FEBS Letters | |
Involvement of the hydrophobic stack residues 39–44 of factor VIIa in tissue factor interactions | |
Petersen, Lars C.1  Schiødt, Jakob2  Christensen, Ulla3  | |
[1] Biopharmaceuticals Research, Novo Nordisk A/S Hagedornsvej 1, DK-2820 Copenhagen Gentofte, Denmark;Department of Chemistry, Carlsberg Laboratory, Gamle Carlsberg Vej 10, DK-2500 Copenhagen Valby, Denmark;Department of Chemistry, University of Copenhagen, Universitetsparken, DK-2100 Copenhagen, Denmark | |
关键词: Factor VII; Tissue factor; Des(1–38) factor VIIa; Des(1–44) factor VIIa; Ca2+-ion binding; Gla-; γ-carboxy-glutamic acid; EGF domain; epidermal growth factor-like domain; TF; tissue factor; (H-d-Ile-Pro-Arg-pNA); H-d-isoleycyl-l-prolyl-l-arginine-p-nitroanilide; tPA; tissue type plasminogen activator; uPA; urokinase-type plasminogen activator; PC; phosphatidyl choline; PS; phosphatidyl serine; | |
DOI : 10.1016/0014-5793(94)00513-3 | |
学科分类:生物化学/生物物理 | |
来源: John Wiley & Sons Ltd. | |
【 摘 要 】
Des(1–38) factor VIIa and des(1–44) factor VIIa were obtained by limited proteolysis. The binding of tissue factor to these factor VIIa-derivatives was assessed from its stimulation of the proteolytic activity on chromogenic obgopeptide substrates. Compared to native factor VIIa (K TF = 0.6 ± 0.1 nM), Tissue factor binds to des(1–38) factor VIIa with a lower, but still significant affinity (K TF = 4.8 ± 0.3 nM). The activity of des(1–44) factor VIIa was only slightly stimulated by TF (K TF ∼ 200 nM). Binding of TF depends critically on the presence of Ca2+ ions. Ca2+ ions stimulated the activity of factor VIIa/TF with an apparent K Ca = 0.16 ± 0.02 mM. Factor VIIa in the absence of tissue factor was stimulated by Ca2+ with an apparent K Ca = 0.05 ± 0.01 mM, and similar K Ca values were obtained for the truncated derivatives of factor VIIa. Measurements of Ca2+-induced changes in intrinsic protein fluorescence suggest a conformational change. The Ca2+ ion concentration at which this change occurred was higher for des(1–44) factor VIIa (apparent K Ca = 0.14 mM) than for des(1–38)- and native factor VIIa (apparent K Ca = 0.04 mM). The Tb3+ ion luminescence technique was used to further investigate the Ca2+ binding sites. Tb3+ ions bound with a lower affinity to des(1–44) factor VIIa than to des(1–38)-and native factor VIIa. The observed drastic decrease in affinity for tissue factor as a result of truncation of the ‘hydrophobic stack’ residues 39–44, suggest that this region of factor VIIa provides a structural determinant that together with other regions participates in tissue factor binding.
【 授权许可】
Unknown
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