2010 Fiscal Year Final Research Report
Molecular dissection of Streptomyces trypsin on the recognition mechanism of structural protein substrates
| Project/Area Number |
21750183
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| Research Category |
Grant-in-Aid for Young Scientists (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Chemistry related to living body
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| Research Institution | Kyoto University |
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Principal Investigator |
UESUGI Yoshiko Kyoto University, 再生医科学研究所, 研究員 (30416416)
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| Project Period (FY) |
2009 – 2010
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| Keywords | 酵素 / 基質認識 / 高分子基質 / セリンプロテアーゼ / 放線菌 / RIBSシャフリング |
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| Research Abstract |
Serine proteases have recently received much attention in relation to degradation of recalcitrant animal proteins, such as collagen, keratin, blood clots, and amyloid prion proteins for beneficial use of industrial waste and medical applications. In this study, a serine protease from Streptomyces omiyaensis (SOT), which is a trypsin-like enzyme, was chosen as a model enzyme for clarifying the recognition mechanism of structural protein substrates in serine proteases. We constructed chimeras between SOT and SGT using an in vivo DNA shuffling system and several mutants to identify the key residues involved in topological specificities. By comparing substrate specificities of chimeras and mutants, we found that residues 71 and 72 of SOT contribute to topological specificity and collagen binding, respectively. Furthermore, we found that Lys217 and Ala221 of SOT, which is located in the C-terminal α-helix domain, as a crucial residues for fibrinolysis.
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