Inhibition of Recombinant Human Blood Coagulation Factor VIIa Amidolytic and Proteolytic Activity by Zinc Ions

Abstract

SummaryAlthough it is well established that calcium is an essential cofactor in blood coagulation, recent experimental evidence suggests that zinc may also play an important role in hemostasis. In the present study, we have examined the effects of zinc ions on the amidolytic and proteolytic activity of recombinant factor VIIa in the presence of physiological levels of calcium ions. The amidolytic activity of factor VIIa was inhibited half-maximally by 20 ΜM zinc. The amidolytic activity of a derivative of factor VIIa lacking the Γ-carboxyglutamic acid domain was also inhibited half-maximally by 20 ΜM zinc, suggesting that the mechanism of zinc inhibition of factor VIIa amidolytic activity did not involve its Γ-carboxyglutamic acid residues. The amidolytic activity of a complex of recombinant tissue factor and factor VIIa was inhibited half-maximally by 70 ΜM zinc. In contrast to the results obtained with factor VIIa, the amidolytic activities of other human vitamin K-dependent coagulation proteases including factor Xa, thrombin and activated protein C were not appreciably affected by 50-100 ΜM zinc. The proteolytic activation of factor X by a complex of factor VIIa and relipidated tissue factor apoprotein was inhibited half-maximally by 40 ΜM zinc, whereas activation of factor IX in this system was inhibited half-maximally by 70 ΜM zinc ions. Considerably higher levels of zinc (∽100 ΜM) were required to inhibit half-maximally the rate of factor X activation by a complex of factor VIIa and functional tissue factor on the surface of either a human bladder carcinoma cell line, J82, or stimulated human umbilical vein endothelial cells. Activation of factor IX by factor VIIa and tissue factor on the surface of J82 cells was not influenced by zinc. However, the activation rate of factor IX on human umbilical vein endothelial cells was inhibited half-maximally at 100 ΜM zinc. The activation of factor X by factor VIIa in the presence of small unilamellar phospholipid vesicles was inhibited half-maximally by 20 ΜM zinc whereas factor IX activation by factor VIIa was not appreciably influenced by 10-100 ΜM zinc. Our data demonstrate that plasma levels of zinc ions inhibit the amidolytic and proteolytic activities of factor VIIa. The mechanism of this inhibition, as well as its possible physiological relevance, is unknown.