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- Title
Novel arguments in favor of the substrate-transport model of glucose-6-phosphatase.
- Authors
Gerin, Isabelle; Noel, Gaetane; Van Schaftingen, Emile; Gerin, I; Noël, G; Van Schaftingen, E
- Abstract
The purpose of this work was to discriminate between two models for glucose-6-phosphatase: one in which the enzyme has its catalytic site oriented toward the lumen of the endoplasmic reticulum, requiring transporters for glucose-6-phosphate, inorganic phosphate (Pi), and glucose (substrate-transport model), and a second one in which the hydrolysis of glucose-6-phosphate occurs inside the membrane (conformational model). We show that microsomes preloaded with yeast phosphoglucose isomerase catalyzed the detritiation of [2-(3)H]glucose-6-phosphate and that this reaction was inhibited by up to 90% by S3483, a compound known to inhibit glucose-6-phosphate hydrolysis in intact but not in detergent-treated microsomes. These results indicate that glucose-6-phosphate is transported to the lumen of the microsomes in an S3483-sensitive manner. Detritiation by intramicrosomal phosphoglucose isomerase was stimulated twofold by 1 mmol/l vanadate, a phosphatase inhibitor, indicating that glucose-6-phosphatase and the isomerase compete for the same intravesicular pool of glucose-6-phosphate. To investigate the site of release of Pi from glucose-6-phosphate, we incubated microsomes with Pb(2+), which forms an insoluble complex with Pi, preventing its rapid exit from the microsomes. Under these conditions, approximately 80% of the Pi that was formed after 5 min was intramicrosomal, compared with <10% in the absence of Pb(2+). We also show that, when incubated with glucose-6-phosphate and mannitol, glucose-6-phosphatase formed mannitol-1-phosphate and that this nonphysiological product was initially present within the microsomes before being released to the medium. These results indicate that the primary site of product release by glucose-6-phosphatase is the lumen of the endoplasmic reticulum.
- Subjects
GLUCOSE-6-phosphatase; ENDOPLASMIC reticulum
- Publication
Diabetes, 2001, Vol 50, Issue 7, p1531
- ISSN
0012-1797
- Publication type
journal article
- DOI
10.2337/diabetes.50.7.1531