Adaptation of Escherichia coli to the uncoupler of oxidative phosphorylation 2,4-dinitrophenol

Abstract

Escherichia coli was found to adapt to the uncoupler of oxidative phosphorylation 2,4-dinitrophenol. The rates of synthesis of 53 proteins were increased following exposure to 2,4-dinitrophenol. Adaptation was accelerated when the cofactor pyrroloquinoline quinone was provided in the growth medium.In Escherichia coli, energy associated with the oxidation of an energy source is conserved as an electrochemical potential difference of hydrogen ions across the cytoplasmic membrane and is termed the proton motive force (Ap). The electrical potential (At) together with the difference in proton concentration between the periplasm and cytoplasm (zApH) make up the proton motive force, which is typically maintained at a steady-state value of -150 to -200 mV (14). Ap is harnessed to drive nutrient transport, ATP synthesis, and other endergonic reactions at the cell membrane.Uncouplers of oxidative phosphorylation, such as 2,4-dinitrophenol (DNP) and carbonyl cyanide m-chlorophenylhydrazone (CCCP), are able to transport protons across biological membranes, which usually have extremely low proton conductance (16,17). By transporting protons down the electrochemical potential gradient, these agents collapse Ap. The collapse of Ap has detrimental effects on the bacterial cell: (i) the rate of proton-coupled ATP synthesis declines; (ii) the pH of the cytoplasm, typically maintained near 7.8 (3), approaches that of the external medium (8); and (iii) nutrient transport is inhibited, as is the translocation of proteins destined for insertion into the cytoplasmic membrane (1, 7, 10).Here we show that wild-type E. coli is able to adapt to the uncoupler of oxidative phosphorylation DNP and that the rate of adaptation is accelerated by the cofactor pyrroloquinoline quinone (PQQ). In addition, we show that E. coli induces approximately 50 proteins, including the heat shock proteins, upon exposure to DNP. E. coli adapts to DNP. E. coli W3110 growing in glucose-rich morpholinepropanesulfonic acid (MOPS) medium (GRM) without methionine (9) at 28°C was exposed to 0.5, 0.75, and 1.0 mM DNP. Following challenge with these concentrations of DNP, there were immediate cutbacks in the growth rate of 45, 75, and 90%, respectively (Fig. 1). Following the cutback in growth rate, cultures exposed to 0.5 and 0.75 mM DNP increased their growth rates and nearly reached the rate at which they grew before exposure to DNP. The growth rate of the culture exposed to 1.0 mM DNP increased somewhat but never reached the pre-DNP growth rate. A similar growth response has been noted when E. coli is exposed to inhibitory concentrations of CCCP (18 (Fig. 2). PQQ did not increase the growth rate of a culture growing in medium without DNP, indicating that PQQ did not act as a general growth enhancer. It was shown that glucose dehydrogenase, and not some other enzyme which utilizes PQQ as a cofactor, was responsible for the accelerated adaptation to DNP by conducting the same experiment using isogenic strain DG300 (gcd::cat), which contains a chloramphenicol resistan...

Glucose starvation stimulon of Escherichia coli: role of integration host factor in starvation survival and growth phase-dependent protein synthesis

“…IHF has previously been shown to inhibit ompC transcription both in vivo and in vitro (14). Interestingly, expression of fumarase and glycerol-6-phosphate kinase is increased in cells exposed to both uncouplers of oxidative phosphorylation (10) and carbon starvation, indicating that similar adjustments in the central metabolic pathways take place during these stresses.…”

Real-time detection of central carbon metabolism in livingEscherichia coliand its response to perturbations

“…1) and the fast formation of increased CO 2 levels in presence of uncoupler likely reflects increased oxidative decarboxylation of pyruvate and 6-phosphogluconate due to reduced reductive power. Such adaptations to chemical uncouplers may be relevant in natural habitats, where bacteria can encounter conditions that are mimicked by chemical uncouplers [26]. As instantaneous pulse-response experiments with hyperpolarized tracer do not require that cells thrive and multiply, the methodology is well suited for studying metabolic adaptations in response to extracellular perturbations, even if these perturbations are detrimental to cellular physiology.…”

Regulation of the Histidine Utilization (Hut) System in Bacteria

“…Glucose dehydrogenase requires an unusual quinone cofactor, pyrroloquinoline quinone (PQQ), for its activity (25). Although E. coli has the gene for glucose dehydrogenase and synthesizes the apoenzyme in a regulated way (39), E. coli lacks the genes or capability for PQQ synthesis and cannot form an active glucose dehydrogenase in pure culture unless PQQ is provided in the medium (98). However, E. coli is chemotactic toward PQQ when glucose is present (22) and thus can probably activate glucose dehydrogenase in mixed cultures.…”