Fine Mapping of Inhibitory Anti-factor V Antibodies Using Factor V C2 Domain Mutants

Summary

Hemorrhagic factor V inhibitors frequently bind to the second C-type (C2) domain of factor V and interfere with phospholipid binding. To define specific residues recognized by inhibitors from four patients (one bovine thrombin-induced and three spontaneous antibodies), epitope mapping was performed using recombinant human factor V lacking most of the B-type domain (FV des B) and alanine-substituted mutants within the C2 domain (FV des B C2 mutants). FV des B C2 mutants located in the region between Lys²⁰⁶⁰ and Glu²⁰⁶⁹ were resistant to inhibition by three IgG preparations including the bovine thrombin-induced antibody in both prothrombinase and phospholipid-binding assays. In contrast, mutations at Lys²⁰⁸⁷ and Lys²⁰⁹²/ Glu²⁰⁹⁶ were significantly resistant to inhibition by the fourth IgG preparation in both prothrombinase and phospholipid-binding assays. These results confirm interference of phospholipid binding by hemorrhagic factor V inhibitors and support the role(s) of these residues in phospholipid binding.

Presented in part at the 41st Annual Meeting of the American Society of Hematology, New Orleans, Louisiana, December 3–7, 1999

Key words

Factor V - hemorrhagic inhibitor - epitope mapping - C2 domain - phospholipid binding

^* Department of Clinical Laboratory Science, College of Health Sciences, Korea University, Korea;

^† Instituto de Biologia Molecular de Barcelona, CSIC, Spain

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