Utilization of gluconate by Escherichia coli. Uptake of D-gluconate by a mutant impaired in gluconate kinase activity and by membrane vesicles derived therefrom.
1. From Escherichia coli strain K2.1.5(c).8.9, which is devoid of 6-phosphogluconate dehydrogenase (gnd) and 6-phosphogluconate dehydratase (edd) activities, a mutant R6 was isolated that was tolerant to gluconate though still edd(-), gnd(-). 2. Measurements of the fate of labelled gluconate, of the conversion of gluconate into 6-phosphogluconate, and of the induction of gluconate kinase by the two organisms show that, although both inducibly form a gluconate-transport system, strain R6 is impaired in its ability to convert the gluconate thus taken up into 6-phosphogluconate; it was therefore used for study of the kinetics and energetics of gluconate uptake. 3. Suspensions of strain R6 induced for gluconate uptake took up this substrate via a ;high affinity' transport process, with K(m) about 10mum and V(max.) about 25nmol/min per mg dry mass; a ;low affinity' system demonstrated to occur in certain E. coli mutants was not induced under the conditions used in this work. 4. The uptake of gluconate was inhibited by lack of oxygen and by inhibitors of electron transport; such inhibitors also promoted the efflux of gluconate taken up. 5. Membrane vesicles prepared from strain R6 also manifested these properties when incubated with suitable electron donors, at rates similar to those observed with whole cells. 6. The results indicate that the active transport of gluconate into the cells is the rate-limiting step in gluconate utilization by E. coli, and that the mechanism of this process can be validly studied with membrane vesicles.
D000654:Amobarbital; D001151:Arsenic / Q000494:pharmacology; D001386:Azides / Q000494:pharmacology; D001692:Biological Transport; D002250:Carbon Radioisotopes; D002498:Centrifugation; D003486:Cyanides / Q000494:pharmacology; D004140:Dinitrophenols / Q000494:pharmacology; D004579:Electron Transport / Q000187:drug effects; D004926:Escherichia coli / Q000378:metabolism*; D005942:Gluconates / Q000378:metabolism*; D007700:Kinetics; D008566:Membranes / Q000378:metabolism; D009154:Mutation; D010101:Oxygen Consumption; D010734:Phosphogluconate Dehydrogenase / Q000378:metabolism; D010770:Phosphotransferases / Q000378:metabolism*; D013696:Temperature
D001386:Azides; D002250:Carbon Radioisotopes; D003486:Cyanides; D004140:Dinitrophenols; D005942:Gluconates; D010734:Phosphogluconate Dehydrogenase; D010770:Phosphotransferases; D000654:Amobarbital; D001151:Arsenic