Evidence that a nicked C4b, C4b', is a functionally active C4b derivative

Factor I-catalyzed C4b cleavage is a regulatory reaction for the classical pathway of the complement system. Although the reaction was shown to be a two-step reaction, production of a nicked form of C4b, C4b', as an intermediate cleavage product and subsequent splitting of C4b' into C4c and C4d, it is not known which of the two steps represents the inactivation of the C4b function in the assembly of C3 convertase, C4b,2a. We have purified C4b' and assessed the ability of C4b' to assemble C3 convertase with C2 by utilizing size exclusion high performance liquid chromatography. Evidence was obtained demonstrating that C4b' still retains the function of C4b to assemble C3 convertase. Thus, the substantial step for the inactivation of the C4b function appears to be the second cleavage reaction, that is, the cleavage of C4b' into C4c and C4d.