Characterization of the interaction between the light chain of factor VIIa and tissue factor.
Factor VIIa (fVIIa) consists of a heavy chain (serine protease domain) and a light chain (gamma-carboxyglutamic acid (Gla)-rich and epidermal growth factor (EGF)-like domains). The light chain, primarily the first EGF-like domain, appears to provide most of the binding energy in the interaction with tissue factor (TF). The Ca2+-binding sites in the protease domain and in the first EGF-like domain influence activity and interaction with TF, but the contribution from the Ca2+-binding sites in the Gla domain has not been established. We have compared the soluble TF (sTF)-binding properties of intact fVIIa to those of a fragment comprising almost the entire light chain and a small disulphide-linked peptide from the protease domain. Half-maximal binding of fVIIa and the light chain to sTF occurred around 0.3 and 1 mM Ca2+, respectively. The Ca2+ dependence of light-chain binding indicates an influence of Ca2+ binding to the Gla domain on the interaction between fVIIa and sTF. Comparison of the sTF-binding properties of fVIIa and a truncated variant lacking the Gla domain suggests that this domain interferes with sTF association at suboptimal Ca2+ concentrations. The light chain of fVIIa associated 5-fold slower with sTF than did fVIIa at saturating Ca2+ concentrations, whereas the dissociation of its complex with sTF was at least 100-fold faster than that of fVIIa:sTF. This gave a dissociation constant of 1-2 microM for the interaction between the light chain and sTF compared to about 3 nM for the fVIIa:sTF interaction.
Persson
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Vessel Wall Biology, Health Care Discovery, Novo Nordisk A/S, Gentofte, Denmark. egpe@novo.dk
D015055:1-Carboxyglutamic Acid / Q000378:metabolism; D000818:Animals; D015374:Biosensing Techniques; D002118:Calcium / Q000494:pharmacology; D002478:Cells, Cultured; D003029:Coagulants / Q000378:metabolism*; D006224:Cricetinae; D015942:Factor VIIa / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D013925:Thromboplastin / Q000378:metabolism*
D003029:Coagulants; D011994:Recombinant Proteins; D015055:1-Carboxyglutamic Acid; D013925:Thromboplastin; D015942:Factor VIIa; D002118:Calcium