Molecular characterization of the catalytic domains of human complement serine protease C.hivin.1r

Abstract

Limited cleavages of human C.hivin.1r by extrinsic proteases of various specificity (plasmin, elastase, chymotrypsin, thermolysin) yield dimeric associations of two globular domains, each comprised of the intact B chain disulfide linked to .gamma., the C-terminal fragment of the A chain. These (.gamma.-B)2 domains, which are homologous to those obtained from C.hivin.1r by autolytic cleavage [Villiers, C. L., Arlaud, G. J., and Colomb, M. G. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 4477-4481], represent the core of the C.hivin.1r molecule and are associated with the catalytic properties of the serine active site. V8 protease also yields (.gamma.-B)2 associations, although additional cleavages occur in the B chain. Sequence analysis shows that all cleavages generating the .gamma. fragments occur within a 13-residue sequence extending from positions 274 to 286 of the C.hivin.1r A chain. Chemical cross-linking with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide of the (.gamma.-B)2 catalytic domains obtained from C.hivin.1r autolytic cleavage indicates that each .gamma.-B domain interacts with its neighbor in a "head to tail" configuration, the .gamma. region of one domain interacting with the B chain of the other domain, and conversely. No evidence is found of .gamma.-.gamma. or B-B interactions. Such a head to tail configuration, placed in the context of the model proposed for the C1s-C1r-C1r-C1s catalytic subunit of C1 [Colomb, M. G., Arlaud, G. J., and Villiers, C. L. (1984) Philos. Trans. R. Soc. London, B 306, 283-292], is compatible with autolytic activation of C1r through an intramolecular cross-mechanism and with subsequent activation of C1s by activated C.hivin.1r.