title: Synergistic inhibition of the intrinsic factor X activation by protein S and C4b-binding protein.
abstract: The complement protein C4b-binding protein plays an important role in the regulation of the protein C anticoagulant pathway. C4b-binding protein can bind to protein S, thereby inhibiting the cofactor activity of protein S for activated protein C. In this report, we describe a new role for C4b-binding protein in coagulation. We observed inhibition of the intrinsic factor X activating reaction by the complex of C4b-binding protein and protein S. At the plasma concentration of protein S, the factor X activation was inhibited for 50% and addition of C4b-binding protein led to a potentiation of the inhibition to almost 90%. Because C4b-binding protein alone had no effect on the activation of factor X, we hypothesized that binding of C4b-binding protein to protein S was a prerequisite for optimal inhibition of factor X activation. C4b-binding protein lacking the beta-chain, which is unable to bind to protein S, did not potentiate the inhibitory effect of protein S. In an earlier study, we observed that C4b-binding protein increased the binding affinity of protein S for factor VIII. Therefore, a possible interaction of C4b-binding protein with factor VIII was investigated. C4b-binding protein bound to factor VIII and to thrombin activated factor VIII in a saturable and specific way. Also, factor VIII in complex with von Willebrand factor was able to bind C4b-binding protein. The beta-chain of C4b-binding protein was not required for the interaction with factor VIII because C4b-binding protein lacking the beta-chain also bound to factor VIII. Monoclonal antibodies directed against the alpha-chain of C4b-binding protein inhibited the binding to factor VIII, whereas monoclonal antibodies directed against the beta-chain had no effect on the binding to factor VIII. This finding indicates that the binding site for factor VIII on C4b-binding protein is localized on the alpha-chains of C4b-binding protein. The potentiation by C4b-binding protein of the inhibition of the factor X activation by protein S was blocked by a monoclonal antibody directed against the alpha-chain of C4b-binding protein. This finding indicates that the potentiation of the inhibitory effect of protein S was mediated via an interaction of C4b-binding protein with factor VIII. C4b-binding protein did not bind to factor V and was not able to potentiate the inhibitory effect of protein S on prothrombinase activity.(ABSTRACT TRUNCATED AT 400 WORDS)
authors.affiliation: Department of Haematology, University Hospital, Utrecht, The Netherlands.
authors.forename: S J
authors.initials: SJ
authors.lastname: Koppelman
authors.forename: C
authors.initials: C
authors.lastname: van't Veer
authors.forename: J J
authors.initials: JJ
authors.lastname: Sixma
authors.forename: B N
authors.initials: BN
authors.lastname: Bouma
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D003169:Complement Inactivator Proteins; D004357:Drug Synergism; D015949:Factor IXa / Q000494:pharmacology; D005169:Factor VIII / Q000378:metabolism / Q000494:pharmacology; D005170:Factor X / Q000378:metabolism*; D015951:Factor Xa / Q000378:metabolism; D006023:Glycoproteins; D006801:Humans; D007700:Kinetics; D017293:Protein S / Q000378:metabolism / Q000494:pharmacology*; D014841:von Willebrand Factor / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D003169:Complement Inactivator Proteins; D006023:Glycoproteins; D017293:Protein S; D014841:von Willebrand Factor; D005169:Factor VIII; D005170:Factor X; D015949:Factor IXa; D015951:Factor Xa