2003 Fiscal Year Final Research Report Summary
Analysis of genes of C-type lectin domain-containing proteins with useful biological activities.
| Project/Area Number |
14580655
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | MEIJI PHARMACEUTICAL UNIVERSITY |
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Principal Investigator |
ATODA Hideko MEIJI PHARMACEUTICAL UNIVERSITY, FACULTY OF PHARMACY, Associate professor, 薬学部, 助教授 (20221046)
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| Project Period (FY) |
2002 – 2003
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| Keywords | C-type lectin / anticoagulant / heterodimer / genomic / calcium-binding protein / IX / X-bp |
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| Research Abstract |
Many proteins that contain C-type lectin-like domains are isolated from snake venom. They express a great variety of biological activities. IX/X-bp is one of the heterodimeric proteins isolated from the venom of habu snake (Trimeresurus flavoviridis) consisting of two C-type lectin-like domains. Two C-type lectin-like domains are swapped. This protein binds to blood coagulation factors IX and X in the presence of Ca ions and inhibits the blood coagulation cascade. Complementary DNA cloning of IX/X-bp indicated that each chain of IX/X-bp is encoded by respective gene.. Gene structure of snake venom C-type lectin-like protein is not yet reported. In the present study, we cloned the gene encoding IX/X-bp A chain. Total genomic DNA was prepared from the blood of Trimeresurus flavoviridis, and partially digested with Sau3Al. A genomic library was constructed using a Lambda EMBL3 cloning vector predigested by BamHl according to the manufacture's protocol. E. coli clone containing IX/X-bp A chain cDNA plasmid was cultured and the plasmid was prepared. Inserted DNA was cleaved off from the plasmid by the digestion with Sall and Notl and labeled with DIG to use as probe to screen the genomic library. One gene clone encoding the IX/X-bp A chain was isolated from the genomic library and the nucleotide sequence was determined by the dideoxynucleotide chain-termination method. The gene structure of IX/X-bp A chain is different from that of many vertebrate C-type lectin domains such as asialoglycoprotein receptors and lgE Fc ε receptor whose protein structures are very similar to that of IX/X-bp but are classified in the groups other than group VII by Drickamer. One of introns is inserted in the hinge region suggesting the contribution of the intron insertion to the domain swapping mechanism.
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