RU2012116529A - Варианты субтилаз - Google Patents
Варианты субтилаз Download PDFInfo
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- RU2012116529A RU2012116529A RU2012116529/10A RU2012116529A RU2012116529A RU 2012116529 A RU2012116529 A RU 2012116529A RU 2012116529/10 A RU2012116529/10 A RU 2012116529/10A RU 2012116529 A RU2012116529 A RU 2012116529A RU 2012116529 A RU2012116529 A RU 2012116529A
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- Prior art keywords
- residue
- replacing
- residues
- subtilisin
- replacement
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- 101710135785 Subtilisin-like protease Proteins 0.000 title 1
- 108090000787 Subtilisin Proteins 0.000 claims abstract 13
- 238000006467 substitution reaction Methods 0.000 claims abstract 8
- 238000005406 washing Methods 0.000 claims abstract 7
- 102000039446 nucleic acids Human genes 0.000 claims abstract 6
- 108020004707 nucleic acids Proteins 0.000 claims abstract 6
- 150000007523 nucleic acids Chemical class 0.000 claims abstract 6
- 229920001184 polypeptide Polymers 0.000 claims abstract 5
- 102000004196 processed proteins & peptides Human genes 0.000 claims abstract 5
- 108090000765 processed proteins & peptides Proteins 0.000 claims abstract 5
- 238000000034 method Methods 0.000 claims abstract 4
- 238000012986 modification Methods 0.000 claims abstract 4
- 230000004048 modification Effects 0.000 claims abstract 4
- 102000040430 polynucleotide Human genes 0.000 claims abstract 4
- 108091033319 polynucleotide Proteins 0.000 claims abstract 4
- 239000002157 polynucleotide Substances 0.000 claims abstract 4
- 102220036452 rs137882485 Human genes 0.000 claims abstract 4
- 102200029981 rs28936700 Human genes 0.000 claims abstract 4
- 102220099575 rs878853725 Human genes 0.000 claims abstract 4
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract 3
- 230000003197 catalytic effect Effects 0.000 claims abstract 2
- 238000004140 cleaning Methods 0.000 claims abstract 2
- 238000004851 dishwashing Methods 0.000 claims abstract 2
- 239000013604 expression vector Substances 0.000 claims abstract 2
- 102220225954 rs1064795276 Human genes 0.000 claims abstract 2
- 102200118280 rs33918343 Human genes 0.000 claims abstract 2
- 102200034374 rs6092 Human genes 0.000 claims abstract 2
- 102220289974 rs757282628 Human genes 0.000 claims abstract 2
- 102220120384 rs886042566 Human genes 0.000 claims abstract 2
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 claims 1
- 238000003780 insertion Methods 0.000 claims 1
- 230000037431 insertion Effects 0.000 claims 1
- 238000010412 laundry washing Methods 0.000 abstract 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
- C12N15/52—Genes encoding for enzymes or proenzymes
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
- C11D3/00—Other compounding ingredients of detergent compositions covered in group C11D1/00
- C11D3/16—Organic compounds
- C11D3/38—Products with no well-defined composition, e.g. natural products
- C11D3/386—Preparations containing enzymes, e.g. protease or amylase
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/52—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from bacteria or Archaea
- C12N9/54—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from bacteria or Archaea bacteria being Bacillus
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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Wood Science & Technology (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Molecular Biology (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Plant Pathology (AREA)
- Detergent Compositions (AREA)
- Enzymes And Modification Thereof (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
1. Вариант исходного субтилизина, включающий замены 9{R,K,H}, 15{G,A,S,T,M}, 68{G,A,S,T,M}, 218{D,S,G,V} и 245{R,K,H}, где данный вариант дополнительно включает, по меньшей мере, одну из следующих модификаций: 61{D,E}, 62{D,E}, 76{D,E}, *97aG, 98{G,S}, 99G, 101G, 120{V,Q,D}, 131{T,S}, 137H, 194P, 228V, 230V, 261D, где указанные положения соответствуют положениям в последовательности зрелого полипептида SEQ ID NO:2 [BPN'].2. Вариант по п.1, где указанный вариант включает замену G61E.3. Вариант по п.1, где указанный вариант включает замену A98S.4. Вариант по п.1, где указанный вариант включает замену S99G.5. Вариант по п.1, где указанный вариант включает следующие замены S9R, A15T, G61E, V68A, A98S, S99G, N218D и Q245R.5. Вариант по п.5, где исходный субтилизин является полипептидом, включающим аминокислотную последовательность, имеющую, по меньшей мере, 80% идентичность с последовательностью SEQ ID NO:1.7. Вариант по п.1, где исходный субтилизин принадлежит к подгруппе I-S2.8. Вариант по п.1, где указанный вариант обладает одним или несколькими улучшенными свойствами по сравнению с исходным субтилизином, где улучшенные свойства включают моющую способность, стабильность, каталитическую активность и эффективность мытья посуды.9. Выделенный полинуклеотид, кодирующий вариант по любому из пп.1-8.10. Конструкт нуклеиновой кислоты, включающий полинуклеотид по п.9.11. Вектор экспрессии, включающий конструкт нуклеиновой кислоты по п.11.12. Клетка-хозяин, содержащая конструкт нуклеиновой кислоты по п.11.13. Чистящая или моющая композиция, предпочтительно композиция для стирки или мытья посуды, содержащая вариант по любому из пп.1-8.14. Способ получения варианта по п.1 путем введения следующих замен в исходный субтилизин:i. замену остатка в положении 9 остатками {R
Claims (16)
1. Вариант исходного субтилизина, включающий замены 9{R,K,H}, 15{G,A,S,T,M}, 68{G,A,S,T,M}, 218{D,S,G,V} и 245{R,K,H}, где данный вариант дополнительно включает, по меньшей мере, одну из следующих модификаций: 61{D,E}, 62{D,E}, 76{D,E}, *97aG, 98{G,S}, 99G, 101G, 120{V,Q,D}, 131{T,S}, 137H, 194P, 228V, 230V, 261D, где указанные положения соответствуют положениям в последовательности зрелого полипептида SEQ ID NO:2 [BPN'].
2. Вариант по п.1, где указанный вариант включает замену G61E.
3. Вариант по п.1, где указанный вариант включает замену A98S.
4. Вариант по п.1, где указанный вариант включает замену S99G.
5. Вариант по п.1, где указанный вариант включает следующие замены S9R, A15T, G61E, V68A, A98S, S99G, N218D и Q245R.
5. Вариант по п.5, где исходный субтилизин является полипептидом, включающим аминокислотную последовательность, имеющую, по меньшей мере, 80% идентичность с последовательностью SEQ ID NO:1.
7. Вариант по п.1, где исходный субтилизин принадлежит к подгруппе I-S2.
8. Вариант по п.1, где указанный вариант обладает одним или несколькими улучшенными свойствами по сравнению с исходным субтилизином, где улучшенные свойства включают моющую способность, стабильность, каталитическую активность и эффективность мытья посуды.
9. Выделенный полинуклеотид, кодирующий вариант по любому из пп.1-8.
10. Конструкт нуклеиновой кислоты, включающий полинуклеотид по п.9.
11. Вектор экспрессии, включающий конструкт нуклеиновой кислоты по п.11.
12. Клетка-хозяин, содержащая конструкт нуклеиновой кислоты по п.11.
13. Чистящая или моющая композиция, предпочтительно композиция для стирки или мытья посуды, содержащая вариант по любому из пп.1-8.
14. Способ получения варианта по п.1 путем введения следующих замен в исходный субтилизин:
i. замену остатка в положении 9 остатками {R, K, H};
ii. замену остатка в положении 15 остатками {G, A, S, T, M};
iii. замену остатка в положении 68 остатками {G, A, S, T, M};
iv. замену остатка в положении 245 остатками {R, K, H}; и
v. замену остатка в положении 218 остатками {D, S, G или V},
и одной или нескольких из следующих модификаций: замену остатка в положении 61 остатками {D, E}, замену остатка в положении 62 остатками {D, E}, замену остатка в положении 76 остатками {D, E}, включение остатка G в положение 97, замену остатка в положении 98 остатками {G, S}, замену остатка в положении 99 остатком G, замену остатка в положении 101 остатком G, замену остатка в положении 120 остатками {V, Q, D}, замену остатка в положении 131 остатками {T, S}, замену остатка в положении 137 остатком H, замену остатка в положении 194 остатком P, замену остатка в положении 228 остатком V, замену остатка в положении 230 остатком V, замену остатка в положении 261 остатком D, где указанные положения соответствуют положениям в последовательности зрелого полипептида SEQ ID NO:2 [BPN'].
15. Способ по п.14, где в исходном субтилизине осуществляют следующие замены:
i. замену остатка S в положении 9 остатком R;
ii. замену остатка A в положении 15 остатком T;
iii. замену остатка V в положении 68 остатком A;
iv. замену остатка Q в положении 245 остатком R;
v. замену остатка N в положении 218 остатками {D, S, G или V}
и одну или несколько из следующих модификаций: замену остатка G в положении 61 остатком E, замену остатка N в положении 62 остатком D, замену остатка N в положении 76 остатком D, вставку остатка G в положение 97, замену остатка A в положении 98 остатком S, замену остатка S в положении 99 остатком G, замену остатка S в положении 101 остатком G, замену остатка H в положении 120 остатком D, замену остатка P в положении 131 остатком T, замену остатка Q в положении 137 остатком H, замену остатка A в положении 194 остатком P, замену остатка A в положении 228 остатком V, замену остатка A в положении 230 остатком V, замену остатка N в положении 261 остатком D.
16. Способ по п.14 или 15, где исходный субтилизин принадлежит к подгруппе I-S2, предпочтительно, где исходный субтилизин включает аминокислотную последовательность, имеющую, по меньшей мере, 80% идентичность с последовательностью SEQ ID NO:1.
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP09171308.1 | 2009-09-25 | ||
EP09171308 | 2009-09-25 | ||
PCT/EP2010/064171 WO2011036263A1 (en) | 2009-09-25 | 2010-09-24 | Subtilase variants |
Publications (2)
Publication Number | Publication Date |
---|---|
RU2012116529A true RU2012116529A (ru) | 2013-10-27 |
RU2651525C2 RU2651525C2 (ru) | 2018-04-19 |
Family
ID=43033187
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
RU2012116529A RU2651525C2 (ru) | 2009-09-25 | 2010-09-24 | Варианты субтилаз |
Country Status (10)
Country | Link |
---|---|
US (1) | US20120172280A1 (ru) |
EP (1) | EP2480650B1 (ru) |
JP (2) | JP5947213B2 (ru) |
CN (1) | CN102648273B (ru) |
AU (1) | AU2010299799B2 (ru) |
BR (1) | BR112012006487A8 (ru) |
CA (1) | CA2775045A1 (ru) |
MX (1) | MX339581B (ru) |
RU (1) | RU2651525C2 (ru) |
WO (1) | WO2011036263A1 (ru) |
Families Citing this family (273)
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BR112012006487A8 (pt) * | 2009-09-25 | 2018-04-24 | Novozymes As | variante de uma subtilisina precursora, polinucleotídeo isolado, construção de ácido nucleico, vetor de expressão, célula hospedeira, composição de limpeza ou detergente, e, método para produzir uma variante |
DE102011118037A1 (de) | 2011-06-16 | 2012-12-20 | Henkel Ag & Co. Kgaa | Geschirrspülmittel mit Bleichkatalysator und Protease |
BR112013033816A2 (pt) | 2011-07-01 | 2017-02-14 | Novozymes A / S | composição de detergente líquido sem boro |
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JP2015500006A (ja) | 2011-11-25 | 2015-01-05 | ノボザイムス アクティーゼルスカブ | サブチラーゼ変異体およびこれをコードするポリヌクレオチド |
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EP2863759A2 (en) | 2012-06-20 | 2015-04-29 | Novozymes Biopolymer A/S | Use of polypeptides having protease activity in animal feed and detergents |
BR112015003724A2 (pt) | 2012-08-22 | 2017-08-08 | Novozymes As | polipeptídeo isolado, composição, uso de uma composição, polinucleotídeo isolado, constructo de ácido nucleico ou vetor de expressão, célula hospedeira recombinante, e, métodos de produção de um polipeptídeo e de produção de uma proteína. |
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CN104903443A (zh) | 2013-01-03 | 2015-09-09 | 诺维信公司 | α-淀粉酶变体以及对其进行编码的多核苷酸 |
EP2970830B1 (en) | 2013-03-14 | 2017-12-13 | Novozymes A/S | Enzyme and inhibitor contained in water-soluble films |
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- 2010-09-24 EP EP10757210.9A patent/EP2480650B1/en active Active
- 2010-09-24 MX MX2012003386A patent/MX339581B/es active IP Right Grant
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- 2010-09-24 US US13/496,240 patent/US20120172280A1/en not_active Abandoned
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- 2010-09-24 AU AU2010299799A patent/AU2010299799B2/en not_active Ceased
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WO2011036263A1 (en) | 2011-03-31 |
RU2651525C2 (ru) | 2018-04-19 |
BR112012006487A2 (pt) | 2016-11-22 |
EP2480650A1 (en) | 2012-08-01 |
JP2013505711A (ja) | 2013-02-21 |
MX2012003386A (es) | 2012-06-27 |
CN102648273A (zh) | 2012-08-22 |
US20120172280A1 (en) | 2012-07-05 |
JP2016052334A (ja) | 2016-04-14 |
CA2775045A1 (en) | 2011-03-31 |
JP5947213B2 (ja) | 2016-07-06 |
MX339581B (es) | 2016-06-01 |
AU2010299799B2 (en) | 2015-10-29 |
CN102648273B (zh) | 2017-04-26 |
AU2010299799A1 (en) | 2012-04-05 |
BR112012006487A8 (pt) | 2018-04-24 |
EP2480650B1 (en) | 2017-03-22 |
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