| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1994: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1993: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Research Abstract |
High and low molecular weight kininogens strongly inactivate cysteine proteases such as cathepsin B,H and L,calpain I and II,papain, and ficin in noncovalent and noncompetitive manners. It is thought that both kininogens play important physiological roles in the regulation of cysteine proteases. There are two inhibitory domains on the common heavy chain of both kininogens. It is speculated that domains 2 and 3 contain two region as potential candidates for the reactive site toward cysteine proteases : Gln-Val-Val-Ala-Gly (QVVAG) located at two-thirds of the distance from the N-terminus of the domains and Glycine (Gly) located at 44 amino acid residues upstream from the GVVAG region.
In this project, by using a cDNA coding for human low molecular weight kininogen, we had attempted to express the wild type protein and proteins with deletions of domain 2 and 3, and with deletions or substitutions of the QVVAG and Gly regions, in the eukaryotic cells such as BHK and CHO,in Bacureo virus system cells and in the prokaryotic cells such as E.coli.
Although the amounts of the wild type of low molecular weight kininogen with deletion of domains 2 or 3 (<less than or equal>50ng/ml) expressed by the induction with dexamethsone in BHK cells were very low. As the next steps, using site-directed cDNA for low molecular weight kininogen, we have been attemptting to express the wild type and other proteins with deletion and/or substitution in Bacuro virus system and in E
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