The effect of fructose on pyruvate kinase activity in isolated hepatocytes. Inhibition by allantoin and alanine

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Abstract
Incubation of isolated [rat] hepatocytes with fructose at concentrations above 3 mM resulted in an apparent inhibition of pyruvate kinase assayed in crude extracts at sub-optimal phosphoenolpyruvate concentrations. Fructose at concentrations below 3 mM caused an activation of the enzyme. Increases in the hepatocyte contents of the positive effectors fructose 1,6-bisphosphate and fructose 1-phosphate were found at all concentrations of fructose up to 10 mM. Removal of the extrahepatocyte medium from the hepatocytes by washing resulted in an activation of the enzyme at all concentrations of fructose examined. Inhibitors of the enzyme accumulated in the hepatocytes depsite the depletion of ATP (a known negative effector) caused by higher concentrations of fructose. Indeed the inhibition of pyruvate kinase appeared to be correlated to the depletion of ATP. Alanine (a known inhibitor) accumulated in hepatocytes as a consequence of incubation with fructose. Allantoin and uric acid were inhibitors of a partially purified pyruvate kinase preparation assayed both in the presence and in the absence of fructose 1,6-bisphosphate. Allantoin, but not uric acid, accumulated in the extrahepatocyte medium as a result of incubation of the cells with 10 mM-fructose.