Analysis of the reactive site peptide bond in C1-inhibitor by chemical modification of tyrosyl, lysyl, and arginyl residues: the essential role of lysyl residues in the functional activity of C1-INH
Minta, J.O.; Aziz, E.
Journal of Immunology 126(1): 250-255
1981
ISSN/ISBN: 0022-1767 PMID: 6450235 Document Number: 172216
Chemical modification of human C.hivin.1-INH [complement component 1 esterase inhibitor] with group-specific reagents known to modify the tyrosyl residues (tetranitromethane, TMN), arginyl residues (cyclohexanedione, CHD; phenylglyoxal, PGO) and lysyl residues (2,4,6-trinitrobenzenesulfonic acid, TNBS; acetic anhydride; citraconic anhydride, CA) and their effects on its C.hivin.1s inhibitory activity were studied. Modification of 60% of the tyrosyl residues in C.hivin.1-INH did not markedly affect the capacity of C.hivin.1-INH to inhibit C.hivin.1s activity; apparently tyrosine residues are not an integral part of C.hivin.1-INH function. Modification of the guanidinium groups of arginine with cyclohexanedione was without appreciable effect on C.hivin.1s inhibitory activity of C.hivin.1-INH. When PGO was utilized for modification of arginyl residues, C.hivin.1-INH activity was destroyed. Regeneration of PGO-blocked arginyl residues in C.hivin.1-INH was associated with only a minor recovery of C.hivin.1s inhibitory activity. By amino acid analyses, the destruction of C.hivin.1-INH activity by PGO was due to a side reaction with lysyl residues. Modification of .epsilon.-amino groups of lysine residues with TNBS, acetic anhydride and citraconic anhydride resulted in extensive destruction of C.hivin.1-INH activity. When citraconyl groups were deblocked from CA-modified and inactivated C.hivin.1-INH, complete recovery of C.hivin.1-INH activity was achieved. Other than the acetylation of C.hivin.1-INH with acetic anhydride, all other chemically modified C.hivin.1-INH derivatives studied retained their electrophoretic homogeneity and immunologic reactivities against antisera to native C.hivin.1-INH. C.hivin.1-INH is a lysyl-type inhibitor with a lys-X bond at its reactive site with C.hivin.1s.