| Project/Area Number |
11304058
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
動物生理・代謝
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| Research Institution | Osaka University |
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Principal Investigator |
TOKUNAGA Fumio Osaka University, Graduate School of Science, Professor, 大学院・理学研究科, 教授 (80025452)
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| Co-Investigator(Kenkyū-buntansha) |
SASAKI Jun Osaka University, Graduate School of Science, Research associate, 大学院・理学研究科, 助手 (70314359)
HISATOMI Osamu Osaka University, Graduate School of Science, Associate Professor, 大学院・理学研究科, 助教授 (60231544)
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| Project Period (FY) |
1999 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥39,060,000 (Direct Cost: ¥38,100,000、Indirect Cost: ¥960,000)
Fiscal Year 2001: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2000: ¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 1999: ¥30,900,000 (Direct Cost: ¥30,900,000)
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| Keywords | Vision / Retina / Visual cells / Atomic force microscopy / Rhodopsin / Functional protein / Information exchange / Light information exchange / 視物質 / 細胞内情報伝達 / 1分子測定 |
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| Research Abstract |
Visual cells convert light signals into ele ctric signals which consist membrane potentials of visual cell. When visual pigments absorbs photons, they activate transducin, which activates phosphodiesterase. Phosphodiesterase catalyses hydrolysis of cyclic guanyl monophosphate into guanyl monophosphate which closes GMP-depending sodiun channels. This is visual excitation. Guanylate cyclase, rhodopsin kinase. arrestin, s-moduline, phosducin, guanylate cyclase activating protein are interacted and produce the most suitable electrical signals against photon energies. In this study, we tried to observe directly the situations of the molecules and the interactions between the molecules and analyze them. Though resolution of atom ; force microscope is theoretically less than an angstrom, it was neccessary to improve cantilevers and sample preparatic. for observing them under an atomic force microscope. These improvements will be continued. The following projects has been performed within this year. The monoclonal antibodies raised against bovine rhodopsin were prepared. We isolated the monoclonal antibodies which have higher reactivity after the light illumination. The epitope of one of the antibodies was determined to be the region between Arg 21 and Glu 25 in bovine rhodopsin. The replicas of crayfish visual rhabdoms and newt rod outer segments were prepared and were observed by an electron microscope and an atomic force microscope. We cloned cDNAs encoding guanylate cyclase, rhodopsin kinase, arrestin, s-modulin, phosducin and guanylate cyclase activating protein (GCAP). We cloned their cDNAs into the expression vectors, expressed their proteins in Escherichia coli and confirmed that the expressed proteins showed the same properties as the authentic proteins. The chimera protein of bacteriorhodopsin and sensory rhodopsin and the chimera protein combined with the transducer protein were expressed in E coli and were observed the atomic force microscope.
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