Reactive site peptide structural similarity between heparin cofactor II and antithrombin III

• Published in: The Journal of biological chemistry 1985-02, Vol.260 (4), p.2218-2225
• Main Authors: Griffith, M J, Noyes, C M, Church, F C
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Antithrombin III - metabolism; Binding Sites; Biological and medical sciences; Blood coagulation. Blood cells; Chromatography, High Pressure Liquid; Fundamental and applied biological sciences. Psychology; General aspects, investigation methods, hemostasis, fibrinolysis; Glycoproteins - metabolism; Heparin Cofactor II; Humans; Kinetics; Molecular and cellular biology; Peptide Fragments; Thrombin - metabolism; Trypsin
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)89541-2

Heparin cofactor II (Mr = 65,600) was purified 1800-fold from human plasma to further characterize the structural and functional properties of the protein as they compare to antithrombin III (Mr = 56,600). Heparin cofactor II and antithrombin III are functionally similar in that both proteins have been shown to inhibit thrombin at accelerated rates in the presence of heparin. There was little evidence for structural homology between heparin cofactor II and antithrombin III when high performance liquid chromatography-tryptic peptide maps and NH2-terminal sequences were compared. A partially degraded form of heparin cofactor II was also obtained in which a significant portion (Mr = 8,000) of the NH2 terminus was missing. The rates of thrombin inhibition (+/- heparin) by native and partially degraded-heparin cofactor II were not significantly different, suggesting that the NH2-terminal region of the protein is not essential either for heparin binding or for thrombin inhibition. A significant degree of similarity was found in the COOH-terminal regions of the proteins when the primary structures of the reactive site peptides, i.e. the peptides which are COOH-terminal to the reactive site peptide bonds cleaved by thrombin, were compared. Of the 36 residues identified, 19 residues in the reactive site peptide sequence of heparin cofactor II could be aligned with residues in the reactive site peptide from antithrombin III. While the similarities in primary structure suggest that heparin cofactor II may be an additional member of the superfamily of proteins consisting of antithrombin III, alpha 1-antitrypsin, alpha 1-antichymotrypsin and ovalbumin, the differences in structure could account for differences in protease specificity and reactivity toward thrombin. In particular, a disulfide bond which links the COOH-terminal (reactive site) region of antithrombin III to the remainder of the molecule and is important for the heparin-induced conformational change in the protein and high affinity binding of heparin does not appear to exist in heparin cofactor II. This observation provides an initial indication that while the reported kinetic mechanisms of action of heparin in accelerating the heparin cofactor II/thrombin and antithrombin III/thrombin reactions are similar, the mechanisms and effects of heparin binding to the two inhibitors may be different.