Interaction between protein S and complement C4b-binding protein (C4BP) - Affinity studies using chimeras containing C4BP beta-chain short consensus repeats

Human C4b-binding protein (C4BP) is a regulator of the complement system an d plays an important role in the regulation of the anticoagulant protein C pathway. C4BP can bind anticoagulant protein S, resulting in a decreased co factor function of protein S for activated protein C, C4BP is a multimeric protein containing several identical alpha chains and a single beta-chain ( C4BP beta), each chain being composed of short consensus repeats (SCRs), Pr evious studies have localized the protein S binding site to the NH2-termina l SCR (SCR-1) of C4BP beta. To further localize the protein S binding site, we constructed chimeras containing C4BP beta SCR-1, SCR-2, SCR-3, SCR-1+2, SCR-1+3, and SCR-2+3 fused to tissue-type plasminogen activator. Binding a ssays of protein S with these chimeras indicated that SCR-S contributes to the interaction of protein S with SCR-1, since the affinity of protein S fo r SCR-1+2 was up to B-fold higher compared with SCR-1 and SCR-1+3. Using an assay that measures protein S cofactor activity, we showed that cofactor a ctivity was decreased due to binding to constructs that contain SCR-1. SCR- 1+2 inhibited more potently than SCR-1 and SCR-1+3. SCR-3 had no additional effect on SCR-1, and therefore the effect of SCR-S was specific. In conclu sion, beta-chain SCR-2 contributes to the interaction of C4BP with protein S.