Uncoupling of Substrate-Level Phosphorylation in Escherichia coli during Glucose-Limited Growth

Abstract

<jats:title>ABSTRACT</jats:title><jats:p>The respiratory chain of<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">Escherichia coli</jats:named-content>contains three different cytochrome oxidases. Whereas the cytochrome<jats:italic>bo</jats:italic>oxidase and the cytochrome<jats:italic>bd</jats:italic>-I oxidase are well characterized and have been shown to contribute to proton translocation, physiological data suggested a nonelectrogenic functioning of the cytochrome<jats:italic>bd</jats:italic>-II oxidase. Recently, however, this view was challenged by an<jats:italic>in vitro</jats:italic>biochemical analysis that showed that the activity of cytochrome<jats:italic>bd</jats:italic>-II oxidase does contribute to proton translocation with an H<jats:sup>+</jats:sup>/e<jats:sup>−</jats:sup>stoichiometry of 1. Here, we propose that this apparent discrepancy is due to the activities of two alternative catabolic pathways: the pyruvate oxidase pathway for acetate production and a pathway with methylglyoxal as an intermediate for the production of lactate. The ATP yields of these pathways are lower than those of the pathways that have so far always been assumed to catalyze the main catabolic flux under energy-limited growth conditions (i.e., pyruvate dehydrogenase and lactate dehydrogenase). Inclusion of these alternative pathways in the flux analysis of growing<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">E. coli</jats:named-content>strains for the calculation of the catabolic ATP synthesis rate indicates an electrogenic function of the cytochrome<jats:italic>bd</jats:italic>-II oxidase, compatible with an H<jats:sup>+</jats:sup>/e<jats:sup>−</jats:sup>ratio of 1. This analysis shows for the first time the extent of bypassing of substrate-level phosphorylation in<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">E. coli</jats:named-content>under energy-limited growth conditions.</jats:p>

Molecular physiology of the dynamic regulation of carbon catabolite repression in Escherichia coli

“…At the maximal growth rate, the calculated H + /e 2 stoichiometry is much higher than the theoretical maximum (i.e. 4) (Puustinen et al, 1989;Wikström, 1984; see also Sharma et al, 2012). This overestimate may be due to multiple factors, one of which is an error in the estimate of the ATP costs of biomass formation.…”

Cytochrome bd from Escherichia coli catalyzes peroxynitrite decomposition

“…The enzyme catalyzes the reduction of molecular oxygen to water, using quinols as the physiological electron donors. The reaction is associated with the generation of a proton motive force via transmembrane charge separation, without involving a proton pumping mechanism [15][16][17][18][19][20][21][22]. Cytochrome bd is composed of two major membrane-spanning polypeptides, subunits I (CydA, 57 kDa) and II (CydB, 43 kDa), and a small protein, named CydX (4 kDa).…”

Refined method to study the posttranslational regulation of alternative oxidases from Arabidopsis thaliana in vitro

“…A suitable approach to characterize the enzymatic properties and the posttranslational regulation of a single quinol oxidase is its heterologous expression in Escherichia coli , which exhibits an extreme respiratory flexibility (Lengeler et al ). The branched, aerobic respiratory chain of E. coli consists of a set of five NADH dehydrogenases (NDH‐I, NDH‐II, WrbA, YhdH and oxido reductase), which transfer electrons into the UQ pool in the cytoplasmic membrane (Calhoun et al , Thorn et al , Sulzenbacher et al , Patridge and Ferry , Bekker et al ), and a set of three UQH 2 oxidases (cytochrome bo , cytochrome bd ‐I and cytochrome bd ‐II), which directly use the electrons from UQH 2 for the reduction of O 2 to H 2 O (Puustinen et al , Dassa et al , Calhoun et al , Bekker et al , Borisov et al , Sharma et al , Giuffrè et al ). In contrast to the respiratory chain in eukaryotes, aerobically growing E. coli cells lack cytochrome c as well as complex III (cytochrome bc 1 complex) and complex IV (cytochrome c oxidase), whereas NDH‐1 is homologous to eukaryotic mitochondrial complex I (NADH dehydrogenase) (Calhoun et al , Efremov and Sazanov ).…”