Evidence for the participation of independent translocation for phosphate and glucose 6-phosphate in the microsomal glucose-6-phosphatase system. Interactions of the system with orthophosphate, inorganic pyrophosphate, and carbamyl phosphate

• Published in: The Journal of biological chemistry Vol. 255; no. 21; pp. 10396 - 10406
• Main Authors: Arion, W J, Lange, A J, Walls, H E, Ballas, L M
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 10.11.1980
• Subjects: Animals; Antiporters; Carbamyl Phosphate - pharmacology; Diphosphates - pharmacology; Glucose - pharmacology; Glucose-6-Phosphatase - metabolism; Intracellular Membranes - enzymology; Kinetics; Microsomes, Liver - enzymology; Monosaccharide Transport Proteins; Multienzyme Complexes - metabolism; Phosphates - pharmacology; Phosphotransferases - metabolism; Rats
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)70478-5

The interactions of Pi, PPi, and carbamyl-P with the hepatic glucose-6-phosphatase system were studied in intact and detergent-disrupted microsomes. Penetration of PPi and carbamyl-P into intact microsomes was evidenced by their reactions with the enzyme located exclusively on the luminal surface. Lack of effects of carbonyl cyanide m-chlorophenylhydrazone and valinomycin + KCl indicated that pH gradients and/or membrane potentials that could influence the kinetics of the system are not generated during metabolism of PPi and glucose-6-P by intact microsomes. With disrupted microsomes, only competitive interactions were seen among glucose-6-P, Pi, PPi, and carbamyl-P. With intact microsomes, Pi, PPi, and carbamyl-P were relatively weak, noncompetitive inhibitors of glucose-6-phosphatase, and PPi hydrolysis was inhibited competitively by Pi and carbamyl-P but noncompetitively by glucose-6-P. Analysis of the kinetic data in combination with findings from other studies that a variety of inhibitors of the glucose-6-P translocase (T1) does not affect PPi hydrolysis provide compelling evidence that permeability of microsomes to Pi, PPi, and carbamyl-P is mediated by a second translocase (T2). Some properties of the microsomal anion transporters are described. If the characteristics of the glucose-6-phosphatase system as presently defined in intact microsomes apply in vivo, glucose-6-P hydrolysis appears to be the predominant, if not the exclusive, physiologic function of the system. Both the "noncompetitive character" and the relative ineffectiveness of Pi as an inhibitor of glucose-6-phosphatase of intact microsomes result from the rate limitation imposed by T1 that prevents equilibration of glucose-6-P across the membrane. In microsomes from fed rats, where T1 is less rate restricting, about one-half as much Pi was required to give 50% inhibition compared with microsomes from fasted or diabetic rats. Thus, any treatment or agent that alters the kinetic relationship between transport and hydrolysis of glucose-6-P (e.g. endocrine or nutritional status) is an essential consideration in analyses of kinetic data for the glucose-6-phosphatase system.