| Project/Area Number |
63560113
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
発酵・醸造
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| Research Institution | UNIVERSITY OF OSAKA PREFECTURE |
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Principal Investigator |
ARAI Motoo UNIVERSITY OF OSAKA PREFECTURE, COLLEGE OF AGRICULTURE, PROFESSOR, 農学部, 教授 (80081537)
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| Project Period (FY) |
1988 – 1989
|
| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1989: ¥300,000 (Direct Cost: ¥300,000)
Fiscal Year 1988: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Keywords | alpha-AMYLASE / INHIBITOR / CLONING / アミラーゼ / インヒビター / クローニング |
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| Research Abstract |
Proteinaceous alpha-anylase inhibitor(T-76-I) produced by Streptomyces nitrosporeus T-76 was corboxymethylated and digested by TPCK trypsin. The resulting fragments were separated using HPLC on reverse-phase colum and each of then was applied to gas phase amino acid sequencer. Fifty-four amino acid residues(approx, 70%) of mature T-76-I were determined.
T-76-I gene was cloned in a 1.2kb Sau 3AI fragment using a synthetic oligonucleotide probe, which contained inosine at the position of 3rd letter of codons, corresponding to the N-teminal end of the inhibitor. The nucleotide sequence data of the inserted fragment showed that the fragment contained sole open reading frame of 330 nucleotide encoding a polypeptide of 110 amino acids with a Mr of 11,306 dalton. The amino acid sequence deduced from the nucleotide sequence indicated that the presumptive pre-T-76-I protein extends 33 amino acids of signal peptide to the N-teminus of the mature T-76-I protein consisting of 77 amino acids with a Mr of 8,082 dalton. The pre-T-76-I protein is demonstrated to be processed during secrerion.
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