Binding of protein S to factor Va associated with inhibition of prothrombinase that is independent of activated protein C.
Since plasma protein S serves an anticoagulant function by mechanisms which are not completely understood, its possible interaction with Factor Va was investigated. Human protein S bound to immobilized human Factor Va in a calcium-dependent, saturable, and reversible manner and Factor Va bound similarly to immobilized protein S. Binding of protein S to immobilized Factor V was greatly enhanced by pretreatment of the surface-bound Factor V with increasing doses of thrombin up to 1 unit/ml. Binding of protein S to Factor Va was also demonstrated in fluid phase with a Kd of 33 +/- 9 nM. Biotin-labeled heavy chain of Factor Va bound to immobilized protein S, and this binding was reversed by a 17-fold molar excess of intact unlabeled Factor Va. Protein S competed efficiently with prothrombin for binding to immobilized Factor Va. The prothrombinase activity in a reaction mixture of purified clotting factors was inhibited by protein S and exhibited a pattern of mixed inhibition. The concentration of protein S needed for 50% inhibition of the prothrombinase activity of a mixture containing 1 nM Factor Xa, 20 pM Factor Va, and 50 microM phospholipids was about 16 nM. Since not all protein S preparations exhibited this degree of prothrombinase inhibitory activity, extensive control experiments were performed to verify that the inhibitory activity was associated with protein S during immunoaffinity chromatography and was not caused by traces of activated protein C in the protein S preparations. These data show that protein S has an anticoagulant function which is independent of activated protein C and, at least in part, that this is because of its competition with prothrombin for direct binding to Factor Va.
Heeb
M J
MJ
Committee on Vascular Biology, Scripps Research Institute, La Jolla, California 92037.
Mesters
R M
RM
Tans
G
G
Rosing
J
J
Griffin
J H
JH
D001667:Binding, Competitive; D001777:Blood Coagulation; D004789:Enzyme Activation; D005165:Factor V / Q000378:metabolism; D015943:Factor Va / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D011486:Protein C / Q000378:metabolism*; D017293:Protein S / Q000378:metabolism*; D011516:Prothrombin / Q000378:metabolism; D012995:Solubility; D013925:Thromboplastin / Q000037:antagonists & inhibitors*
D011486:Protein C; D017293:Protein S; D015943:Factor Va; D005165:Factor V; D011516:Prothrombin; D013925:Thromboplastin