tailieunhanh - Báo cáo khoa học: Role of CCP2 of the C4b-binding protein b-chain in protein S binding evaluated by mutagenesis and monoclonal antibodies

Complement regulator C4b-binding protein (C4BP) and the anticoagulant vitamin K-dependent protein S form a high affinity complexin human plasma. C4BP is composed of sevena-chains and a uniqueb-chain, each chain comprising repeating complement control protein (CCP) modules. The binding site for protein S mainly involves the first of the threeb-chain CCPs (CCP1). However, recently it has been suggested that CCP2 of theb-chain also contributes to the binding of proteinS. | Eur. J. Biochem. 270 93-100 2003 FEBS 2003 doi Role of CCP2 of the C4b-binding protein b-chain in protein S binding evaluated by mutagenesis and monoclonal antibodies Joanna H. Webb1 Bruno O. Villoutreix2 Bjorn Dahlback1 and Anna M. Blom1 Division of Clinical Chemistry Department of Laboratory Medicine Lund University Sweden 2INSERM U428 University of Paris V France Complement regulator C4b-binding protein C4BP and the anticoagulant vitamin K-dependent protein S form a high affinity complex in human plasma. CBBP is comoosed Iff seven a-chains and a unique b-chain each chain comprising repeating complement control protein CCP modules. The binding site for protein S mainly involves the first of the three b-chain CCPs CCP1 . However recently it has been suggested that CCP2 of the b-chain also contributes to the binding of protein S. To elucidate the structural background for the involvement of CCP2 in the protein S binding several recombinant b-chain CCP1-2 variants having mutations in CCP2 were expressed and tested for protein S binding. Mutations were chosen based on analysis of a homology model of the b-chain and included R60A R101A D66A L105A F114A I116A and H108A. All mutant proteins bound equally well as recombinant wild type to protein S. Several monoclonal antibodies against the b-chain CCP2 were raised and their influence on protein S binding characterized. Taken together the results suggest that the role of CCP2 in protein S binding is to orient and stabilize CCP1 rather than to be directly part of the binding site. Keywords binding site C4BP complement protein S structure-function relationship. C4b-binding protein C4BP is an important regulator of the classical pathway of complement. C4BP also affects the regulation of the coagulation system as it binds protein S which serves as a cofactor to the anticoagulant activated protein C 1 . C4BP and protein S form a high-affinity noncovalent 1 1 complex the interaction being .