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.

This research article investigates the inhibitory effects of certain analogues on mammalian phospholipases A2 enzymes from pig, ox, and horse pancreas. The study revealed surprising affinity differences and highlighted optimal conditions for inhibitory power.

Inhibition of Pancreatic Phospholipases A2

The study began by testing the inhibitory power, denoted by “Z”, of several (R)-1-alkyl-2-acylamino phospholipid analogues on three mammalian phospholipases A2. These enzymes were sourced from the pancreas of pigs, oxen, and horses. It was observed that:

• All three studied enzymes showed a clear preference for anionic (phosphoglycol) inhibitors over the zwitterionic (phosphocholine) derivatives.
• This preference was most significant in the bovine (oxen) enzyme.

Variation of the 1-Alkyl Chain Length

Upon variation of the 1-alkyl chain length, important observations were noted:

• The bovine and equine (horse) phospholipases, in line with the porcine (pig) enzyme in previous studies, displayed an optimal Z at a six-carbon alkyl group.
• The introduction of a double bond in the 2-acylamino group typically improved inhibitory power compared with a fully saturated acyl chain.

Results for the Horse Enzyme

The horse enzyme yielded some fascinating results:

• The presence of an (R)-2-undecenoylamino group in the phosphocholine- and phosphoglycol-containing inhibitors resulted in affinities that were nearly 4 and 5 orders of magnitude higher, respectively, than the substrate molecule.

Dissociation Constant Ki* Determination

The dissociation constant Ki* of several inhibitors, incorporated in a host lipid/water interface of non-inhibitory n-octadecenylphosphocholine micelles, was directly determined using ultraviolet difference spectroscopy. During the process:

• 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.

By extrapolating the experimentally found linear relationship between Z and 1/Ki*, the researchers were able to make predictions about the Ki* numbers for much stronger inhibitors with Z values between 10,000 and 100,000.