The uptake and acylation of exogenous lysophosphatidylethanolamine by Escherichia coli cells

1980 ◽  
Vol 58 (12) ◽  
pp. 1381-1386 ◽  
Author(s):  
P. Hellion ◽  
F. Landry ◽  
P. V. Subbaiah ◽  
P. Proulx
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Isotope Ratio ◽  
Inorganic Phosphate ◽  
Cell Envelope ◽  
Membrane Lipid ◽  
Water Soluble ◽  
Absorption Process ◽  
Outer Membranes ◽  
Escherichia Coli Cells

Escherichia coli envelopes were fractionated to yield inner and outer membrane fractions. Both these fractions were found to convert [14C]lysophosphatidylethanolamine to its diacyl analogue. Intact Escherichia coli cells were capable of absorbing exogenous labelled lysophosphatidylethanolamine and converting it to phosphatidylethanolamine. When the 14C- and 32P-labelled lyso analogue was used, both the absorption process and the conversion to diacyl analogue proceeded without a significant change in isotope ratio either in the presence or in the absence of added inorganic phosphate. The absorption process was not markedly stimulated by Ca2+ in the medium; it proceeded to an amount representing 25–30% of the endogenous membrane lipid and was accompanied by some degradation to water-soluble products which accumulated in the cell mainly, but also in the incubation medium. The absorbed lipid was recovered in both the inner and outer membrane fractions of the cell envelope. The results indicate that Escherichia coli inner and outer membranes are capable of absorbing exogenous lysophosphoglyceride and converting it into structurally useful diacyl analogue.

scholarly journals Kinetic Analysis of the Assembly of the Outer Membrane Protein LamB in Escherichia coli Mutants Each Lacking a Secretion or Targeting Factor in a Different Cellular Compartment

2006 ◽  
Vol 189 (2) ◽  
pp. 446-454 ◽  
Author(s):  
Alejandro R. Ureta ◽  
Robert G. Endres ◽  
Ned S. Wingreen ◽  
Thomas J. Silhavy
Keyword(s):  
Escherichia Coli ◽  
Kinetic Analysis ◽  
Outer Membrane ◽  
Cell Envelope ◽  
Normal Growth ◽  
Reaction Rates ◽  
Assembly Process ◽  
Cellular Compartment ◽  
Outer Membranes ◽  
Additional Role

ABSTRACT Outer membrane β-barrel proteins in gram-negative bacteria, such as Escherichia coli, must be translocated from their site of synthesis in the cytoplasm to the periplasm and finally delivered to the outer membrane. At least a dozen proteins located in the cytoplasm, the periplasm, and both the inner and outer membranes are required to catalyze this complex assembly process. At normal growth temperatures and conditions the transport and assembly processes are so fast that assembly intermediates cannot be detected. Using cells grown at a low temperature to slow the assembly process and pulse-chase analysis with immunodetection methods, we followed newly synthesized LamB molecules during their transit through the cell envelope. The quality and reproducibility of the data allowed us to calculate rate constants for three different subassembly reactions. This kinetic analysis revealed that secB and secD mutants exhibit nearly identical defects in precursor translocation from the cytoplasm. However, subsequent subassembly reaction rates provided no clear evidence for an additional role for SecD in LamB assembly. Moreover, we found that surA mutants are qualitatively indistinguishable from yfgL mutants, suggesting that the products of both of these genes share a common function in the assembly process, most likely the delivery of LamB to the YaeT assembly complex in the outer membrane.

scholarly journals MCE domain proteins: conserved inner membrane lipid-binding proteins required for outer membrane homeostasis

2017 ◽  
Author(s):  
Georgia L. Isom ◽  
Nathaniel J. Davies ◽  
Zhi-Soon Chong ◽  
Jack A. Bryant ◽  
Mohammed Jamshad ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Cell Envelope ◽  
Membrane Lipid ◽  
Lipid Binding ◽  
Single Domain ◽  
Inner Membrane ◽  
Bacterial Phyla ◽  
Single Domain Protein ◽  
Lipid Binding Proteins

Bacterial proteins with MCE domains were first described as being important forMammalianCellEntry. More recent evidence suggests they are components of lipid ABC transporters. InEscherichia coli, the single-domain protein MlaD is known to be part of an inner membrane transporter that is important for maintenance of outer membrane lipid asymmetry. Here we describe two multi MCE domain-containing proteins inEscherichia coli, PqiB and YebT, the latter of which is an orthologue of MAM-7 that was previously reported to be an outer membrane protein. We show that all three MCE domain-containing proteins localise to the inner membrane. Bioinformatic analyses revealed that MCE domains are widely distributed across bacterial phyla but multi MCE domain-containing proteins evolved in Proteobacteria from single-domain proteins. Mutants defective inmlaD,pqiABandyebSTwere shown to have distinct but partially overlapping phenotypes, but the primary functions of PqiB and YebT differ from MlaD. Complementing our previous findings that all three proteins bind phospholipids, results presented here indicate that multi-domain proteins evolved in Proteobacteria for specific functions in maintaining cell envelope homeostasis.

scholarly journals Biosynthesis and turnover of outer-membrane proteins in Escherichia coli ML308-225

1979 ◽  
Vol 182 (2) ◽  
pp. 407-412 ◽  
Author(s):  
R J Allen ◽  
G K Scott
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Outer Membrane ◽  
Bacterial Growth ◽  
Growth Phase ◽  
Exponential Growth ◽  
Outer Membrane Proteins ◽  
Exponential Growth Phase ◽  
Outer Membranes ◽  
Dilution Experiments

Isolated outer membranes and outer-membrane extracts from Escherichia coli ML308-225 in the early-exponential growth phase contain more protein than do corresponding preparations from late-exponential- or stationary-phase bacteria. Isotope-dilution experiments show that this is due to a loss of protein from the membrane during the exponential growth phase. Inhibition of bacterial growth and protein synthesis stabilizes the outer-membrane-protein concentration. Protein synthesis in the absence of bacterial growth results in higher concentrations of protein in the outer membrane.

scholarly journals Mechanisms of Resistance to the Contact-Dependent Bacteriocin CdzC/D inCaulobacter crescentus

2019 ◽  
Vol 201 (8) ◽  
Author(s):  
Leonor García-Bayona ◽  
Kevin Gozzi ◽  
Michael T. Laub
Keyword(s):  
Outer Membrane ◽  
Caulobacter Crescentus ◽  
Cell Envelope ◽  
Target Cells ◽  
Dependent Manner ◽  
Content Type ◽  
Repeat Protein ◽  
Outer Membranes ◽  
Oligotrophic Bacterium ◽  
Small Hydrophobic

ABSTRACTThe Cdz bacteriocin system allows the aquatic oligotrophic bacteriumCaulobacter crescentusto kill closely related species in a contact-dependent manner. The toxin, which aggregates on the surfaces of producer cells, is composed of two small hydrophobic proteins, CdzC and CdzD, each bearing an extended glycine-zipper motif, that together induce inner membrane depolarization and kill target cells. To further characterize the mechanism of Cdz delivery and toxicity, we screened for mutations that render a target strain resistant to Cdz-mediated killing. These mutations mapped to four loci, including a TonB-dependent receptor, a three-gene operon (namedzerRABforzipperenveloperesistance), andperA(forpentapeptideenveloperesistance). Mutations in thezerRABlocus led to its overproduction and to potential changes in cell envelope composition, which may diminish the susceptibility of cells to Cdz toxins. TheperAgene is also required to maintain a normal cell envelope, but our screen identified mutations that confer resistance to Cdz toxins without substantially affecting the cell envelope functions of PerA. We demonstrate that PerA, which encodes a pentapeptide repeat protein predicted to form a quadrilateral β-helix, localizes primarily to the outer membrane of cells, where it may serve as a receptor for the Cdz toxins. Collectively, these results provide new insights into the function and mechanisms of an atypical, contact-dependent bacteriocin system.IMPORTANCEBacteriocins are commonly used by bacteria to kill neighboring cells that compete for resources. Although most bacteriocins are secreted, the aquatic, oligotrophic bacteriumCaulobacter crescentusproduces a two-peptide bacteriocin, CdzC/D, that remains attached to the outer membranes of cells, enabling contact-dependent killing of cells lacking the immunity protein CdzI. The receptor for CdzC/D has not previously been reported. Here, we describe a genetic screen for mutations that confer resistance to CdzC/D. One locus identified,perA, encodes a pentapeptide repeat protein that resides in the outer membrane of target cells, where it may act as the direct receptor for CdzC/D. Collectively, our results provide new insight into bacteriocin function and diversity.

High-Pressure Transient Sensitization of Escherichia coli to Lysozyme and Nisin by Disruption of Outer-Membrane Permeability

1996 ◽  
Vol 59 (4) ◽  
pp. 350-355 ◽  
Author(s):  
KRISTEL J. A. HAUBEN ◽  
ELKE Y. WUYTACK ◽  
CARINE C. F. SOONTJENS ◽  
CHRIS W. MICHIELS
Keyword(s):  
Escherichia Coli ◽  
High Pressure ◽  
Outer Membrane ◽  
High Pressures ◽  
Ethylenediaminetetraacetic Acid ◽  
Viable Count ◽  
Water Soluble ◽  
Pressure Treatment ◽  
E Coli ◽  
Glucose Agar

Escherichia coli MG1655 suspensions in 10 mM phosphate buffer (pH 7.0) were subjected to high pressures in the range of 180 to 320 MPa for 15 min. Cell death was evident at 220 MPa and increased exponentially with pressure. Surviving populations were sublethally injured, as demonstrated by their reduced ability to form colonies on violet red bile glucose agar, a selective growth medium containing crystal violet and bile salts. During exposure to high pressure (> 180 MPa), cells were sensitive to lysozyme, nisin, and ethylenediaminetetraacetic acid (EDTA), as was apparent from an increased lethality of pressure in the presence of these agents. Sublethal injury in the surviving population was lower in the presence of nisin and lysozyme, but higher in the presence of EDTA. Combinations of EDTA with nisin or lysozyme present during pressure treatment increased lethality in an additive manner. However, the addition of lysozyme, nisin and/or EDTA to pressurized cell suspensions immediately after pressure treatment did not cause any viable count reduction. Finally, we observed leakage of the periplasmic enzyme β-lactamase from an ampicillin-resistant recombinant E. coli MG1655 under high pressure. These results suggest that high pressure transiently disrupts the permeability of the E. coli outer membrane for water-soluble proteins.

Formation of radioactive diacylglycerol from radioisotope-labelled phosphatidylethanolamine by Escherichia coli extracts

1989 ◽  
Vol 67 (6) ◽  
pp. 288-292 ◽  
Author(s):  
H. Aubry ◽  
P. Proulx
Keyword(s):  
Escherichia Coli ◽  
Phospholipase C ◽  
Substrate Concentration ◽  
Incubation Medium ◽  
Isotope Ratio ◽  
Enzyme Concentration ◽  
Water Soluble ◽  
Ph Optimum ◽  
Direct Incorporation

Radioisotope-labelled phosphatidylethanolamine can be converted to radioactive diacylglycerol in the presence of added unlabelled diacylglycerol. With [14C-glycerol; 3H-acyl]phosphatidylethanolamine as substrate, the conversion to double-labelled diacylglycerol occurred without change in isotope ratio indicating that the whole diacylglycerol moiety of phosphatidylethanolamine was directly involved. With [3H-acyl; 32P]phosphatidylethanolamine, formation of [3H]diacylglycerol occurred without production of labelled water-soluble products and consequently no phospholipase C activity could be detected. Under similar conditions, a conversion of [14C-acyl]- or [3H-acyl]-diacylglycerol to labelled phosphatidylethanolamine could also be shown even in the presence of hydroxylamine. [14C-Glycerol; 3H-acyl] diacylglycerol was converted to double-labelled product without change in isotope ratio which again indicated a direct incorporation of the entire diacylglycerol molecule into phosphatidylethanolamine. Both types of conversions were dependent upon time, enzyme concentration, and substrate concentration, and both displayed a pH optimum of approximately 6 and required no added cofactors. In the presence of increasing concentrations of [14C-acyl]diacylglycerol, added to incubation medium containing [3H-acyl]phosphatidylethanolamine, equal amounts of [14C]phosphatidylethanolamine and [3H]diacylglycerol were formed which matched the decrease in [3H]phosphatidylethanolamine. From these results, we conclude that Escherichia coli has an enzyme that catalyses the exchange between the diacylglycerol moiety of phosphatidylethanolamine and free diacylglycerol, with complete sparing of the phosphoethanolamine moiety.Key words: diacylglycerol, phosphatidylethanolamine, exchange, Escherichia coli.

scholarly journals The Type IV Pilus Assembly Complex: Biogenic Interactions among the Bundle-Forming Pilus Proteins of Enteropathogenic Escherichia coli

2002 ◽  
Vol 184 (13) ◽  
pp. 3457-3465 ◽  
Author(s):  
Sandra W. Ramer ◽  
Gary K. Schoolnik ◽  
Cheng-Yen Wu ◽  
Jaiweon Hwang ◽  
Sarah A. Schmidt ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Cell Compartment ◽  
Inner Membrane ◽  
Outer Membranes ◽  
Type Iv ◽  
Pilin Subunit ◽  
Membrane Compartments

ABSTRACT Production of type IV bundle-forming pili (BFP) by enteropathogenic Escherichia coli (EPEC) requires the protein products of 12 genes of the 14-gene bfp operon. Antisera against each of these proteins were used to demonstrate that in-frame deletion of individual genes within the operon reduces the abundance of other bfp operon-encoded proteins. This result was demonstrated not to be due to downstream polar effects of the mutations but rather was taken as evidence for protein-protein interactions and their role in the stabilization of the BFP assembly complex. These data, combined with the results of cell compartment localization studies, suggest that pilus formation requires the presence of a topographically discrete assembly complex that is composed of BFP proteins in stoichiometric amounts. The assembly complex appears to consist of an inner membrane component containing three processed, pilin-like proteins, BfpI, -J, and -K, that localize with BfpE, -L, and -A (the major pilin subunit); an outer membrane, secretin-like component, BfpB and -G; and a periplasmic component composed of BfpU. Of these, only BfpL consistently localizes with both the inner and outer membranes and thus, together with BfpU, may articulate between the Bfp proteins in the inner membrane and outer membrane compartments.

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