2007 Fiscal Year Final Research Report Summary
Proline-peptide synthesis with a mutant of prolyl aminopeptidase via aminolysis reaction.
| Project/Area Number |
18550162
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Chemistry related to living body
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| Research Institution | Okayama Prefectural Technology Center for Agriculture |
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Principal Investigator |
HATANAKA Tadashi Okayama Prefectural Technology Center for Agriculture, Dept.of Enzymology, Group Leader (00344408)
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| Project Period (FY) |
2006 – 2007
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| Keywords | prolyl aminopeptidase / proline / proline-peptides / synthesis / Ser / Cys mutant |
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| Research Abstract |
We specifically examined an exopeptidase, prolyl aminopeptidase (PAP), as a target to synthesize proline-peptides conveniently. Using sequence-based screening, a PAP from Stieptomyces thermoluteus subsp. fuscus NBRC14270 (PAP14270) was obtained. From the PAP14270, 144Ser was replaced by Cys (scPAP14270) to give aminolysis activity. In contrast to wild type PAP14270, scPAP14270 produced polymer of proline benzyl ester and cyclo[Pro-Pro]. The products of mass were confirmed using LC/MS. Several factors affecting the reaction such as pH, concentration of substrate, and reaction time were measured for their effects. Furthermore, the correlation between substrate specificity in the proline-peptide synthesis and the log D value of acyl acceptors on aminolysis catalyzed by scPAP14270 was demonstrated. The results showed that dipeptide synthesis proceeded in a weakly acidic environment and that cyclization and polymerization took place in alkaline conditions. Furthermore, results suggest that almost all amino acid esters whose log D value is greater than 0 can be recognized as acyl acceptors except Hyp-OBzl. These findings support the use of PAPs as a leading tool for production of physiologically active proline-peptides.
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