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(Arteriosclerosis, Thrombosis, and Vascular Biology. 1997;17:925-931.)
© 1997 American Heart Association, Inc.

Articles

Structural Aspects of Heparin Responsible for Interactions With von Willebrand Factor

Lawrence F. Poletti; Karyn E. Bird; Dalila Marques; Robert B. Harris; Yasuo Suda; ; Michael Sobel

From the Division of Vascular Surgery (L.F.P., K.E.B., D.M., M.S.) and the Department of Biochemistry and Biophysics (R.B.H.), Medical College of Virginia, Virginia Commonwealth University, and H.H. McGuire Veterans Affairs Medical Center (L.F.P., K.E.B., D.M., M.S.), Richmond, Va and the Department of Chemistry, Osaka University, Osaka, Japan (Y.S.).

Correspondence to Michael Sobel, MD, Box 980108, Medical College of Virginia, Richmond, VA 23298. E-mail sobel{at}gems.vcu.edu

Abstract Unfractionated heparin (UFH) binds von Willebrand factor (vWF) and inhibits the vWF–platelet GP Ib interaction. For vWF, a heparin-binding domain has been identified, but for heparin, the structures that confer such activity are unknown. To investigate this, UFH was depolymerized by methods that yield structurally distinct fragments. The glycosaminoglycans (GAGs) produced were separated into five groups of homogeneous molecular weight (MW). Anti-Xa activity, vWF binding affinity, and vWF-dependent platelet agglutination were measured. Periodate oxidation but not heparinase digestion destroyed anti-Xa activity. At all MWs, periodate conferred greater vWF binding affinity and greater ability to inhibit platelet agglutination than heparinase. As an example, at MW 6100, the binding IC₅₀ was 100±19 µmol/L for a periodate-derived GAG and 527±70 µmol/L for a heparinase-derived GAG. At the same MW, the agglutination IC₅₀ was 17±5 µmol/L for periodate and 135±18 µmol/L for heparinase. This suggests that the disaccharide GlcNS[6S]-IdoA2S, destroyed by heparinase but not periodate, is crucial to heparin-vWF interactions. An MW dependency was also noted, with a minimum dodecasaccharide required for activity inhibition. To further investigate the heparin/vWF interaction, affinity fractionation of heparins was performed with an immobilized peptide derived from a heparin-binding domain of vWF. Disaccharide analysis of high-affinity heparins revealed an increased ratio of IdoA2S-GlcN[S/Ac]6S to IdoA2S-GlcN[S/Ac]. Affinity fractionation of oligosaccharides (MW 3500) diminished the relative content of all disaccharides except IdoA2S-GlcNS6S, which was increased. These data suggest that the disaccharide structures IdoA2S-GlcNS6S and GlcNS6S-IdoA2S are crucial to heparin/vWF interactions. Understanding the structural aspects that confer such activity may be useful in designing heparin-based antithrombotic drugs.

Key Words: heparin • von Willebrand factor • platelets • thrombosis • platelet aggregation

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