Basic residues of human group IIA phospholipase A(2) are important for binding to factor Xa and prothrombinase inhibition - Comparison with other mammalian secreted phospholipases A(2)

Human secreted group IIA phospholipase A(2) (hGIIA) was reported to inhibit prothrombinase activity because of binding to factor Xa. This study furthe r shows that hGIIA and its catalytically inactive H48Q mutant prolong the l ag time of thrombin generation in human platelet-rich plasma with similar e fficiency, indicating that hGIIA exerts an anticoagulant effect independent ly of phospholipid hydrolysis under ex vivo conditions. Charge reversal of basic residues on the interfacial binding surface (IBS) of hGIIA leads to d ecreased ability to inhibit prothrombinase activity, which correlates with a reduced affinity for factor Xa, as determined by surface plasmon resonanc e. Mutation of other surface-exposed basic residues, hydrophobic residues o n the IBS, and His48, does not affect the ability of hGIIA to inhibit proth rombinase activity and bind to factor Xa. Other basic, but not neutral or a cidic, mammalian secreted phospholipases A(2) (sPLA(2)s) exert a phospholip id-independent inhibitory effect on prothrombinase activity, suggesting tha t these basic sPLA(2)s also bind to factor Xa. In conclusion, this study de monstrates that the anticoagulant effect of hGIIA is independent of phospho lipid hydrolysis and is based on its interaction with factor Xa, leading to prothrombinase inhibition, even under ex vivo conditions. This study also shows that such an interaction involves basic residues located on the IBS o f hGIIA, and suggests that other basic mammalian sPLA(2)s may also inhibit blood coagulation by a similar mechanism to that described for hGIIA.