| Project/Area Number |
63480021
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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 |
動物発生・生理学
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| Research Institution | Kanazawa University |
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Principal Investigator |
SUZUKI Norio Kanazawa University, Faculty of Science Noto Marine Laboratory, Professor, 理学部, 教授 (20082133)
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| Co-Investigator(Kenkyū-buntansha) |
YAMAGUCHI Masaaki Kanazawa University, Faculty of Science Noto Marine Laboratory, Instructor, 理学部, 助手 (60182458)
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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 |
¥7,100,000 (Direct Cost: ¥7,100,000)
Fiscal Year 1989: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1988: ¥5,100,000 (Direct Cost: ¥5,100,000)
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| Keywords | sea urchin / spermatozoa / acrosome reaction / fucose sulfate glycoconjugate / disulfide bond / oocytes / cAMP / egg jelly / フコ-ス硫酸 / タンパク質 / 卵外被物質 / 卵ゼリー / フコース硫酸タンパク質複合体(FSG) / 抗体 / シアル酸 / 精子活性化ぺプチド |
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| Research Abstract |
A fucose sulfate glycoconjugate (FSG) was isolated from the egg jelly of the sea urchin Hemicentrotus pulcherrimus by gel chromatogrqaphy on a Sepharose 2B columnn. FSG induced the acrosome reaction in H. pulcherrimus spermatozoa in a concentration-dependent manner, although it showed about half the activity of the original unfractionated jelly. Synthetic sperm-activating peptide I (SAP-I:Gly- Phe-Asp-Leu-Asn-Gly-Gly-Gly-Val-Gly) increased the rate of the acrosoine reaction induced by FSG; the maximal rate of the acrosome reaction with FSG and SAP-I being that of the unfractionated jelly. Pronase digestion of FSG resulted in an 50% decrease in induction of the acrosome reaction and also in the elevation of cAMP in sperm cells.
A sialoglycoprotein was also purified from the egg jelly of the sea urchin H. pulcherrimus. Sialoglycoprotein which consisted of sialic acid (90%, w/w) and protein (10%, w/w) did not cause induction of the acrosome reaction and sperm isoagglutination.
FSG which con
… More
tained one mol sulfate/mol fucose possessed 2.0 times protein to fucose by weight. The proteins in intact FSG were separated to two major (258 kDa and 237 kDa) and one minor (120 kDa) proteins by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) in the presence of 2- mercaptoethanol (2-ME) while the proteins could not be separated by HPLC in the presence of 0.1% SDS or SDS-PAGE without 2-ME. Wliell FSG was first carboxyniethylated with non-radioactive iodoacetic acid and then reduced with 2-ME and finally carboxymetliylated with carbon-14 lapeled iodoacetic acid, the most of radioactivity was detected in proteins corresponding to a 120 kDa protein. Carboxymetiiylated-FSG was less potent than intact FSG in induction of the acrosome reaction. Flicoidan, a fucose sulfate polymer, did not induce the acrosome reaction.
These results indicate that SAP-I promotes induction of the acrosome reaction by acting a specific co-factor of FSG and the protein moiety of FSG plays an important role in induction of the acrosome reaction in H. pulcherrimus spermatozoa.
Immunohistochemical study with various stages of oocytes using a mouse anti-FSG anti-serum indicated that FSG is formed in the cytoplasm of a young oocyte and secreted to the cell surface during the oogenesis. Less
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