Mechanisms of Active Transport in Isolated Bacterial Membrane Vesicles

The role of functional sulfhydryl groups in active transport in Escherichia coli membrane vesicles

“…NEM Inactivation of Active Transport. Although D-lactate-dependent accumulation of many solutes is inhibited by NEM, p-HMB, and p-HMBS (Kaback and Barnes, 1971;Kerwar et al, 1972;Gordon et al, 1972;Lombardi and Kaback, 1972), as noted by Kaback and Barnes (1971), this observation does not provide unequivocal evidence that the porters require functional sulfhydryl groups, because D-lactate figure 4: Effect of dithiothreitol on lactose transport and NEM reactivity in diamide-treated vesicles. Two aliquots (1.0 mL each) of membrane vesicles containing about 8.0 mg of membrane protein were diluted to a final volume of 2.0 mL containing, in final concentrations, 0.05 M potassium phosphate (pH 6.6) and 0.01 M magnesium sulfate.…”

“…In contrast to sulfhydryl reagents such as NEM, p-hydroxymercuribenzoate (p-HMB), and p-hydroxymercuribenzenesulfonate (p-HMBS) which inactivate both D-lactate oxidation and transport (Barnes and Kaback, 1971; Kaback and Barnes, 1971;Kerwar et al, 1972;Gordon et al, 1972; Lombardi and Kaback, 1972), diamide has essentially no effect on D-lactate oxidation or on the ability of NEM (Table I) or p-HMBS (not shown) to block D-lactate oxidation. Importantly, moreover, neither NEM nor p-HMBS inhibits the " Oxygen uptake was measured with a Clark oxygen electrode as described previously (Barnes and Kaback, 1971).…”

Photoaffinity inhibition of dipeptide transport in Escherichia coli

“…3). Arginine transport was chosen as the control because arginine, like dipeptides, is transported exclusively via a shocksensitive transport system (Lombardi & Kaback, 1972) and is therefore likely to be metabolically coupled in the same way as is dipeptide transport. The observed photoinactivation of dipeptide transport is therefore likely to be due to an effect on the transport system per se rather than to some light-dependent metabolic uncoupling.…”

Properties and characterization of binding protein dependent active transport of glutamine in isolated membrane vesicles of Escherichia coli

“…The pH profile of glutamine transport clearly does not reflect the ability of the glutamine binding protein to bind glutamine since this property is unaffected over a pH range of 5-8(Table III). Likewise, it cannot be explained by an inability of vesicles to take up succinate or pyruvate, since the ability of vesicles to take up substances via osmotic shock resistant, ApH+-driven transport systems is, in general, not affected over this pH range (Lombardi & Kaback, 1972). This is especially true of succinate uptake in vesicles (Rayman et al, 1972).…”