Exogenously added human group X secreted phospholipase A(2) but not the group IB, IIA, and V enzymes efficiently release arachidonic acid from adherent mammalian cells

Mammalian secreted phospholipases A(2) (sPLA2s) comprise a group of at leas t eight enzymes, including the recently identified group X sPLA2, A bacteri al expression system was developed to produce human group X sPLA2 (hGX). In hibition studies show that the sPLA2 inhibitor LY311727 binds modestly more tightly to human group IIA sPLA2 than to hGX and that a pyrazole-based inh ibitor of group IIA sPLA2 is much less active against hGX. The phospholipid head group preference of vesicle-bound hGX was determined, hGX binds tight ly to phosphatidylcholine vesicles, which is thought to be required to act efficiently on cells. Tryptophan 67 hGX makes a significant contribution to interfacial binding to zwitterionic vesicles. As little as 10 ng/ml hGX re leases arachidonic acid for cyclooxygenase-2-dependent prostaglandin E-2 ge neration when added exogenously to adherent mammalian cells. In contrast, h uman group IIA, rat group V, and mouse group IB sPLA2s are virtually inacti ve at releasing arachidonate when added exogenously to adherent cells. Disl odging cells from the growth surface enhances the ability of all the sPLA2s to release fatty acids. Studies with CHO-K1 cell mutants show that binding of sPLA2s to glycosaminoglycans is not the basis for poor plasma membrane hydrolysis by group IB, IIA, and V sPLA2s.