Characterisation of new intracellular membranes in Escherichia coli accompanying large scale over‐production of the b subunit of F1Fo ATP synthase

Aréchaga, Ignacio; Miroux, Bruno; Karrasch, Simone; Huijbregts, Richard P. H.; Kruijff, Ben de; Runswick, Michael J.; Walker, John E.

doi:10.1016/s0014-5793(00)02054-8

Cited by 149 publications

(137 citation statements)

References 28 publications

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“…For example, it is possible that increased expression of an ICM protein(s) is sufficient to drive increased membrane synthesis. This is illustrated by observations in Escherichia coli, where overexpression of individual integral membrane proteins, including fumarate reductase (46), the glycerol acyl transferase PlsB (47), or the ATP synthase b subunit (48), is sufficient to induce formation of intracellular membranes. Indeed, we found that when synthesis of SCs was increased by mutation under high O 2 tension, when they are not normally present, membrane lipid content also increased ( Fig.…”

Section: Discussionmentioning

confidence: 99%

Oxygen-Dependent Regulation of Bacterial Lipid Production

et al. 2015

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Understanding the mechanisms of lipid accumulation in microorganisms is important for several reasons. In addition to providing insight into assembly of biological membranes, lipid accumulation has important applications in the production of renewable fuels and chemicals. The photosynthetic bacterium Rhodobacter sphaeroides is an attractive organism to study lipid accumulation, as it has the ability to increase membrane production at low O 2 tensions. Under these conditions, R. sphaeroides develops invaginations of the cytoplasmic membrane to increase its membrane surface area for housing of the membrane-bound components of its photosynthetic apparatus. Here we use fatty acid levels as a reporter of membrane lipid content. We show that, under low-O 2 and anaerobic conditions, the total fatty acid content per cell increases 3-fold. We also find that the increases in the amount of fatty acid and photosynthetic pigment per cell are correlated as O 2 tensions or light intensity are changed. To ask if lipid and pigment accumulation were genetically separable, we analyzed strains with mutations in known photosynthetic regulatory pathways. While a strain lacking AppA failed to induce photosynthetic pigment-protein complex accumulation, it increased fatty acid content under low-O 2 conditions. We also found that an intact PrrBA pathway is required for low-O 2 -induced fatty acid accumulation. Our findings suggest a previously unknown role of R. sphaeroides transcriptional regulators in increasing fatty acid and phospholipid accumulation in response to decreased O 2 tension. IMPORTANCELipids serve important functions in living systems, either as structural components of membranes or as a form of carbon storage. Understanding the mechanisms of lipid accumulation in microorganisms is important for providing insight into the assembly of biological membranes and additionally has important applications in the production of renewable fuels and chemicals. In this study, we investigate the ability of Rhodobacter sphaeroides to increase membrane production at low O 2 tensions in order to house its photosynthetic apparatus. We demonstrate that this bacterium has a mechanism to increase lipid content in response to decreased O 2 tension and identify a transcription factor necessary for this response. This is significant because it identifies a transcriptional regulatory pathway that can increase microbial lipid content. Lipids serve important functions in living systems, either as structural components of membranes or as a form of carbon storage. Lipids derived from oil-rich microorganisms (including bacteria, yeasts, and microalgae) also offer a promising source of renewable fuels and chemicals (1, 2). However, the genetic and biochemical mechanisms regulating lipid accumulation in microorganisms are poorly understood, have been difficult to study, and are typically linked to stress conditions that hinder growth (3-5). Much effort has been directed at increasing lipid accumulation through alteration of enzymes involved in ...

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Section: Discussionmentioning

confidence: 99%

Oxygen-Dependent Regulation of Bacterial Lipid Production

et al. 2015

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“…In other examples of E. coli membrane protein overexpression (3,11,43), extended membrane structures form that are enriched in the overexpressed protein.…”

Section: Discussionmentioning

confidence: 99%

Electron Microscopic Analysis of Membrane Assemblies Formed by the Bacterial Chemotaxis Receptor Tsr

et al. 2003

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The serine receptor (Tsr) from Escherichia coli is representative of a large family of transmembrane receptor proteins that mediate bacterial chemotaxis by influencing cell motility through signal transduction pathways. Tsr and other chemotaxis receptors form patches in the inner membrane that are often localized at the poles of the bacteria. In an effort to understand the structural constraints that dictate the packing of receptors in the plane of the membrane, we have used electron microscopy to examine ordered assemblies of Tsr in membrane extracts isolated from cells engineered to overproduce the receptor. Three types of assemblies were observed: ring-like "micelles" with a radial arrangement of receptor subunits, two-dimensional crystalline arrays with approximate hexagonal symmetry, and "zippers," which are receptor bilayers that result from the antiparallel interdigitation of cytoplasmic domains. The registration among Tsr molecules in the micelle and zipper assemblies was sufficient for identification of the receptor domains and for determination of their contributions to the total receptor length. The overall result of this analysis is compatible with an atomic model of the receptor dimer that was constructed primarily from the X-ray crystal structures of the periplasmic and cytoplasmic domains. Significantly, the micelle and zipper structures were also observed in fixed, cryosectioned cells expressing the Tsr receptor at high abundance, suggesting that the modes of Tsr assembly found in vitro are relevant to the situation in the cell.The serine receptor (Tsr), one of four methyl-accepting chemotaxis proteins (MCPs) that span the inner membrane of Escherichia coli, initiates responses and governs adaptation to changes in the serine concentration. MCPs belong to a large class of transducers (21, 46), which sense a variety of environmental cues and are the inputs to sensory pathways that bias cell movement toward favorable environments (12). The chemotaxis pathways belong to the two-component superfamily of signal transduction pathways (17, 42), which are chiefly found in prokaryotes. A two-component pathway consists of a sensor, which is frequently an integral membrane protein possessing kinase activity, and one or more cytoplasmic phosphate-accepting response regulator proteins. The transmembrane sensor-kinases of the chemotaxis pathways are often noncovalent complexes between MCPs (which have no enzyme activity) and two soluble cytoplasmic proteins, namely, an adaptor protein (CheW) and a kinase (CheA) (15, 39).Elucidation of the structure and distribution of receptors in the membrane of the cell is integral to understanding the molecular basis of signaling by the transmembrane sensor (MCP-CheW-CheA) complexes. X-ray structure determination of the soluble domains has clearly defined the dimeric organization of the 60-kDa receptor subunits (19,31,45), and functional studies have helped to elucidate the role of dimer organization in the mechanism of transmembrane signaling (references 32 and 12 and refer...

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“…Overexpression of some membrane proteins, including ones that synthesize lipids (38,39) and cell wall precursors (40) or depletion of some membrane assembly proteins (41), may result in the formation of new membrane structures in the cytoplasm. Given the NB character of MGlcDAG, it was also important to establish whether the large amount of this lipid was properly integrated into the membranes.…”

Section: Mglcdag-containing Strain Is Less Dependent On Mg 2ϩ -mentioning

confidence: 99%

Monoglucosyldiacylglycerol, a Foreign Lipid, Can Substitute for Phosphatidylethanolamine in Essential Membrane-associated Functions in Escherichia coli

Wikström

Xie²,

Bogdanov³

et al. 2004

Journal of Biological Chemistry

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The mechanisms by which lipid bilayer properties govern or influence membrane protein functions are little understood, but a liquid-crystalline state and the presence of anionic and nonbilayer (NB)-prone lipids seem important. An Escherichia coli mutant lacking the major membrane lipid phosphatidylethanolamine (NBprone) requires divalent cations for viability and cell integrity and is impaired in several membrane functions that are corrected by introduction of the "foreign" NBprone neutral glycolipid ␣-monoglucosyldiacylglycerol (MGlcDAG) synthesized by the MGlcDAG synthase from Acholeplasma laidlawii. Dependence on Mg 2؉ was reduced, and cellular yields and division malfunction were greatly improved. The increased passive membrane permeability of the mutant was not abolished, but protein-mediated osmotic stress adaptation to salts and sucrose was recovered by the presence of MGlcDAG. MGlcDAG also restored tryptophan prototrophy and active transport function of lactose permease, both critically dependent on phosphatidylethanolamine. Three mechanisms can explain the observed effects: NB-prone MGlcDAG improves the quenched lateral pressure profile across the bilayer; neutral MGlcDAG dilutes the high anionic lipid surface charge; MGlcDAG provides a neutral lipid that can hydrogen bond and/or partially ionize. The reduced dependence on Mg 2؉ and lack of correction by high monovalent salts strongly support the essential nature of the NB properties of MGlcDAG.The lipid bilayer of biological membranes acts as a permeability barrier permitting maintenance of essential ion gradients and is also the local environment for integral and peripheral membrane proteins. Important bilayer structural features are a liquid-crystalline state, an optimal length of the lipid chains, and critical fractions of anionic and nonbilayer-(NB) 1 prone lipids. Because of their small head groups the NB-prone lipids, when embedded in a bilayer, cause a curvature elastic stress of each monolayer with closer acyl chain packing (i.e. higher chain order) that changes the lateral pressure profile across the membrane (1). Increasing the proportion of NBprone lipids leads to a lower temperature for the bilayer to NB phase transition, which in many cases is fairly close to the growth temperature of organisms (2, 3). The balance of bilayer to NB-prone lipids may affect the passive membrane permeability of the bilayer (e.g. (4)), the folding and assembly of membrane proteins (5), and the function of important cellular processes such as cell division, transport, and osmotic responses. The essential character of such NB-prone lipids for cell membrane functions has been little studied.NB-prone lipids substantially affect the activity of several different types of proteins when studied in vitro. For example, a critical conformational change of transmembrane rhodopsin (6) and the activity of the interface-bound CTP:phosphocholine cytidylyltransferase (7) are modulated by variations in NBprone lipids and membrane curvature elastic stress, respectively. Likewise...

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Characterisation of new intracellular membranes in Escherichia coli accompanying large scale over‐production of the b subunit of F₁F_o ATP synthase

Abstract: Recombinant membrane proteins in Escherichia coli are either expressed at relatively low level in the cytoplasmic membrane or they accumulate as inclusion bodies. Here, we report that the abundant over-production of subunit b of E. coli

Cited by 149 publications

References 28 publications

Oxygen-Dependent Regulation of Bacterial Lipid Production

Oxygen-Dependent Regulation of Bacterial Lipid Production

Electron Microscopic Analysis of Membrane Assemblies Formed by the Bacterial Chemotaxis Receptor Tsr

Monoglucosyldiacylglycerol, a Foreign Lipid, Can Substitute for Phosphatidylethanolamine in Essential Membrane-associated Functions in Escherichia coli

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