Fine mapping of inhibitory anti-factor V antibodies using factor V C2 domain mutants. Identification of two antigenic epitopes involved in phospholipid binding.
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.
Izumi
T
T
Department of Medicine, Duke University Medical Center, Durham, North Carolina, USA.
Kim
S W
SW
Greist
A
A
Macedo-Ribeiro
S
S
Fuentes-Prior
P
P
Bode
W
W
Kane
W H
WH
Ortel
T L
TL
D000368:Aged; D000906:Antibodies / Q000276:immunology* / Q000494:pharmacology; D001665:Binding Sites; D001780:Blood Coagulation Tests; D018604:Epitope Mapping / Q000379:methods*; D005165:Factor V / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D006470:Hemorrhage / Q000209:etiology; D006801:Humans; D008297:Male; D008567:Membranes, Artificial; D009154:Mutation / Q000276:immunology*; D010743:Phospholipids / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000235:genetics; D013925:Thromboplastin
D000906:Antibodies; D008567:Membranes, Artificial; D010743:Phospholipids; D005165:Factor V; D013925:Thromboplastin