Open AccessRole of CCP2 of the C4b‐binding protein β‐chain in protein S binding evaluated by mutagenesis and monoclonal antibodies
Author(s) -
Webb Joanna H.,
Villoutreix Bruno O.,
Dahlbäck Björn,
Blom Anna M.
Publication year - 2003
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1033.2003.03365.x
Subject(s) - binding protein , complement control protein , recombinant dna , protein a/g , protein g , plasma protein binding , monoclonal antibody , binding site , mutant , binding domain , protein s , biochemistry , microbiology and biotechnology , chemistry , biology , classical complement pathway , complement system , antibody , gene , protein c , genetics , fusion protein
Complement regulator C4b‐binding protein (C4BP) and the anticoagulant vitamin K‐dependent protein S form a high affinity complex in human plasma. C4BP is composed of seven α‐chains and a unique β‐chain, each chain comprising repeating complement control protein (CCP) modules. The binding site for protein S mainly involves the first of the three β‐chain CCPs (CCP1). However, recently it has been suggested that CCP2 of the β‐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 β‐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 β‐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 β‐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.