Influence of the gamma-carboxyglutamic acid-rich domain and hydrophobic stack of factor VIIa on tissue factor binding.
Des (1-38)- and des(1-44)-factor VIIa (fVIIa) were inhibited with Phe-Phe-Arg-chloromethyl ketone (FFR-cmk). Des(1-38)-FFR-fVIIa inhibited tissue factor (TF)-enhanced fVIIa amidolytic activity with an IC50 value of 15 nM, whereas 3- and 6-fold higher values were obtained with des(1-44)-FFR-fVIIa using soluble and full-length TF, respectively. The value for FFR-fVIIa was 8-10 nM. Clotting time was prolongated with IC50 values for inactivated des(1-38)- and des(1-44)-fVIIa of 50 nM and 1 mu M, respectively, whereas FFR-fVIIa yielded a value of 10 nM. From binding experiments in a BIA-core instrument, dissociation constants for the complexes between TF1-218 and fVIIa, des(1-38)-fVIIa and des(1-44)-fVIIa of about 3, 20 and 70 nM, respectively, could be estimated.
Persson
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Vessel Wall Biology-Hemostasis, Health Care Discovery, Novo Nordisk A/S, Gentofte, Denmark.
D015055:1-Carboxyglutamic Acid / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D001780:Blood Coagulation Tests; D002478:Cells, Cultured; D006224:Cricetinae; D015942:Factor VIIa / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012995:Solubility; D013925:Thromboplastin / Q000737:chemistry*; D014867:Water / Q000737:chemistry
D014867:Water; D015055:1-Carboxyglutamic Acid; D013925:Thromboplastin; D015942:Factor VIIa