96LA,LG; G97E,D,W,A,GG,GA,GV,N,GS; A98S,D,E,T,AS,AD,AV,AE,AH,Q,N,M,L,G,R,V,S; S99D,L,A,AD,SD,SM,SG,DA,P,G,N,C,M,V,I; G100S,GE,C; S101SA, SK; G102D,S; S103D,E,Y,L,Q,H,T; V104T,S,R,I,N,M,L,D; S106D,E,T,M,G,A,L,F,I; I107T,V,M; A108V,T,S; L111I,V; A114V; N116S,D; G118D; M119L,I,V,A,S; H120N,D,Q,K,E,Y,S; V121A; L124C; L126I; G127E; S128N,I,G,C; P129PSN,T,E,D,S,N,A; S130P,T,C,*; P131M,F,W,L,A,H,T,*,PA,S,Q,R,E,G,D,C; S132G,T; A133ASA; T134A; Q137H,E,D; A138G,V; V139L,I; N140D, K; T143A; S144D, N,P; R145G; V150I; A151V,G; A152P; A158T,V,C,E,L,D, M; G160A,D; S163G,C,N,A; Y167K,A,I; A168G; A169G; R170C,S,H,L; Y171C; A172V; N173D; A174V; M175L,I,V,A,S,T; N183D; N184D,S; N185S,D; R186L,C,H; S188G; S190A; Y192H; G195F,E; V203S,A,L,Q,M, F,I; N204T,D,S; Q206L; Y209C,H; G211D; S212N,L; T213A; Y214C,H; A215D,T; N218D,S; M222L,I,V,A,S; A223G; T224A,S; A228T; A230V; A232S,L,T,P; V234I; Q236A,L,D,T,C,M,F,S; K237R; N238D; P239T,S; S240F; S242T; V244I, M,A; Q245R,K,E,D,T,F,N,V,W,G,I,S,C,L,A,M; N248P,D,S; K251E,R; N252G,H,D,V,M,S,T,E,Y,S,Q,K,A,L; A254S; T255A,S; S256N,R,G; L257G; G258K, S259A,N,G; T260A,R; N261D; L262S, Q,V; Y263H,F; G264E; S265G,R,N; V268L,I; N269T; N296K; E271A; T274S,L,A,R, or
b) one of the following combination variants
A108T+L111V; L124I+S125A; P129S+S130AT; L96LA+A151G+V203A; S49N+V203L+N218D; S3T+A16P+R45C+G100S+A230V; I8V+R19K+V139I; N76D+A174AL+A194P+A230V; N185R; N62NE; H120Q+Q137E, G61GE, G61GS, G100L, A133D, V68A, N123D, L111F+Y263H, V11A+G61GE+V227A+S240F, A133E+S144K+N218D, S128A+P129S+S130SP, S9R+A15T+T22TQ+S101P, S9R+A15T+H120R+Q137D+N173S, G97E, Q245W, S9R+A15T+L96LG+Q137E+Y209H, S9R+A15T+L111V+Q137E+G211D, S9R+A15T+L111I+Q137E, S9R+A15T+L111I+H120N+Q137E, S9R+A15T+L96LG+H120Q+Q137E, S9R+A15T+T260M, S9R+A15T, Q245I, S9R+A15T+H120G+Q137E+N218D, S9R+A15T+S130P, Q245F, S9R+A15T+N218D, G63E+N76D+A194P+A230V, S9R+A15T+T224A, G100S, S9R+A15T+D60DG, A138V+V139I+A194P+N218D+A230V, A108V+A169G+R170A+Y171H, I8V+P14L+R19L+V30I+I35V+S57P+P129S+Q137D+S144D+S256N, A133D+T134S+Q137A, Q137D, A98AH, V51D, Q12E+P14L+A15T, G63E+N76D+A194P+A230V, Q12E+P14L+A15T, G97GS, V51A+S163T; V139I+A151G; S9R+A15T+L96LG+S130*; A169G+R170H or
c) one or more modifications in position 68, wherein said modification(s) comprise(s): deletion, insertion and/or substitution of an amino acid residue selected from the group consisting of K,H,R,E,D,Q,N,C,V,L,I,P,M,F,W,Y,G,A,S and T.
2 . A protease variant of claim 1 comprising the combination of one or more of the modifications
X62D,E,S,XD,XE,XG,DE
X68A,S,L,I
X97E,D,W,A,N,XG,XA,XV,XS
X98S,D,E,T,N,M,L,G,R,V,XS,XD,XV
X99D,L,A,P,G,N,C,M,V,I,AD,XD,XM,XG,DA
X106D,E,T,M,G,A,L,F,I
X131M,F,W,L,A,H,T,*,S,Q,R,E,G,D,C,XA
X170C,S,H,L
X245R,K,E,D,T,F,N,V,W,G,I,S,C,L,A,M
X252G,H,D,V,M,S,T,E,Y,S,Q,K,A,L
with at least one of the following modifications
*0AQSVPWG; A1T,V; Q2L; S3T,A,L; V4L,A; I8V,T; S9G,D,R,K,L,V; R10H,K; V11A; Q12D; A13V; P14S,T,D,A,M,V,K,Q,L,H,R,I; A15M,T; A16P; H17R; N18S,H; R19W,K,L,F,G,I; G20*,R,A; L21F,LP,LW,LA,LG; T22S,A,K,TV,TG,TL,TW,TV,G,L,TY; G23S; S24P; K27R, V28I; V30I; I35T,V;T38S; P40L; N43D; R45H,K; G46D; A48T; S49N; F50S; V51A,I,D; P52V,A; P55S,A; S57P; G61E,D,S,R,GP; N62D,ND,NE,DE,NG,E,S; V68A,S,L,I; T71A; I72V; L75I; N76S,D; N77S; S78T; V81A; A85T; S87C; A88V,T; E89G; K94N; V95C,T; L96LA,LG; G97E,D,W,A,GG,GA,GV,N,GS; A98S,D,E,T,AS,AD,AV,AE,AH,Q,N,M,L,G,R,V,S; S99D,L,A,AD,SD,SM,SG,DA,P,G,N,C,M,V,I; G100S,GE,C; S101SA, SK; G102D,S; S103D,E,Y,L,Q,H,T; V104T,S,R,I,N,M,L,D; S106D,E,T,M,G,A,L,F,I; I107T,V,M; A108V,T,S; L111I,V; A114V; N116S,D; G118D; M119L,I,V,A,S; H120N,D,Q,K,E,Y,S; V121A; L124C; L126I; G127E; S128N,I,G,C; P129PSN,T,E,D,S,N,A; S130P,T,C,*; P131M,F,W,L,A,H,T,*,PA,S,Q,R,E,G,D,C; S132G,T; A133ASA; T134A; Q137H,E,D; A138G,V; V139L,I; N140D, K; T143A; S144D, N,P; R145G; V150I; A151V,G; A152P; A158T,V,C,E,L,D, M; G160A,D; S163G,C,N,A; Y167K,A,I; A168G; A169G; R170C,S,H,L; Y171C; A172V; N173D; A174V; M175L,I,V,A,S,T; N183D; N184D,S; N185S,D; R186L,C,H; S188G; S190A; Y192H; G195F,E; V203S,A,L,Q,M, F,I; N204T,D,S; Q206L; Y209C,H; G211D; S212N,L; T213A; Y214C,H; A215D,T; N218D,S; M222L,I,V,A,S; A223G; T224A,S; A228T; A230V; A232S,L,T,P; V234I; Q236A,L,D,T,C,M,F,S; K237R; N238D; P239T,S; S240F; S242T; V244I, M,A; Q245R,K,E,D,T,F,N,V,W,G,I,S,C,L,A,M; N248P,D,S; K251E,R; N252G,H,D,V,M,S,T,E,Y,S,Q,K,A,L; A254S; T255A,S; S256N,R,G; L257G; G258K, S259A,N,G; T260A,R; N261D; L262S, Q,V; Y263H,F; G264E; S265G,R,N; V268L,I; N269T; N296K; E271A; T274S,L,A,R.
3 . The protease variant of claim 1 , comprising one or more of the following alterations:
G97E + A98S
V28I + A98AD + T224S
G97D + A98D
S99AD + M175V + P131F
V95C + G97W + A98E
S99AD + P131L
V95T + G97A + A98D
S9R + S99AD + P131W
S103Y + V104M + S106D
V68A + N116S + V139L + Q245R
V104T + S106D
S3T + A16P + R45C + G100S + A230V
S3T + A16P + S99SD + S144D + A158T +
I8V + S9R + A15T + R19W + V30I + G61D +
A230V + T260R
S99SD + S256N
S103D + V104T + S106T
V30I + S99SD + S256R
S103D + V104L + S106M
G61S + S99SD + V244I
S103D + V104T + S106G
V68A + V139L + S163G + N185S
S103D + V104S + S106A
S99SD + Y263H
S103H + V104N + S106D
V104N + S106T
S103E + V104I + S106T
S99SG + S144D
S103Q + V104T + S106E
V30I + S99SD
S103E + S106T
N18H + S99SD
S103E + V104R + S106A
S9R + T22S + S99SD + K251E
A108T + L111V
A48T + V68A + P131M
L124I + S125A
A15M + S99SM + V139I + V244I
L124C + P131*
P14T + A15M + S99SD
P129S + S130AT
I8V + S99SD + S144D + A228T
L96LA + A151G + V203A
I8V + R19K + V139I
S99SD + A108V + V139L
I35T + N62D
S99SD + S190A
N62D + S265G
S99SD + V203A
Q2L + N62D
S99SD + V139I
N62D + N76D
S99SD + A108V
R45H + G61E + V68A
S99SD + S106A + A151G
N62D + V121A
V68A + S106A
N62D + A215D
V68A + N185D + V203S
N62D + N238D
V68A + V139L
N62D + R145G
V68A + V139I
V4L + N62D + E89G
V68A + A158V
N62D + S188G + K251R
V68A + V203A
S49N + N62D
V68A + V203S
N62NE
V68A + V203L + S259A
V11A + N62DE
V68A + S106L
N62ND + N184S + S256G
V30I + V68A + V203S
N18S + N62D + I107T + A254S
V51A + V68A + S106T + A168G
S57P + N62ND
V51A + V68A + S106T + A168G
N62NE + V234I
V68A + N76S + V203M + P239T
Q137H + R170C + G195E
V68A + V203L
S99A + S101SA
V68A + L75I + V203Q
R10K + P14A + R19K + A98AS + S128N
V68A + T71A + V139L
T22A + R45K + A98AS + S128N
Y192H + V68A
A98AV + S99D + Y167K
V68A + S106A + A108T
S9G + P14K + Y167A + R170S
V68A + S106T + A108T
S9D + P14T + Y167A + R170S
V68S + A108S
S9R + P14M + A98AD
V68A + N76S + G211D
S9R + R19L + A98AD + E271A
V68A + S106T + A108T
S9R + P14S + R19F + A98AD
A151V + R170C
S99DA + P129PSN + P131A
P14D + A98AS + H120D + G195F +
S99AD + V244M + Q245K + N248D +
S212N + M222S
K251R + T255A + S256N
S49N + V203L + N218D
S9R + P14V + R19G + A98AD
V68A + S106M + N184D
S99AD + N248P + T255A + S256G
P55S + V68L + A158E + G160A
*0AQSVPWG + A98AD
V68A + A158C
T22A + S99AD
V68A + A158L + Y214C
K94N + A98T + S99L
A88V + S99AD + P131F
N76D + A174AL + A194P + A230V
P14T + A16P + I72V + S99SD + V244I + T260A
P40L + N218D + A232S + Q236L + Q245E + S259N
S99AD + P131F
A232L + Q236D + Q245E
R10H + N62D
A232T + Q236L + Q245D
V28I + A98AD + T224S
R170H + Q236A + Q245R
S9K + T22K + S99AD
A232L + Q236T + Q245D
P14S + S99AD + P131W
G97GG + P131H + Q137E + V268L
V68A + I72V + P131F
A88V + G97GV + P131H
S9R + S99AD
G97GA + H120Q + S130P + G264E
S9K + S99AD
G97GG + V139L
V28I + A88V + G100S + P131M
G97GG + Q137D
S103L + V104S + S106G
G97GG + H120D + Q137H
V68A + T224A
N185R
V68A + P131F
P131H + Q137E
A48T + V68A + P131M
V104I + H120N + P131H + Q137E
V68A + I72V + P131F
H120Q + Q137E
G100GE + P131F
S9R + A15T + G97GV + H120D
S99AD + P131F + T260A
G100S + H120Q + Q137H
R19G + A98AS
V68A + H120K + Q137E
G61R + N62D
G97GA + H120E
V68A + S106M + N184D
H120D + S128I + Q137D
P55S + V68L + A158E + G160A
G97GG + P131H
V68A + A158C
G97GG + H120N + L126I
R19W + G61S + S99SD + N204T + Y263H +
S9R + A15T + G97GA + H120D + P131H +
S265R
Q137E
A232T + Q236C
S9R + A15T + G97GV + P131T + Q137H
N62D + A232T + Q236C
S9R + A15T + G20* + L21F + N62D + Q245N
A232P + Q236L + Q245E
S9L + A15T + T22TV + V139L + Q245F
A232S + Q236L + Q245T + K251E
S132G + Q245F
S163C + Q236M + Q245T + S256G
S9R + A15T + T22TG + N62D + V139L + Q245V
N218D + A232L + Q236F + Q245F
S9L + A15T + T22TV + V139L + Q245F + L262S
S163N + A232L + Q236S + Q245E
S9R + A15T + T22TL + N62D + Q245W
A232S + Q236S + Q245E
V68A + A158L + Y214C
V68A + V203L
N62D + V150I
V68S + A158D
S3T + P14Q + A15M + R19K + N62D + S144D
I8V + A15T + R19K + A85T + S99SD + A114V +
P14Q + R19W + V51I + G61E + S99SD +
V244I + S256N + Y263H
V139I + T260R
L111F + Y263H
S3T + P14L + H17R + S99SD + V139I + S144D
P52V + S78T + S99SD
S3A + V30I + S99SD + S106G + N248S
A15M + S99SD + V268I
I8V + A15T + S99SD
S99G + S128N + N183D + A232L + Q236T +
S3T + S9R + P14H + A15M + R19L + S99SD +
Q245R
V139I
S99R + S101SA
S9R + A15T + G97GG + H120D + Q137E
L96LA + A98T + P131AA
S9R + A15T + G20A + G97GV + H120D + P131H
A98E + S99P
S163N + A232L + Q236A + Q245G
V28I + S99AD + P131F
N173D + A232L + Q236A + Q245N
S9R + A15T + G97GV + Q137H
P55S + V68A + S106M + A108T + P129T
V81A + P131T + A133S + Q137E
K27R + V68L + G118D + A158E
N43D + V68A + S106F + N238D
A98E + S99A + S101SK
V68A + V203F
V68A + N140D + T143A + S144N
V68A + S106E
N62D + N140K + T143A + S144D
V68A + S106I
S9F + P14T + R19L + A98AD
V68A + A158M + R170C
S9V + P14R + R19F + A98AD
V68A + P129T + N218D
S99A + S99SD + G258K + L262Q
V68S + P129E
S87C + S99SA + S99D + P131A
V68S + P129D
S99A + S99SD + G258K + L262Q
V68L + P129E + N261D
V28I + S99A + *99aD + P131F
G97GV + H120D
A85T + G102D + S106T + K237R
P131A + A133ASA
V68A + T71A
L111F + Y263H
G61GS
V11A + G61GE + V227A + S240F
G100L
A133E + S144K + N218D
A133D
S128A + P129S + S130SP
V68A
G61GE
N123D
S9R + A15T + T22TW + N204D + Q245I
Q245W + N252V
S9R + A15T + G97GG + P131S + Q137H
R45H + Y171C + Q245W + N252S
S9R + A15T + T22TG + N62D + V139L + Q245G
G20R + A48T + R170C + Q245W + N252Q
S9R + A15T + T22TL + N62D + I107V + V139L +
S9R + A15T + A16P + G97GA + P131S + Q137D +
Q245W
N204S
S9G + A15T + G97GA + Q137H
N218D + Q245W + N252E
S9R + A15T + V68A + Q245R
G20R + R170C + Q245R + N252V
S9R + A15T + G97GA + H120N + S212L
S9R + P14I + R19K + A98AD + T274S
S9R + A15T + L96LG + H120D + P131H + R186L
A98AE + V203I
S9R + A15T + G97GA + H120D + Q137D
V51A + V68A + S163G + V203A
N62D + N252T
N62D + Q245W + N252H
V4A + S9R + A15T + G97GV + H120D
N62D + Q245W + N252A
S9R + A15T + G97GV + H120D + Q137H
G20R + N62D + V244I + Q245W + N252E
S9R + A15T + L96LG + H120N + P131H + Q137E
N204D + Q245S
S9R + A15T + L96LG + H120D + P131S + Q137E
N62D + Q245W + N252E
S9R + A15T + H120N + P131T + N218D
N62D + Q245R + N252V
S9R + A15T + L21LP + T22TV + M119I + N218D +
S9R + A15T + S24P + G61E + A85T + P239S +
Q245I
Q245A
S9R + A15T + L96LG + H120D + G160D
G102S + M222S + Q245L + N252D
V68A + S106A + G118D + Q245R + T255S +
A15M + V30I + N62D + S99N + L111I + V244A +
L257G + T274L
S265N
S9R + A15T + G61E + A85T + P239L + Q245C
S9R + A15T + T22TG + N62D + V139L + Q245S
S9R + A15T + P131H + S144P
S3T + Q12D + R19W + V30I + S106G + I107M
S9R + A15T + G97GA + Q137E
V68A + A88T + V139L
S9R + A15T + G97GA + H120Q + P131H + Q137E
V51I + L111I + G118D + Q245R
S9R + A15T + L21LW + G100S + V139L + Q245V
V68A + V203L
S9R + A15T + G97GA + Q137H + N218S
A1T + V68A + N116D + G118D
S9R + A15T + L96LG + H120N + P131S + Q137H
V68A + G118D + Q245R
S9R + A15T + G97GA + H120N + Q137E
N62D + V139I + N183D + N185S + V203I +
Q245R + L262S
S9R + A15T + L96LG + P131T + Q137H
N62D + I72V
S9R + A15T + L96LG + H120N + P131S
N62D + V81A + Q245R
S9R + A15T + V68A + Q137D
T22A + V68A + S106T + G118D
S9R + A15T + G97GA + H120Y + Q137H
V68A + L111I + V203I
S9R + A15T + G97GA + Q137D
G61E + V68A + A169G
S9R + A15T + K94N + H120N + P131H
V68A + L111V
S9R + A15T + L96LG + P131H + Q137D
V68A + G118D + V203A + K251R
S9R + A15T + F50S + H120D + P131H
V68A + G118D
S9R + A15T + G97GA + H120N + Q137D + N248D
A1V + V51A + V68A + V203I
S9R + A15T + L96LG + P131Q + Q137D
V68A + V139L + A223G
S9R + A15T + T22G + V139L + Q245L
N62D + Y214H + K237R
V139L + Q245R
V68A + S106A + G118D + Q245R
S9R + A15T + Q245F
S9R + A15T + T22A + N62D
S9R + A15T + Q245S
A98Q + S99D
S9R + A15T + G97GV + H120Q
S9R + P14I + R19K + A98AD
S9R + A15T + G97GA + Q137E + L262V
S9R + A15M + A16P + T22S + S99AD
S9R + A15T + G127E + P131R + Q137H
S99AD + T255R + S256N
S9R + A13V + A15T + I35V + N62D + Q245F
S9R + A15T + T22TQ + S101P
S9R + A15T + Q245V
S9R + A15T + H120R + Q137D + N173S
V139L + Q245F
G97E
S9R + A15T + T22A + V139L + Q245E
Q245W
S9R + A15T + T22L + V139L + Q245V + A254S
S9R + A15T + L96LG + Q137E + Y209H
S9R + R19L + A98AD
S9R + A15T + L111V + Q137E + G211D
P14R + A98AD
S9R + A15T + L111I + Q137E
S9R + A15T + Q245L
S9R + A15T + L111I + H120N + Q137E
S9R + A15T + G61E + A85T + P239S + Q245V
S9R + A15T + L96LG + H120Q + Q137E
S9R + A15T + G61E + A85T + Q206L + Q245R
S9R + A15T + T260M
P239T + Q245R
S9R + A15T
S9R + A15T + N62NG + Q245T
Q245I
S9R + A15T + G61GP + Q245L
S9R + A15T + H120G + Q137E + N218D
S9R + A15T + G61E + A85T + Q137H + Y209C +
I8V + P14L + R19L + V30I + I35V + S57P +
Q245G
P129S + Q137D + S144D + S256N
S9R + A15T + G61E + A85T + P239S + Q245C
Q245F
V68I + A98AD
S9R + A15T + N218D
V68A + N269K
G63E + N76D + A194P + A230V
N62D + Q245A + N252G + S265G
S9R + A15T + T224A
N218D + Q245G + N252H
G100S
S9R + A15T + G102S + M175T + Q245R + N252D
S9R + A15T + D60DG
S9R + A15T + N62D + Q245W + N252V
A138V + V139I + A194P + N218D + A230V
S9R + A15T + N62D + Q245R + N252M
A108V + A169G + R170A + Y171H
S9R + A15T + N62D + Q245W + N252S
S9R + A15T + S130P
S99SD + N204S + Q245R
A133D + T134S + Q137A
N62D + Q245R
Q137D
N62D + A151G
A98AH
V68A + S106T
V51D
S99A + S99SD + V203L
Q12E + P14L + A15T
A98AD + A215T
G63E + N76D + A194P + A230V
N62D + Q245G + N252T
Q12E + P14L + A15T
A152P + Q245R + N252T
G97GS
S163N + T213A + Q245R
Q245W + N252Y
S106L + Q245R + N252E
A169G + R170H
S9V + P14R + R19F + A98AD
Q12E + P14L + A15T
S9R + A15T + L111I + Q137E
P14R + A98AD
S9R + A15T + G97GA + Q137E
G100S
S9R + A15T + L96LG + Q137E + Y209H
A169G + R170H
S9R + A15T + L96LG + H120N + P131S
A98AD + A169G
S9R + A15T + G97GV + H120Q
A138V + V139I + A194P + N218D + A230V
S9R + A15T + L96LG + H120Q + Q137E
S99A + S99SD + V203L
S9R + A15T + G97GV + P131S
V68A + S106T
S9R + A15T + K94N + H120N + P131H
A98AD + A215T
S9R + A15T + N76S + L111V + P131H + Q137D
A108V + A169G + R170A + Y171H
S9R + A15T + F50S + H120D + P131H
S3L + N62D + S163A + S190A
S9R + A15T + L96LG + S130*
S9R + P14I + R19K + A98AD + T274S
S9R + A15T + T22TL + N62D + I107V + V139L +
S9R + A15T + G61E + A85T + N218D + P239S +
Q245W
Q245L
S9R + A15T + G97GA + H120D + Q137H +
S9R + A15T + S24P + G61E + A85T + P239S +
M222V
Q245A
S9R + A15T + G97GA + H120N + Q137D + N248D
S99SD + P131F
S9R + A15T + L21LW + G100S + V139L + Q245V
N62D + P131F + A172V
S9R + A15T + G20* + L21F + N62D + Q245N
N62D + P131F
S9R + A15T + L21LC + V139L + R186H + Q245M
V68A + A88T + V139L
S132G + Q245F
V68A + G118D + V203A
S9R + A15T + T22TG + N62D + V139L + Q245G
P40L + V68A + A108T + A138V + V203I
S9R + A15T + L96LG + P131Q + Q137D
I8T + A98AD + T274R
S9R + A15T + T22TQ + S101P
A98AE + V203I
S9R + A15T + T22TG + N62D + V139L + Q245V
V51A + V68A + S163G + V203A
S9R + A15T + T22TL + N62D + Q245W
A1V + V51A + V68A + V203I
S9R + A15T + T22TW + N204D + Q245I
V68A + G100S
S9R + A15T + T22TG + N62D + V139L + Q245S
V68A + V203L
S9R + A15T + S130P
A1T + V68A + N116D + G118D
Q245W
N62D + A169G + V203I + Q245R
S9R + A15T + L21LP + T22TY + V139L + G160D +
G23S + S99SD + A194P + S242T + Q245R +
Q245L
T274R
S9R + A15T + G61E + A85T + P239L + Q245C
S99SD + N204S + Q245R
S9R + A15T + L21LP + T22TV + M119I + N218D +
N62D + V139I + N183D + N185S + V203I +
Q245I
Q245R + L262S
S9R + A15T + V68A + Q245R
V68A + S106A + G118D + Q245R
S9R + A15T + T22A + V139L + Q245E
V51I + L111I + G118D + Q245R
V139L + Q245R
N62D + Q245R
S9R + A15T + Q245F
N62D + I72V
S9R + A15T + Q245S
S9R + R19L + A98AD
S9R + A15T + T260M
S9G + P14R + R19I + A98AD
S9R + A15T
S9R + A15T + T22L + V139L + Q245V + A254S
S9R + A15T + L21LG + T22TV + V139L + N204D +
S99G + S128N + N183D + A232L + Q236T +
Q245N
Q245R
V139L + Q245F
S9R + A15T + Q245L
S9R + A15T + T22G + V139L + Q245L
S9R + A15T + N62NG + Q245T
S9R + A15T + Q245V
S9R + A15T + N62ND + V139L + Q245E
Q245F
S9R + A15T + N62ND + V139L + N261D
S9R + Q245C
Y167I + R170L + Q245E
S9R + A15T + N218D
Y167I + R170L + Q245R
S9R + A13V + A15T + I35V + N62D + Q245F
Y167I + R170L + Q245M
S9R + A15T + T224A
Y167I + R170L
S163N + A232L + Q236A + Q245G
S99SE + Q245R
S9R + A15T + A16P + G97GA + P131S + Q137D +
S9R + A15T + G61E + A85T + Q137H + Y209C +
N204S
Q245G
N218D + A232L + Q236F + Q245F
S9R + A15T + G61E + A85T + P239S + Q245C
S163N + A232L + Q236S + Q245E
G102S + M222S + Q245L + N252D
G97GA + H120E
N62D + Q245A + N252G + S265G
G97GG + P131H
N62D + Q245G + N252T
S9R + A15T + G97GA + H120D + P131H + Q137E
S9R + A15T + N62D + Q245W + N252V
S9R + A15T + G97GV + Q137H
S9R + A15T + N62D + Q245R + N252M
S9R + A15T + G97GV + H120N
S9R + A15T + N62D + Q245W + N252S
S9R + A15T + G97GG + P131S + Q137H
S163N + T213A + Q245R
S9R + A15T + G97GG + H120N + Q137D
S106L + Q245R + N252E
S9R + A15T + H120Q + P131C + Q137H
Q245W + N252Y
S9R + A15T + G97GV + H120D + Q137H
Q245W + N252V
S163C + Q236M + Q245T + S256G
G20R + A48T + R170C + Q245W + N252Q
S9R + A15T + G97GG + H120D + P131H +
N62D + N252T
Q137H
S9R + A15T + G97GV + H120E + Q137H
N218D + Q245W + N252E
S9R + A15T + G97GV + P131T + Q137H
G20R + R170C + Q245R + N252V
S9R + A15T + G97GV + H120Q + Y263F
N62D + Q245W + N252H
S9R + A15T + G97GV + S106A + P131H
N62D + Q245W + N252A
S9R + A15T + G97GG + L111I + P131T + Q137H
G20R + N62D + V244I + Q245W + N252E
S9R + A15T + G97GV + P131H + Q137H
N204D + Q245S
S9R + A15T + G20A + G97GV + H120D + P131H
N62D + Q245W + N252E
S9R + A15T + G97GA + H120D + P131S + Q137E
N62D + Q245R + N252V
S9G + A15T + G97GA + Q137H
A98L + S99C + Q245R
S9R + A15T + H120R + Q137D + N173S
N62D + A98R + Q245R
S9R + A15T + L96LG + H120N + P131H + Q137E
S9R + A15T + V68A + S99G + Q245R + N261D
S9R + A15T + L96LG + H120D + P131S + Q137E
S9R + A15T + G20* + L21F + N62D + Q245R
S9R + A15T + H120N + P131T + N218D
S9R + A15T + G20* + L21F + N62E + Q245R
S9R + A15T + G97GA + H120D + Q137D
V68I + A98AD
S9R + A15T + L96LG + H120D + P131H + R186L
S9R + A15T + H120D + Q137D
S9R + A15T + G97GA + R186C
S9R + A15T + N77S + L96LG + H120D + P131Q
V4A + S9R + A15T + G97GV + H120D
S9R + A15T + G97GA + H120N + Q137E
S9R + A15T + L96LG + H120D + G160D
S9R + A15T + G97GA + Q137E + L262V
S9R + A15T + G97GA + H120N + S212L
S9R + A15T + P131H + S144P
S9R + A15T + G97GA + Q137H + N218S
S9R + A15T + G127E + P131R + Q137H
M222S + Q245G + N252G
S9R + A15T + V68A + S99G + Q245R + N261D
V68I + V203L
N62D + P131F + A172V
V51A + S163T
N62D + P131F
S106A + A138G
S99SD + Q245R
V139I + A151G
S9R + A13T + S99A + S99SD + P131F
A98R + G100C + Q245R
S9R + A15T + N62S + H120N + P131T + N218D
S9R + A15T + S99G + G100S + H120N + P131S +
S9R + A15T + S99C + H120N + P131S + Q137H +
Q137H
M222S
A15T + N185D + M222S + Q245R + N252V
A98G + S99C + Q245R
S9R + A15T + T22TL + G61E + L96LG + Q137D +
A98T + S99G + G100S + S240F + Q245R
Q245R
S9R + T22TL + G61E + G97GG + M119I + P131T
S9R + A15T + H120N + P131T + N218D + N269T
Y209H + M222S + Q245G + N252L
M222S + Q245M + N252E
S9R + A15T + G61E + H120S + Q137D + V139L +
S9R + A15T + L96LG + H120N + P131S + Q137H +
N218D
M222S
S9R + A15T + N62D + H120N + P131T
S9R + A15T + G61E + A98S + S99M + Q245R
S9R + A15T + V68A + N218D + Q245R
A98G + G100S + Q245R + N261D
S9R + A15T + V68A + H120N + N218D + Q245R
S9R + A15T + V68A + A98L + Q245R
S9R + A15T + V68A + A174V + Q245R
S9R + A15T + V68A + A98G + S99V + Q245R
S9R + A15T + G46D + V68A + N218D + Q245R
S9R + A15T + V68A + A98M + Q245R + N248D
G97D + A98N + S128G + S130T + P131D + T134A
S9R + A15T + G61E + V68A + A98S + S99G +
Q245R
S9R + A15T + V68A + A98M + S99G + Q245R +
S9R + A15T + A88V + A98R + S99G + G100C +
T274A
H120N + P131S + Q137H
S9R + A15T + V68A + A98L + S99G + Q245R
A98V + S99C + Q245R
S9R + A15T + A98G + S99C + H120N + P131S +
S9R + A15T + G20* + L21F + G61E + *61aP +
Q137H
Q245R
S9R + A15T + T38S + A98R + S99C + G100S +
S9R + A15T + V68A + A98G + S99I + K237R +
H120N + P131S + Q137H
Q245R
S9R + A15T + A98C + G100S + H120N + P131S +
S9R + A15T + V68A + H120N + P131S + Q137H +
Q137H
Q245R
S9R + A15T + A98S + G100S + H120N + P131S +
S9R + A15T + V68A + H120D + P131S + Q137H +
Q137H
Q245R
S9R + A15T + G20* + L21F + N62D + Q245R
A98S + S99G + G100S + Q245R
S9R + A15T + G20* + L21F + N62D + Q245R +
S9R + A15T + A98S + S99G + G100S + H120N +
S259G
P131S + Q137H
A98S + G100S + Q245R
A98T + S99G + G100S + Q245R
S9R + A15T + G20* + L21F + *61aA + V68A +
S9R + A15T + G20* + L21F + P52T + N62D +
Q245R
Q245R
S9R + A15T + G20* + L21F + N62E + Q245R
A98L + S99C + Q245R
V68A + S105G + S106A
V68A + S106A + T213A
S9R + A15T + Y167I + R170L
S9R + A15T + V68A
V68A + S106A + N252M + Y263C
V68A + S106A + Q245W
V68A + S106A + Q245R + N252D
V68A + S106A + Q245W + N252K
V68A + S106A + A174V + Q245R + N252D
S9R + A15T + V68A + Q245R + N252S
S9R + A15T + V28I + V68A + Q245R + N252A
S9R + A15T + V68A + A194T + Q245R + N252E
S9R + A15T + G20* + L21F + *63aG + Q245R +
S9R + A15T + G20* + L21F + *62aS + N218D +
N272V
Q245R
S9R + A15T + G20* + L21F + *61aS + V68A +
S9R + A15T + V68A + H120N + P131S + Q137H +
G160D + Q245R
Q245M
wherein
(a) the variant has protease activity, and
(b) each position corresponds to a position of the amino acid sequence of subtilisin BPN′, shown in SEQ ID NO: 1.
4 . The protease variant of claim 1 , wherein the parent subtilase belongs to the sub-group I-S1.
5 . The protease variant of claim 1 , wherein the parent subtilase belongs to the sub-group 1-S2, and wherein the parent subtilase preferably is subtilisin 309.
6 . The protease variant of claim 1 , wherein said variant further comprises one or more of the modifications K27R, *36D, S56P, N62D, V68A, N76D, S87N, G97N, S99SE, S101G, S103A, V104A, V104I, V104N, V104Y, S106A, H120D, H120N,N123S, G159D, Y167A, R170S, R170L, A194P, N204D, V205I, Q206E, L217D, N218S, N218D, M222S, M222A, T224S, A232V, K235L, Q236H, Q245R, N248D, N252K, T274A, S101G+V104N, S87N+S101G+V104N, K27R+V104Y+N123S+T274A, N76D+S103A+V104I, S99D+S101R+S103A+V104I+G160S, S3T+V4I+S99D+S101R+S103A+V104I+G160S+V199M+V205I+L217D, S3T+V4I+S99D+S101R+S103A+V104I+G160S+A194P+V199M+V205I+L217D, S3T+V4I+S99D+S101R+S103A+V104I+G160S+V205I and N76D+V104A.
7 . The protease variant of claim 1 , comprising the following substitutions: S101G+S103A+V104I+G159D+A232V+Q236H+Q245R+N248D+N252K.
8 . A cleaning or detergent composition, preferably a laundry or dish wash composition, comprising a protease variant of claim 1 and a surfactant.
9 . A composition of claim 8 , which additionally comprises one or more of an amylase, cellulase, cutinase, esterase, beta-galactosidase, glycoamylase, hemicellulase, lactase, ligninase, lipase, polygalacturonase, and protease.
10 . An isolated DNA sequence encoding a protease variant of claim 1
11 . An expression vector comprising the isolated DNA sequence of claim 10 .
12 . A microbial host cell transformed with the expression vector of claim 11 .
13 . A microbial host cell of claim 12 , which is a bacterium, preferably a Bacillus , especially a B. lentus.
14 . A microbial host cell of claim 12 , which is a fungus or yeast, preferably a filamentous fungus, especially an Aspergillus.
15 . A method for producing a protease variant, comprising
(a) culturing a host of claim 12 under conditions conducive to the expression and secretion of the variant, and (b) recovering the protease variant.Cited by (0)
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