had a diauxic pattern of glucose and lactose utilization, and both of these sugars were transported by the sugar phosphotransferase system (PTS). Lactose catabolism was inducible, and used the tagatose pathway to ferment lactose. Since a mutant that was deficient in glucose PTS activity transported lactose as fast as the wild-type, it appeared that has separate enzyme lls for glucose and lactose. The nonmetabolizable glucose analogue 2-deoxyglucose (2-DG) was a noncompetitive inhibitor of methyl β-D-thiogalactopyranoside (TMG) transport, and cells that were provided with either glucose or 2-DG were unable to transport TMG or lactose. Because the glucose-PTS-deficient mutant could ferment glucose, but could not exclude TMG, it appeared that enzyme II rather than glucose catabolism se was the critical feature of inducer exclusion. Cells that had accumulated TMG as TMG 6-phosphate expelled free TMG when glucose was added, but 2-DG was unable to cause TMG expulsion. The glucose-PTS-deficient mutant could still expel TMG in the presence of exogenous glucose. Membrane vesicles also exhibited glucose-dependent TMG exclusion and TMG expulsion. Membrane vesicles that were electroporated with phosphoenolpyruvate (PEP) and HPr retained TMG for more than 3 min, but vesicles that were electroporated with PEP plus HPr and fructose 1,6-diphosphate (FDP) (or glycerate 2-phosphate) lost their ability to retain TMG. Because FDP was able to trigger the ATP-dependent phosphorylation of HPr, it appeared that inducer expulsion was mediated by an FDP-activated protein kinase. This conclusion was further supported by the observation that mutant forms of HPr differed in their ability to faciliate inducer expulsion. S46DHPr, a mutant HPr with aspartate substituted for serine at position 46, promoted TMG expulsion from membrane vesicles in the absence of FDP better than wild-type HPr or S46AHP a mutant form with alanine substituted for serine at position 46. Based on these results, it appeared that glucose catabolism was needed for inducer expulsion, but not inducer exclusion.


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