An allosteric model for the inhibition of glucokinase by long chain acyl coenzyme A.

Rat liver glucokinase is specifically inhibited by palmitoyl-CoA and other long chain acyl-CoAs at concentrations well below their critical micelle concentrations (Tippett, P. S., and Neet, K. E. (1982) J. Biol. Chem. 257, 12839-12845). Kinetic studies of this inhibition indicate that the long chain acyl-CoAs act on glucokinase through an allosteric site. The inhibition is competitive with both glucose and MgATP at either saturating or half-maximal fixed substrate concentrations and the apparent Ki is the same (1.8 PM for palmitoyl-CoA) with respect to either substrate. The positive cooperativity which glucokinase normally displays with glucose is unaffected by the inhibition. Palmitoyl-CoA (as well as the other long chain acyl-CoAs) inhibition is itself positively cooperative, yet this cooperativity is unchanged (Hill coefficient = 1.78 k 0.3 (n = 32)) over a full range of concentrations of either substrate. Inhibition by arachidoyl-CoA in an assay mix containing low concentrations of free magnesium did not continue toward zero activity with increasing concentrations of inhibitor but instead reached a limiting value. This leveling off was not due to the formation of micelles, since the nature of the inhibition was unchanged even after the critical micelle concentration of arachidoyl-CoA was raised &fold in the assay mixture by the addition of 30% glycerol. Data for this limited inhibition for arachidoyl-CoA in the presence of either low M&+ concentrations or a high M&+ concentration assay mixture fit an equation for partial (allosteric) competitive inhibition. An allosteric model is proposed which assumes that an acyl-CoA binds glucokinase at an allosteric site, causes a conformational change (the extent of which is dependent on the particular ligand), and lowers the binding affinity (by a factor a) for both glucose and ATP with no effect on the catalytic apacity (Vmax). The inhibition of glucokinase by all of the long chain acyl-CoAs tested (saturated, even number of carbons from C-12 to C-20, and oleoyl-CoA) under all conditions can be explained by this allosteric model. The effect of 5 mM free M 8 ’ on the inhibition of palmitoylCoA and arachidoyl-CoA (and the other acyl-CoAs) is interpreted as lowering a, Le. decreasing the extent of conformational change induced by the acyl-CoA but with little effect on the Ki itself. Finally, in keeping with predictions based on this model, low concentrations of arachidoyl-CoA, normally an inhibitor with an a of