CONFORMATION OF MICELLAR PHOSPHOLIPID BOUND TO THE ACTIVE-SITE OF PHOSPHOLIPASE A(2)

Transferred NOE techniques have been used to determine the structure o f phospholipid analogues bound to the active site of cobra venom phosp holipase A(2) (PLA(2)). These experiments were carried out on PLA(2) w ith a substrate analogue which serves as an inhibitor, noylamino)-1,2- dideoxy-sn-glycero-3-phosphocholine (PC9). Because this inhibitor bind s tightly to the enzyme and forms micelles at millimolar concentration s, experiments could be carried out to determine the conformation of t he inhibitor when bound to the enzyme at the lipid-water interface. NO Es of the micellar lipid develop inefficiently in the absence of enzym e. NOESY experiments in the presence of PLA(2) were used to determine the inhibitor structure and conformation when bound to the enzyme. The inhibitor adopts an active site conformation in which the end of the sn-2 chain is within 5 Angstrom of the alpha-methylene protons of the sn-1 chain. However, NOE cross-peaks in the experiments indicate that the backbone conformation of the bound lipid is different from that of a shorter chain lipid which forms monomers [Plesniak et al. (1993) Bi ochemistry 32, 5009-5016].