Specific inhibition of glucokinase by long chain acyl coenzymes A below the critical micelle concentration.

Rat liver glucokinase (EC 2.7.1.2) is inhibited specifically by the free form of palmitoyl-CoA (Ki apparent of approximately 1.8 PM) and other related long chain fatty acyl-CoAs under conditions where the micellar form of these compounds does not exist. This inhibition is instantaneous, and is immediately reversible by dilution, bovine serum albumin, and aor B-cyclodextrins. Inhibition is specific for long chain fatty acylCoAs. Related compounds which also form micelles, palmitate and myristate, as well as the structurally related compounds, acetyl-coA and CoASH, are not inhibitory. The extent of inhibition is independent of the molar ratio of glucokinase to acyl-CoA. The inhibition is competitive with both ATP and glucose and does not affect the positive cooperativity which glucokinase normally displays with glucose (Hill coefficient = 1.5 under all inhibited conditions). The critical micelle concentrations (cmc) of oleoylCoA and the saturated long chain acyl-CoAs (C-12 to C-20) were determined under actual assay conditions by a pinacyanol chloride dye binding method. The cmc for palmitoyl-CoA under several conditions was also estimated by ultracentrifugation and ultrafiltration. Significant inhibition by these compounds was found to occur well below their cmc values in all cases. The apparent Ki for palmitoyl-CoA, for example, was between 2.5and 14-fold below its cmc in assay mixtures lacking glycerol and between 10and 52-fold below its cmc in mixtures containing 30% glycerol. The reversible, instantaneous, specific, and sensitive nature of this inhibition, coupled with the lack of micelle formation at inhibitory concentrations of these long chain acylCoAs, suggest that inhibition of glucokinase is not related to detergent effects but rather represents the binding of monomeric acyl-CoA to the enzyme.