Functional domains in the heavy-chain region of factor XI: a high molecular weight kininogen-binding site and a substrate-binding site for factor IX.
To probe the molecular interactions of factor XI we have prepared two monoclonal antibodies (MoAbs; 5F7 and 3C1), each of which binds the heavy chain of reduced and alkylated factor XIa. Competitive solid phase radioimmunoassay (RIA) binding studies revealed that 5F7 and 3C1 are directed against different epitopes within factor XI. One antibody (5F7) blocked the surface-mediated proteolytic activation of factor XI and its binding to HMW kininogen, but had no effect on factor-XIa-catalyzed factor IX activation. The other antibody (3C1) is a competitive inhibitor of factor-IX activation by factor XIa, but blocked factor-XI binding to HMW kininogen only at 1,000-fold higher concentration than 5F7. Moreover, HMW kininogen had no effect on the kinetics of factor-XIa-catalyzed factor-IX activation. Furthermore, factor XI CNBr peptide fragments that bind to the 5F7 and 3C1 antibodies were isolated. The peptides that bound to the 5F7 antibody blocked the binding of HMW kininogen to factor XI but did not inhibit factor-XIa-catalyzed factor-IX activation. However, the peptides isolated by the 3C1 antibody inhibited factor-XIa-catalyzed factor-IX activation and had no effect on factor-XI binding to HMW kininogen. Our results indicate that distinct functional domains within the heavy chain region of factor XI are important for the binding of factor XI to HMW kininogen and for activation of factor IX by factor XIa.
Baglia
F A
FA
Department of Medicine, Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140.
Sinha
D
D
Walsh
P N
PN
D000911:Antibodies, Monoclonal; D000937:Antigen-Antibody Reactions; D001665:Binding Sites; D001667:Binding, Competitive; D004789:Enzyme Activation; D005164:Factor IX / Q000378:metabolism*; D005172:Factor XI / Q000378:metabolism / Q000648:ultrastructure*; D006801:Humans; D007140:Immunoglobulin Fab Fragments; D007158:Immunologic Techniques; D066298:In Vitro Techniques; D007704:Kininogens / Q000378:metabolism*; D046911:Macromolecular Substances; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007704:Kininogens; D046911:Macromolecular Substances; D010446:Peptide Fragments; D005164:Factor IX; D005172:Factor XI