| Budget Amount *help |
¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000)
Fiscal Year 2010: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
Fiscal Year 2009: ¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000)
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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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