| Project/Area Number |
63480515
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
生物物性学
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| Research Institution | Nagoya University (1989)
Kyushu University (1988) |
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Principal Investigator |
GO Mitiko Nagoya University, Department of Biology, Professor, 理学部, 教授 (70037290)
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| Co-Investigator(Kenkyū-buntansha) |
YAMAGAWA Hiroshi Mitsubishi Kasei Institute of Life Science, Senior Researcher, 主任研究員
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| Project Period (FY) |
1988 – 1989
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥6,500,000 (Direct Cost: ¥6,500,000)
Fiscal Year 1989: ¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1988: ¥4,300,000 (Direct Cost: ¥4,300,000)
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| Keywords | Globular protein / Module / Exon shuffling / Barnase / Chemical synthesis of module / Secondary structure of module / Hydrogen-bond network / Protein evolution / タンパク質 / ペプチド合成 / モジュ-ルの2次構造 / 疎水結合 / モジュール / バルネース / リボヌクレアーゼ / モジュールのコンフォメーション / ドメイン |
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| Research Abstract |
A globular protein consists of compact modules and module boundaries correspond close to intron positions in split genes. This fact implies that exon shuffling in genes is module shuffling in protein level and niodules must be primitive enzymes themselves or parts of them. The purpose of this project is to obtain supporting evidences for this module hypothesis. We synthesized modules chemically and carried out experimental study as well as computational work on conformation of each module in solution and its function if any. We chose barnase, an extracellular ribonuclease produced by Bacillus amyloliquefaciens, as a suitable protein for this purpose.
Barnase is a small protein consisting of 110 amino acid residues, it has no disulfide bonds, its X-ray structure was analyzed and the atomic coordinates were kindly supplied by Drs. G. Dodson and C. Hill. Barnase was decomposed into at least six modules (Ml-M6), the polypeptide segments corresponding to each of them was synthesized and purified. CD measurement shows that Ml and M2 have stable alpha-helices in lipophilic solvent. Thus, the helix forming potential of modules disconnected from each other reflects helical content in connected modules as native protein. M5 has a beta-sheet structure in water. The helix making residues in M2 were identified by NMR study using distance geometry method. The atomic forces to make the modules compact in barnase were analyzed by computational method focusing on the hydrogen-bond network. Hydrogen-bond network was found predominantly within each module and quite few between modules. This fact shows the important contribution of hydrogen-bond network in making modules compact in a globular protein. This research project brought out an evidence for the first time that at least three modules of barnase took stable secondary structures in solution. This result prompts us to carry out a further study on the structure and function of modules.
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