title: Protein S is a cofactor for activated protein C neutralization of an inhibitor of plasminogen activation released from platelets.
abstract: Platelets stimulated with thrombin release an inhibitor of plasminogen activator (PAI), which has been shown previously to be neutralized by activated protein C (APC). The requirements for optimal neutralization of PAI activity were investigated. The releasate of gel-filtered human platelets stimulated with thrombin served as a source of PAI. When 6 X 10(8) platelets/mL were incubated with thrombin (1 IU/mL), the releasate contained 18 to 26 ng/mL PAI as determined by incubation of the releasate with urokinase and measurement of residual urokinase activity on plasminogen (S2251). Preincubation of PAI with up to 4 micrograms/mL APC for two hours yielded less than 20% neutralization of PAI activity. In the presence of protein S, phospholipid, and Ca2+, neutralization of PAI activity was time-dependent with 50% neutralization occurring in two hours with 1 microgram/mL APC. The cofactor effects of protein S and phospholipid were concentration-dependent with half-maximal acceleration at approximately 3 micrograms/mL protein S and 10 micrograms/mL phospholipid when the experiments were performed at 1 microgram/mL APC. Diisopropylfluorophosphate-inactivated APC, gla-domainless APC, and thrombin-cleaved protein S had no effect on PAI activity, indicating requirement for preservation of the APC active site and of the Ca2+ binding ability of both APC and protein S. These results suggest coordinate binding of APC and protein S onto phospholipid membrane as a prerequisite for optimal expression of PAI neutralized by APC.
mesh_terms: D001777:Blood Coagulation; D001792:Blood Platelets / Q000502:physiology*; D004789:Enzyme Activation / Q000187:drug effects; D006023:Glycoproteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D006801:Humans; D010743:Phospholipids / Q000502:physiology; D010958:Plasminogen / Q000378:metabolism*; D015849:Plasminogen Inactivators; D011486:Protein C / Q000502:physiology*; D017293:Protein S; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors*
chemical_list: D006023:Glycoproteins; D010743:Phospholipids; D015849:Plasminogen Inactivators; D011486:Protein C; D017293:Protein S; D010958:Plasminogen; D014568:Urokinase-Type Plasminogen Activator