CA2759465C - Method of producing polymers of spider silk proteins - Google Patents
Method of producing polymers of spider silk proteins Download PDFInfo
- Publication number
- CA2759465C CA2759465C CA2759465A CA2759465A CA2759465C CA 2759465 C CA2759465 C CA 2759465C CA 2759465 A CA2759465 A CA 2759465A CA 2759465 A CA2759465 A CA 2759465A CA 2759465 C CA2759465 C CA 2759465C
- Authority
- CA
- Canada
- Prior art keywords
- amino acid
- acid residues
- spider silk
- liquid medium
- silk protein
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/43504—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates
- C07K14/43513—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates from arachnidae
- C07K14/43518—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates from arachnidae from spiders
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Insects & Arthropods (AREA)
- Organic Chemistry (AREA)
- Biochemistry (AREA)
- Gastroenterology & Hepatology (AREA)
- Zoology (AREA)
- Toxicology (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Tropical Medicine & Parasitology (AREA)
- Peptides Or Proteins (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Applications Claiming Priority (7)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| EP09158445.8 | 2009-04-22 | ||
| EP09158445A EP2243792A1 (en) | 2009-04-22 | 2009-04-22 | Methods of producing polymers of spider silk proteins |
| EP10156923.4 | 2010-03-18 | ||
| EP10156927 | 2010-03-18 | ||
| EP10156923 | 2010-03-18 | ||
| EP10156927.5 | 2010-03-18 | ||
| PCT/SE2010/050439 WO2010123450A1 (en) | 2009-04-22 | 2010-04-21 | Method of producing polymers of spider silk proteins |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2759465A1 CA2759465A1 (en) | 2010-10-28 |
| CA2759465C true CA2759465C (en) | 2017-12-12 |
Family
ID=43011348
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA2759465A Expired - Fee Related CA2759465C (en) | 2009-04-22 | 2010-04-21 | Method of producing polymers of spider silk proteins |
Country Status (9)
| Country | Link |
|---|---|
| US (1) | US8642734B2 (enExample) |
| EP (1) | EP2421890B1 (enExample) |
| JP (1) | JP5912077B2 (enExample) |
| CN (1) | CN102395601A (enExample) |
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Families Citing this family (30)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ES2608673T3 (es) * | 2010-03-18 | 2017-04-12 | Spiber Technologies Ab | Producción de proteínas y polipéptidos |
| WO2012055854A1 (en) | 2010-10-27 | 2012-05-03 | Spiber Technologies Ab | Spider silk fusion protein structures for binding to an organic target |
| CN103502516B (zh) * | 2011-06-01 | 2015-04-22 | 丝芭博株式会社 | 人造多肽纤维及其制造方法 |
| EP2844360B1 (en) * | 2012-05-02 | 2018-12-19 | Spiber Technologies AB | Spider silk fusion protein structures without repetitive fragment for binding to an organic target |
| WO2015042164A2 (en) | 2013-09-17 | 2015-03-26 | Refactored Materials, Inc. | Methods and compositions for synthesizing improved silk fibers |
| PE20171791A1 (es) | 2014-12-02 | 2017-12-28 | Silk Therapeutics Inc | Prendas de vestir de seda y productos de alto rendimiento y metodos para elaborarla |
| CN114000218A (zh) | 2015-03-16 | 2022-02-01 | 保尔特纺织品公司 | 改善的丝纤维 |
| US20180080147A1 (en) | 2015-04-09 | 2018-03-22 | Spiber Inc. | Polar solvent solution and production method thereof |
| WO2016163337A1 (ja) | 2015-04-09 | 2016-10-13 | Ｓｐｉｂｅｒ株式会社 | 極性溶媒溶液及びその製造方法 |
| EP3307765B1 (en) * | 2015-06-11 | 2024-04-10 | Bolt Threads, Inc. | Recombinant protein fiber yarns with improved properties |
| CN105031723B (zh) * | 2015-06-23 | 2017-05-24 | 上海交通大学 | 基于蜘蛛丝蛋白的温敏性水凝胶 |
| HK1255640A1 (zh) | 2015-07-14 | 2019-08-23 | Evolved By Nature, Inc. | 丝性能服装和产品及其制备方法 |
| JP2018159137A (ja) | 2015-08-20 | 2018-10-11 | 国立研究開発法人理化学研究所 | クモ糸様構造を有するポリペプチド繊維の製造方法 |
| CN108431027B (zh) | 2015-11-13 | 2022-07-22 | 思百博技术股份公司 | 电荷逆转的n端蜘蛛丝蛋白结构域及其用途 |
| GB201522610D0 (en) * | 2015-12-22 | 2016-02-03 | Univ Southampton | Protein |
| KR20190028381A (ko) * | 2016-05-16 | 2019-03-18 | 스파이버 테크놀러지스 에이비 | 고체 표면의 거미 실크 코팅 |
| EP3469121A4 (en) * | 2016-06-10 | 2020-01-08 | Bolt Threads, Inc. | RECOMBINANT PROTEIN FIBER YARNS WITH IMPROVED PROPERTIES |
| EP3263593A1 (en) | 2016-07-01 | 2018-01-03 | Anna Rising | Engineered spider silk proteins and uses thereof |
| KR20190046993A (ko) | 2016-09-14 | 2019-05-07 | 볼트 쓰레즈, 인크. | 긴 균일한 재조합 단백질 섬유 |
| WO2018087239A1 (en) * | 2016-11-11 | 2018-05-17 | Amsilk Gmbh | Use of a shrinkable biopolymer fiber as sensor |
| CN107475807A (zh) * | 2017-08-30 | 2017-12-15 | 常州豫春化工有限公司 | 一种改性尼龙6纤维的制备方法 |
| AU2018340499B2 (en) * | 2017-09-26 | 2023-09-07 | Spiber Technologies Ab | Structuring of surface-active macromolecules |
| US11390988B2 (en) | 2017-09-27 | 2022-07-19 | Evolved By Nature, Inc. | Silk coated fabrics and products and methods of preparing the same |
| JP2021035330A (ja) | 2017-09-29 | 2021-03-04 | Ｓｐｉｂｅｒ株式会社 | 発現カセット |
| WO2021001840A1 (en) * | 2019-07-04 | 2021-01-07 | Seevix Material Sciences Ltd. | Prokaryotic expression system and methods of using the same |
| CN110551194B (zh) * | 2019-09-19 | 2023-03-21 | 天津大学 | 重组蜘蛛卵鞘丝蛋白复合物及其生成的人造卵鞘丝 |
| AU2021259692A1 (en) | 2020-04-23 | 2022-10-27 | Seevix Material Sciences Ltd. | Cosmetic compositions comprising dragline spider silk |
| WO2024091831A2 (en) * | 2022-10-28 | 2024-05-02 | National Taiwan University | A recombinant spider silk protein and method for tag-free and time-saving purification thereof |
| CN218413490U (zh) | 2022-10-31 | 2023-01-31 | 徐州龙安电子科技有限公司 | 一种新型蜘蛛电子标签 |
| DE102023105437B3 (de) | 2023-03-06 | 2024-06-13 | Dr. Ing. H.C. F. Porsche Aktiengesellschaft | Fahrzeugkomponente mit wenigstens einer Fahrzeuginnenraumkomponente |
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| US20070260039A1 (en) | 2002-01-11 | 2007-11-08 | Karatzas Costas N | Methods of Producing Silk Polypeptides and Products Thereof |
| ES2525095T3 (es) * | 2004-07-22 | 2014-12-17 | Amsilk Gmbh | Proteínas recombinantes de la seda de araña |
| AU2006333611B2 (en) * | 2005-12-30 | 2012-04-26 | Spiber Technologies Ab | Spider silk proteins and methods for producing spider silk proteins |
| BRPI0701826B1 (pt) * | 2007-03-16 | 2021-02-17 | Embrapa - Empresa Brasileira De Pesquisa Agropecuária | proteínas da teia de aranha nephilengys cruentata, avicularia juruensis e parawixia bistriata isoladas da biodiversidade brasileira |
| US9051383B2 (en) * | 2007-06-11 | 2015-06-09 | The Regents Of The University Of California | Spider silk dragline polynucleotides, polypeptides and methods of use thereof |
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- 2010-04-21 US US13/264,938 patent/US8642734B2/en active Active
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- 2010-04-21 JP JP2012507182A patent/JP5912077B2/ja active Active
- 2010-04-21 CN CN2010800169433A patent/CN102395601A/zh active Pending
- 2010-04-21 DK DK10767397.2T patent/DK2421890T3/en active
- 2010-04-21 AU AU2010239773A patent/AU2010239773B2/en not_active Ceased
- 2010-04-21 WO PCT/SE2010/050439 patent/WO2010123450A1/en not_active Ceased
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| CN102395601A (zh) | 2012-03-28 |
| DK2421890T3 (en) | 2015-11-23 |
| EP2421890A1 (en) | 2012-02-29 |
| AU2010239773B2 (en) | 2014-11-06 |
| CA2759465A1 (en) | 2010-10-28 |
| JP5912077B2 (ja) | 2016-04-27 |
| WO2010123450A1 (en) | 2010-10-28 |
| US20120041177A1 (en) | 2012-02-16 |
| AU2010239773A1 (en) | 2011-11-03 |
| EP2421890B1 (en) | 2015-08-26 |
| JP2012531889A (ja) | 2012-12-13 |
| RU2011145429A (ru) | 2013-05-27 |
| EP2421890A4 (en) | 2012-10-24 |
| US8642734B2 (en) | 2014-02-04 |
| RU2560437C2 (ru) | 2015-08-20 |
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