Zinc is an essential trace element for human nutrition and is critical to the structure, stability, and function of many proteins. Zinc ions were shown to enhance activation of the intrinsic pathway of coagulation but down-regulate the extrinsic pathway of coagulation. The protein C pathway plays a key role in blood coagulation and inflammation. At present there is no information on whether zinc modulates the protein C pathway. In the present study we found that Zn(2+) enhanced the binding of protein C/activated protein C (APC) to endothelial cell protein C receptor (EPCR) on endothelial cells. Binding kinetics revealed that Zn(2+) increased the binding affinities of protein C/APC to EPCR. Equilibrium dialysis with (65)Zn(2+) revealed that Zn(2+) bound to the Gla domain as well as sites outside of the Gla domain of protein C/APC. Intrinsic fluorescence measurements suggested that Zn(2+) binding induces conformational changes in protein C/APC. Zn(2+) binding to APC inhibited the amidolytic activity of APC, but the inhibition was reversed by Ca(2+). Zn(2+) increased the rate of APC generation on endothelial cells in the presence of physiological concentrations of Ca(2+) but did not further enhance increased APC generation obtained in the presence of physiological concentrations of Mg(2+) with Ca(2+). Zn(2+) had no effect on the anticoagulant activity of APC. Zn(2+) enhanced APC-mediated activation of protease activated receptor 1 and p44/42 MAPK. Overall, our data show that Zn(2+) binds to protein C/APC, which results in conformational changes in protein C/APC that favor their binding to EPCR.