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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2008;28:329.)
© 2008 American Heart Association, Inc.

Cell Biology/Signaling

Thrombin Mutant W215A/E217A Acts as a Platelet GPIb Antagonist

Michelle A. Berny; Tara C. White; Erik I. Tucker; Leslie A. Bush-Pelc; Enrico Di Cera; András Gruber; Owen J.T. McCarty

From the Departments of Biomedical Engineering (M.A.B., T.C.W., E.I.T., A.G., O.J.T.M.) and Cell and Developmental Biology (O.J.T.M.), Oregon Health and Science University, Portland; and Biochemistry and Molecular Biophysics (L.A.B.-P., E.D.C.), Washington University, St. Louis, Mo.

Correspondence to Owen J.T. McCarty, Department of Biomedical Engineering, Oregon Health & Science University, 3303 SW Bond Ave, Portland, OR 97239. E-mail mccartyo{at}ohsu.edu

Abstract

Objective— Thrombin containing the mutations Trp215Ala and Glu217Ala (WE) selectively activates protein C and has potent antithrombotic effects in primates. The aim of this study was to delineate the molecular mechanism of direct WE–platelet interactions under static and shear conditions.

Methods and Results— Purified platelets under static conditions bound and spread on immobilized wild-type but not WE thrombin. In PPACK-anticoagulated blood under shear flow conditions, platelets tethered and rolled on both wild-type and WE thrombin, and these interactions were abrogated by the presence of a glycoprotein Ib (GPIb)-blocking antibody. Platelet deposition on collagen was blocked in the presence of WE, but not wild-type thrombin or prothrombin. WE also abrogated platelet tethering and rolling on immobilized von Willebrand factor in whole blood under shear flow.

Conclusions— These observations demonstrate that the thrombin mutant WE, while not activating platelets, retains the ability to interact with platelets through GPIb, and inhibits GPIb-dependent binding to von Willebrand factor–collagen under shear.

The thrombin mutant WE selectively activates protein C and has potent antithrombotic effects in primates. We demonstrate that WE, while not activating platelets, retains the ability to interact with platelets through GPIb, and inhibits GPIb-dependent binding to vWF-collagen under shear.

Key Words: platelet • thrombin • von Willebrand factor • GPIb

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New Direction for WE Thrombin

Michael C. Berndt and Robert K. Andrews
Arterioscler. Thromb. Vasc. Biol. 2008 28: 205-207. [Full Text] [PDF]

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