| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1996: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1995: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
beta2-Glycoprotein I (beta2GPI) is a cofactor in the recognition of the phospholipid antigen cardiolipin by anti-cardiolipin antibodies in autoimmune diseases such as systemic lupus erythematosus. beta2GPI (326 amino acids) consists of five repeating units (Domains I-V), each containing 60-80 amino acid residues, corresponding to the short consensus repeat of the complement control protein module. Recent studies suggest that Domain V plays important roles in the binding to cardiolipin and expression of cofactor activity. We studied the structure-function relationship of beta2GPI by preparing various derivatives.
1.We examined the interactions of various forms of bovine beta2GPI with phospholipid liposomes under different conditions. The results indicate that the N-terminal as well as C-terminal domains have an important role in the interaction of beta2GPI with cardiolipin, and that the three residual domains containing sialic acid have no significant effect on the interaction.
2.We constructed a high-level expression system for human Domain V using a methylotrophic yeast, Pichia pastoris. We found that the recombinant protein as the native disulfide bonds and a proper folded structure. For the three dimensional structure determination by NMR,with this expression system, we prepared 15N and 13C labeled Domain V.The NMR studies are on going.
3.Conformation, stability, and liposome-binding activity of the recombinant Domain V were characterized and compared with those of various beta2GPI derivatives. The results suggested that the region including Trp76-Thr78 has a critical role in binding to cardiolipin.
4.A nicked form of domain V in beta2GPI has been shown to have a lower ability to cardiolipin. Using the recombinant Domain V,we found that plasmin can produce the nicked form of domain V,suggesting the biological significance.
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