Competitive inhibition of lipolytic enzymes. IX. A comparative study on the inhibition of pancreatic phospholipases A2 from different sources by (R)-2-acylamino phospholipid analogues.

The inhibitory power (Z) of a number of (R)-1-alkyl-2-acylamino phospholipid analogues was determined for three mammalian phospholipases A2 from pig, ox and horse pancreas. All three enzymes display a clear preference for anionic (phosphoglycol) inhibitors over the zwitterionic (phosphocholine) derivatives; this effect is most pronounced for the bovine enzyme. Upon variation of the 1-alkyl chain length, the bovine and equine phospholipases, like the porcine enzyme in previous studies, show an optimum in Z for a six-carbon alkyl group. The introduction of a double bond in the 2-acylamino group generally improves the inhibitory power as compared with a fully saturated acyl chain. For the horse enzyme, the presence of an (R)-2-undecenoylamino group in the phosphocholine- and phosphoglycol-containing inhibitors resulted in affinities which are nearly 4 and 5 orders of magnitude higher, respectively, than for the substrate molecule. Direct determination of the dissociation constant Ki* of several inhibitors incorporated in a host lipid/water interface of non-inhibitory n-octadecenylphosphocholine micelles, was performed by ultraviolet difference spectroscopy. The progressive binding of a single inhibitor molecule into the active site of the three enzymes was followed quantitatively by an increasing tyrosine perturbation. With moderately strong competitive inhibitors (Z values ranging from about 50 to 10,000), quantitative values for Ki* were obtained. Extrapolation of the experimentally found linear relationship between Z and 1/Ki* yields predicted Ki* numbers for the much stronger inhibitors with Z values between 10,000 and 100,000.