Date of Award

7-2-1989

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biochemistry and Molecular Biology

Abstract

Glucose-6-phosphatase (EC 3.1.3.9) occupies a strategic metabolic location in hepatic glucose metabolism, is multifunctional in nature, consists of multiple components, and exhibits unique kinetic characteristics. These properties of glucose-6-phosphatase make it a likely candidate for regulation of glucose metabolism.The effect of selected intermediate metabolites on the phosphotransferase and phosphohydrolase activities of glucose-6-phosphatase was investigated with intact and detergent-disrupted microsomal preparations. Detergent treatment exposed the catalytic unit of microsomal preparations. The interaction of the metabolite with the catalytic unit was assessed in these preparations. The study of metabolites with the glucose-6-phosphatase of intact microsomes served to characterize the interactions of the metabolites with the substrate transport system as well as the catalytic unit.The metabolites investigated were: fructose-6-P, glucose-1-P, fructose-1,6-diphosphate, 2-phosphoglycerate, 3-phosphoglycerate, glyceraldehyde-3-P, dihydroxyacetone-P, pyruvate, fructose-1-P, phosphoenolpyruvate, glucose-1,6-diphosphate, NAD$\sp+$, NADH, uridine 5$\sp\prime$-diphosphoglucose, lactate, ribose-5-P, NADP$\sp+$, NADPH, 6-phosphogluconic acid, L-leucine, L-glutamine, L-nor-leucine, L-nor-valine, L-alanine, L-asparagine, a combination of L-glutamine + L-leucine, a combination of L-glutamine + L-nor-leucine, a combination of L-glutamine + L-nor-valine, glucuronate, GDP-mannose, inositol, oxaloacetic acid, malate, carbamyl aspartic acid, fructose-2,6-bisphosphate, and arachidonate.Kinetic analysis suggested arachidonate and fructose-1-P as possible regulatory metabolites controlling glucose-6-phosphatase. The inhibition of glucose-6-phosphatase activity by fructose-1-P implied a link between the regulation of glucose metabolism, especially glycogen synthesis, and fructose metabolism. The increased fructose-1-P concentration as a result of a fructose load would cause an increase in glucose-6-P concentration, by inhibition of glucose-6-phosphatase, and thus stimulate glycogen synthesis.Arachidonate exhibited inhibition similar to competitive inhibition of glucose-6-P hydrolysis by disrupted microsomal preparations. Kinetics were characteristic of two molecules of arachidonate binding to one molecule of the enzyme of intact microsomal preparations. The inhibition of glucose-6-phosphatase activity by arachidonate implied that control of arachidonate release could be involved in the regulation of glucose-6-phosphatase activity.

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