Roles of ribosomal binding, membrane potential, and electron transport in bacterial uptake of streptomycin and gentamicin

Abstract

The effects of a set of conditions on aminoglycoside uptake were determined. Membrane vesicles either with a membrane potential (A0,) of -125 mV (adequate to drive lysine uptake) or with succinate, lactate, or phenazine methosulfate did not accumulate gentamicin unless components of protein synthesis were included. Ribosomally resistant (rpsL) Escherichia coli cells demonstrated energy-dependent phase II uptake similar to that of a streptomycin-susceptible strain of E. coli when treated with 100 ,ug of puromycin per ml. Puromycin (100 ,ug/ml) also increased the uptake of the cationic compounds polyamine and arginine. These studies support a role of protein synthesis in aminoglycoside uptake and in the development of energy-dependent phase II. A* of cells did not increase either at the initiation of or during energy-denpendent phase II, showing that energydependent phase II is not due to an elevation of A+. In a Bacillus subtilis system, significant streptomycin uptake requires a threshold value of A* which varies depending upon the concentration of streptomycin used. At 25 ,ug/ml, the uptake of streptomycin reached maximal levels after exceeding the threshold value, whereas at 100 p.g/ml there was a gradual increase of the uptake to the maximal after the threshold value was exceeded. Several studies supported the view that electron transport has a specific role other than its requirement to produce the cellular &*. The uptake of gentamicin was stimulated to a greater extent by phenazine methosulfate-ascorbate than by the ionophore nigericin in strains of E. coli, although nigericin stimulated Ai, to a greater degree. Cells with 25% of the normal quinone concentration had values identical to cells with the normal quinone concentration, but the quinone-deficient cells had a significantly lower rate of gentamicin uptake. KCN prevented gentamicin uptake but did not prevent the development of A+. The effects of ubiquinone depletion in an E. coli strain were more evident on gentamicin uptake than on ATP-driven glutamine transport or proton motive force-driven proline transport, consistent with a specific requirement for quinones in aminoglycoside uptake. A detailed explanation of the mechanism of accumulation of streptomycin and gentamicin and a proposed mechanism for killing bacterial cells by these agents have been provided.The uptake of the aminoglycoside antibiotics streptomycin and gentamicin has been shown to be influenced by a complex set of conditions. It has been shown that the kinetics of uptake involve an initial energy-independent phase associated with ionic binding to the cell surface and cytoplasmic membrane. This is followed by two energy-dependent phases, a slow initial rate of uptake termed energy-dependent phase I (EDP-I) and a second accelerated rate termed energy-dependent phase II (EDP-II). Initiation of the latter phase requires binding to ribosomes (reviewed in references 2 and 11).The most effective energy source has been demonstrated to be electron transport involving quinone oxidation-reducti...

The bactericidal action of streptomycin: membrane permeabilization caused by the insertion of mistranslated proteins into the cytoplasmic membrane of Escherichia coli and subsequent caging of the antibiotic inside the cells due to degradation of these proteins

“…Different groups have reported different effects of some antibiotics on the membrane potential of E. coli or Bacillus subtilis : gentamicin depolarizes B. subtilis (Bryan & Kwan, 1983) and E. coli (our unpublished results) ; by contrast, streptomycin hyperpolarizes these bacteria (Emling & Holtje, 1987;Fig. 6 of this study); in addition, chloramphenicol depolarizes B. subtilis (Emling & Holtje, 1987), but hyperpolarizes E. coli (Fig.…”

“…It is difficult to compare these results with each other, since (i) two organisms were treated with different antibiotics; (ii) the media in which the cells were sospended and the temperature were different; (iii) different probes were used to monitor the membrane potentid [i.e. TPP+ by Bryan & Kwan (1983) and by us, and TPMP+ in the presence of a small concentration of tetraphenylboron by Emling & Holtje (1987)l;and (iv) the concentration of the probe was different. The latter point is important, since both E. coli and B. subtilis possess an efflux system for lipophilic cations like TPP+ and TPMP+ (e.g.…”

“…K+-depleted cells develop higher membrane potentials than do K+-replete cells (Kashket & Barker, 1977;Bakker & Mangerich, 1981 ;Kroll & Booth, 1981). Since the rate at which the polycation streptomycin is taken up by the cells is a steep function of the membrane potential (Damper & Epstein, 1981 ;Bryan & Kwan, 1983;Emling & Holtje, 1987), K+-depleted cells will take up streptomycin faster and will be affected by the antibiotic earlier (Figs 1 and 3). Hence, our data indirectly support the notion that the magnitude of the membrane potential determines the rate at which streptomycin is taken up by the cells (Damper & Epstein, 1981 ;Bryan & Kwan, 1983;Taber et al, 1987;Emling & Holtje, 1987).…”

Efficacy of zincoderm G cream against wound infection by Pseudomonas aeruginosa in Sprague-Dawley rats

“…[7][8][9][10] The uptake of an aminoglycoside antibiotic such as gentamicin into P. aeruginosa has been recognized as a multifactorial process involving an initial ionic interaction with the exterior of the cell, followed by two energy-dependent phases requiring an energized cytoplasmic membrane. [11][12][13] One of the studies suggests that zinc has significantly increased in the MICs of imipenem for P. aeruginosa, and, to a lesser extent, for other Gram-negative bacilli in Muller-Hinton agar.…”

Du fer et du soufre dans les protéines

“…Les bases moléculaires de cette observation surprenante se trouvent dans les mécanismes associant transport et force proton motrice (PMF). En accord avec des résultats anciens [48][49][50][51], nous avons démontré dans des conditions in vivo que les aminoglycosides pénètrent dans la cellule grâce à la force proton motrice, elle-même produite par l'activité des chaînes respiratoires. Or, ces dernières impliquent des complexes multiprotéiques, comme les complexes I et II, riches en centres Fe-S, dont le niveau d'activité est plus faible lorsqu'ils subissent une maturation par une souche utilisant la machinerie SUF au lieu de la machinerie ISC.…”