Insight into the Mechanism of Action of Heparin Cofactor II

Abstract

The anticoagulant activities of glycosammoglycans are mediated by antithrombin and heparin eofaetor II (H C II) (I). When antithronihin is bound to soluble heparin or to heparan sulfate in the vessel wall, it ra pidly inhibits thrombin, faetor Xa, and factor IXa. H C II inhibits throm bin, but not other eoagulation proteases, in the presence o f heparin or dermatan sulfate (2 ,3). This review will loeus on the biochemistry of H C II, with an emphasis on recent evidence that H C II and antithrombin inhibit thrombin by fundamentally different mechanisms. Structure H C II is a member of the a I-antitrypsin family of serine protease in hibitors (serpins). It is a single-chain glycoprotein with a mass of-6 6 kDa (2). The c D N A for human H C II encodes a polypeptide 480 amino acids in length preceded by a 19-residue signal peptide (4 ,5). The pro tein contains three N-linked glycosylation sites and three cysteine resi dues that apparently do not form disulfide bonds (6). H C II is-3 0 % identical in sequence to antithrombin and other serpins, the greatest similarity occurring in the C-terminal two-thirds of the protein. By contrast, the first 80 amino acid residues at the N-lerminal end o f H C II share no homology with other serpins. The N-terminal region includes a tandem repeat of two highly acidic sequences, each o f which contains a tyrosine residue that becomes O-sulfated during biosynthesis (Fig. I A) (7). As discussed below, the N-terminal acidic domain is required for rapid inhibition of thrombin in the presence o f a glycosaminoglycan. Murine plasma contains two forms o f H C II that appear to differ only in the composition of their N-linked oligosaccharides (8). H C II is also present in rabbit, rat, and dog (8-10), but it has not yet been identi fied in non-mammalian species. Human, murine, and rabbit H C II are 87% identical to one another in amino acid sequence. The sequences of the reactive site, the glycosaminoglycan-binding site, and the N-term i nal acidic domain (shown in Fig. IA for human H C II) are even more highly conserved. Some other human serpins, such as a I-antitrypsin and al-antichymotrypsin, are only-6 0 % identical to their murine homologues. The strong conservation of H C II implies that it serves the same physiologic function in different species.

Aspartic Acid Residues 72 and 75 and Tyrosine-sulfate 73 of Heparin Cofactor II Promote Intramolecular Interactions during Glycosaminoglycan Binding and Thrombin Inhibition

“…Heparin cofactor II has an atypical Leu 444 reactive center residue, whereas the more typical Arg is found in thrombin inhibitors like ATIII or protein C inhibitor (10, 12, 13). The inhibition of thrombin by HCII is enhanced by both heparin and dermatan sulfate, and the glycosaminoglycan binding site is primarily contained within the D-helix region of HCII (15,(27)(28)(29)(30)(31)(32)(33)(34). Although HCII is ϳ30% identical in sequence to antithrombin and other serpins, it has a unique N-terminal extension of ϳ80 residues that contains a tandem repeat of negatively charged acidic residues (9 Asp, 5 Glu, and 2 Tyrsulfate) (25,26).…”

“…One mechanism of thrombin regulation is by serine protease inhibitors (serpins) 1 such as antithrombin (ATIII) and heparin cofactor II (HCII) (10 -14). Heparin cofactor II and ATIII belong to a sub-class of serpins whose activity is greatly enhanced upon binding to glycosaminoglycans like heparin and heparan sulfate (HCII and ATIII) and dermatan sulfate (HCII) (15)(16)(17)(18). These glycosaminoglycans are found in vivo on cell surfaces and in extracellular matrix to support these inhibition reactions (19 -23).…”

“…The mechanism of glycosaminoglycan-accelerated thrombin inhibition by HCII is unique among thrombin-inhibiting serpins (15,16,47). HCII appears to employ an allosteric mechanism whereby binding of glycosaminoglycans to the D-helix region is thought to alter the acidic domain, which then serves as a "tethered-ligand" for exosite-1 of thrombin (27, 28, 31-37).…”

The status of new anticoagulants

“…The procoagulant effects of thrombin can be blocked either by inactivating the enzyme itself or by preventing its generation. Indirect thrombin inhibitors, such as heparin and dermatan sulphate, act by catalysing the naturally occurring thrombin inhibitors, antithrombin and/or heparin cofactor II (Hirsh, 1991b;Tollefsen, 1995;Weitz, 1997). Direct thrombin inhibitors bind to thrombin and block its interaction with substrates.…”

Plasma heparin cofactor II activity is inversely associated with left atrial volume and diastolic dysfunction in humans with cardiovascular risk factors

“…HCII potently inhibits thrombin action by forming a bimolecular complex with dermatan sulfate proteoglycans under the endothelial layer in mammalians. 8,9 Although AT inhibits not only the thrombin action, but also the actions of several proteases involved in blood coagulation or fibrinolysis, HCII only inactivates thrombin and has no inhibitory effect on the action of any other proteases. As HCII can counteract the actions of thrombin at injured vascular walls, we, and others, have investigated and confirmed the protective role of HCII against atherosclerosis in clinical examinations and studies using HCII-deficient mice.…”