Mutations in alpha-chain of C4BP that selectively affect its factor I cofactor function.

Abstract C4b-binding protein (C4BP) inhibits all pathways of complement activation, acting as a cofactor to the serine protease factor I (FI) in the degradation of activated complement factors C4b and C3b. C4BP is a disulfide-linked polymer of seven α-chains and a unique β-chain, the α- and β-chains being composed of eight and three complement control protein (CCP) domains, respectively. In previous studies we have localized cofactor activity and binding of C4b to α-chain CCP1–3 of C4BP, whereas the binding of C3b required additionally CCP4. Likewise, introduced point mutations that decreased binding of C4b/C3b caused a decrease in cofactor activity. In the present study, we describe two mutants of C4BP, K126Q/K128Q and F144S/F149S, clustered on α-chain CCP3, which selectively lost their ability to act as cofactors in the cleavage of both C4b and C3b. Both mutants show the same binding affinity for C4b/C3b as measured by surface plasmon resonance and have the same inhibitory effect on formation and decay of the classical pathway C3-convertase as the wild type C4BP. It appears that C4b and C3b do not undergo the same conformational changes upon binding to the C4BP mutants as during the interaction with the wild type C4BP, which then results in the observed loss of the cofactor activity.