1996 Fiscal Year Final Research Report Summary
Development of Highly Sensitive Methods for the Detection of Fungi Utilizing the Horseshoe Crab Hemolymph Coagulation Cascade.
| Project/Area Number |
07557021
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 試験 |
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| Research Field |
Pathological medical chemistry
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
MUTA Tatsushi Kyushu Univ., Biology, Research Associate, 理学部, 助手 (60222337)
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| Co-Investigator(Kenkyū-buntansha) |
TANAKA Shigenori Seikagaku Corp., Technical Div., Group manager, 開発企画部, 診断薬室長
KAWABATA Shun-ichiro Kyushu Univ., Biology, Research Associate, 理学部, 助手 (90183037)
IWANAGA Sadaaki Kyushu Univ., Biology, Professor Emeritus, 理学部, 名誉教授 (90029942)
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| Project Period (FY) |
1995 – 1996
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| Keywords | beta-glucan / pattern Recognition / Coagulation / Invertebrates / Horseshoe Crab / Fungi / Detection / Defense |
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| Research Abstract |
Horseshoe crab hemocyte lysate responds to (1*3)-beta-D-glucans, initiating an enzymatic cascade which culminates in clot formation. We have purified to homogeneity the serine protease zymogen, factor G,which is directly activated by (1*3)-beta-D-glucans and which initiates the hemolymph clotting cascade. Factor G is a heterodimeric protein composed of two noncovalently-associated subunits alpha (72 kDa) and beta (37 kDa). In the presence of (1*3)-beta-D-glucans such as curdlan and paramylon, factor G is autocatalytically activated to an active serine protease, named factor G^^-. This activation is accompanied by limited proteolyses of both subunits : the 72-kDa subunit alpha is cleaved to 55-kDa and 17-kDa fragments, and the 37-kDa subunit beta is shortened to 34-kDa. Longer incubations with (1*3)-beta-D-glucans result in cleavege of the 55-kDa fragment to 46-kDa and the 34-kDa fragment to 32-kDa, with concomitant loss of amidase activity. Reconstitution experiments using purified proteins participating in the hemolymph clotting cascade demonstrate that factor G^^- is capable of activating proclotting enzyme directly, resulting in the conversion of coagulogen to coagulin gel. Thus, purified factor G is shown to be the primary initiator of the (1*3)-beta-D-glucan-sensitive coagulation pathway in the horseshoe crab hemocyte lysate.
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