Heparin cofactor II. Purification and properties of a heparin-dependent inhibitor of thrombin in human plasma.

We have isolated a previously unrecognized heparin-dependent inhibitor of thrombin from human plasma. The inhibitor, designated heparin cofactor II (HCII), was purified to homogeneity with sulfated-dextran, DEAE-Sepharose, heparin-Sepharose and Sephadex G-150. HCII is a glycoprotein consisting of a single polypeptide chain with a Mr = 65,600 as determined by sedimentation equilibrium analysis. Other physical properties include s20,w = 4.31 S; Stokes radius = 34 A; E280 1% = 11.7; and pI = 4.95 to 5.15. The purified inhibitor is not precipitated by antibodies directed against seven other plasma protease inhibitors, including antithrombin III (ATIII). HCII blocks the proteolytic and amidolytic activities of thrombin by forming a covalent, 1:1 molar complex with the protease. The second-order rate constant for inhibition of thrombin by purified HCII increases from 5.0 X 10(5) M-1 min-1 in the absence of heparin to 4.5 x 10(8) M-1 min-1 at optimal heparin concentrations of 0.8 to 1.0 unit/ml. In comparison with ATIII, HCII is a relatively ineffective inhibitor of coagulation factor Xa.