Inhibitors of glucose-1,6-bisphosphate synthase.

Abstract

Glucose-l&P, synthase of brain was sensitive to a number of inhibitors with their associated K, values acting competitively with glycerate-1,3-P,: fructose-1,6-P, (0.3 PM), glucose-1,6-P,(5 PM), glycerate-2,3-P, (15 PM), enolpyru-Vate-P (30 PM), citrate (60 PM), &Ct?rat&-P (150 PM), and Pi (400 PM).Most of these compounds are reported to be at concentrations in whole brain that would be very inhibitory to the enzyine.However, some of them, such as fructose-1,6-P2 and citrate which are known effecters of several important rate-determining enzymes of glycolysis, gluconeogenesis, and lipogenesis, change depending on the physiological state in significant manner.These two inhibitors may act at allosteric sites in view of the very high affinity of the fructose-1,6-P, compared with the other phosphate compounds and the unusual specificity of the tricarboxylic acid.Citrate inhibition was competitive for both substrates, glucose-l-P and glycerate-1,3-P,, when the enzyme was in the Zn*+ form but only for glycerate-1,3-P, when in the Mg*+ form.Lithium was a strong inhibitor (K, = 50 FM) competitive with the cofactor Mg2+ (K,, -1 mM) when Mg2+ was the activating metal.Zn*+, unlike Li+ and Mg2+, is not readily dissociated from the enzyme.Therefore, Li+ does not readily inhibit the Zn*+ enzyme.On the basis of the insensitivity of synthase activity of fresh brain homogenate to Li+ and EDTA, it is suggested that about 65% of the synthase was in the Zn2+ form.On the basis of its strong regulatory effects on hexokinase, phosphofructokinase, pyruvate kinase, its significant concentrations in many tissues, and the sensitive control of its synthesis by regulatory metabolites, it seems that glucose-1,6-P, should be considered a regulatory metabolite of considerable significance.Chemical and kinetic studies (1, 2) indicate that in the synthesis of glucose-1,6-P, by the purified brain synthase the enzyme is first phosphorylated at a serine residue in an essentially irreversible reaction, and that the phosphoryl group is transferred to glucose-l-P in a somewhat more rapid step:Glycerate-1,3-P, + E 4 E-P + glycerate-3-P E-P + glucose-l-P + E + glucose-1,6-P,