Factor I-Dependent Inactivation of Human Complement C4b of the Classical Pathway by C3b/C4b Receptor (CR1, CD35) and Membrane Cofactor Protein (MCP, CD46)1

Proteolytic inactivation of C4b is a crucial step for regulation of the classical complement pathway. A plasma protease factor I and membrane cofactors, C3b/C4b receptor (CR1) and membrane cofactor protein (MCP), participate in the regulation of cell-bound C4b although the physiological potency of these cofactors remains unknown. We have examined the optimal conditions of the factor I-mediated C4b regulatory system using purified cofactors. CR1 being a cofactor at a cofactor/C4b ratio <0.1 (w/w), fluid phase C4b, and methylamine-treated C4 (C4ma) were degraded by factor I into C4bi: minimal C4d was generated in the fluid phase. Liposome-bound C4b (LAC4b), on the other hand, was degraded into C4c and C4d. CR1 showed two optimal pHs (6.0 and 7.5) for fluid phase C4b, but one (6.0) for LAC4b, and in both cases low conductivity conditions enhanced the C4bi generation. CR1 cofactor activity was barely influenced by the NP-40 concentration. On the other hand, MCP degraded C4b and C4ma, as a factor I-cofactor, more efficiently into C4c and C4d. Though MCP cofactor activity, like that of CR1, was enhanced under low conductivity conditions, it has only one optimal pH, 6.0, in both fluid and solid phases. Furthermore, as in the case of C3b cleavage, a sufficient NP-40 concentration to solubilize membrane was needed for MCP to express full cofactor activity for C4b, in contrast to CR1. MCP was less potent for C4b inactivation than for C3b inactivation, while CR1 acted as a slightly more effective cofactor for C4b cleavage than for C3b cleavage. C4 isotypic variants (C4A and C4B) did not modulate MCP/CR1 cofactor activity except that CR1 and factor I more efficiently cleaved C4ma of C4B than that of C4A in the fluid phase. Covalent C4b dimer, an element of a C5 convertase of the C3-independent classical pathway, was degraded similarly by factor I and either of CR1 or MCP. The results imply that although MCP and CR1 serve as C4b regulatory factor I-cofactors, their C4b inhibitory profiles are different from each other, and in fluid and solid phases their activities are not sufficient to block the classical pathway.