scholarly journals CydDC-mediated reductant export in Escherichia coli controls the transcriptional wiring of energy metabolism and combats nitrosative stress

2016 ◽  
Vol 473 (6) ◽  
pp. 693-701 ◽  
Author(s):  
Louise V. Holyoake ◽  
Stuart Hunt ◽  
Guido Sanguinetti ◽  
Gregory M. Cook ◽  
Mark J. Howard ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Energy Metabolism ◽  
Abc Transporter ◽  
Nitrosative Stress ◽  
Growth And Survival ◽  
E Coli

The CydDC ABC transporter of E. coli exports glutathione and cysteine. Loss of cydDC elicits adaptations in energy metabolism and induces sensitivity to NO. CydDC therefore has a likely role in growth and survival during infection.

scholarly journals Enhancement of the Efficiency of Secretion of Heterologous Lipase in Escherichia coli by Directed Evolution of the ABC Transporter System

2005 ◽  
Vol 71 (7) ◽  
pp. 3468-3474 ◽  
Author(s):  
Gyeong Tae Eom ◽  
Jae Kwang Song ◽  
Jung Hoon Ahn ◽  
Yeon Soo Seo ◽  
Joon Shick Rhee
Keyword(s):  
Escherichia Coli ◽  
Directed Evolution ◽  
Abc Transporter ◽  
Microtiter Plate ◽  
Single Amino Acid ◽  
Wild Type ◽  
E Coli ◽  
Single Amino Acid Change ◽  
Secretion Efficiency

ABSTRACT The ABC transporter (TliDEF) from Pseudomonas fluorescens SIK W1, which mediated the secretion of a thermostable lipase (TliA) into the extracellular space in Escherichia coli, was engineered using directed evolution (error-prone PCR) to improve its secretion efficiency. TliD mutants with increased secretion efficiency were identified by coexpressing the mutated tliD library with the wild-type tliA lipase in E. coli and by screening the library with a tributyrin-emulsified indicator plate assay and a microtiter plate-based assay. Four selected mutants from one round of error-prone PCR mutagenesis, T6, T8, T24, and T35, showed 3.2-, 2.6-, 2.9-, and 3.0-fold increases in the level of secretion of TliA lipase, respectively, but had almost the same level of expression of TliD in the membrane as the strain with the wild-type TliDEF transporter. These results indicated that the improved secretion of TliA lipase was mediated by the transporter mutations. Each mutant had a single amino acid change in the predicted cytoplasmic regions in the membrane domain of TliD, implying that the corresponding region of TliD was important for the improved and successful secretion of the target protein. We therefore concluded that the efficiency of secretion of a heterologous protein in E. coli can be enhanced by in vitro engineering of the ABC transporter.

Bioluminescence: A Rapid Indicator of Escherichia Coli O157:H7 in Selected Yogurt and Cheese Varieties

1997 ◽  
Vol 60 (8) ◽  
pp. 891-897 ◽  
Author(s):  
L. M. HUDSON ◽  
J. CHEN ◽  
A. R. HILL ◽  
M. W. GRIFFITHS
Keyword(s):  
Escherichia Coli ◽  
Enterohemorrhagic Escherichia Coli ◽  
Cottage Cheese ◽  
Plate Count ◽  
Escherichia Coli O157 ◽  
Potential Hazard ◽  
Growth And Survival ◽  
E Coli ◽  
Standard Plate Count ◽  
Standard Plate

Outbreaks of enterohemorrhagic Escherichia coli O157:H7 have been commonly associated with products derived from ground beef, but recently the organism has been implicated as the causative agent in outbreaks involving yogurt and cheese. This finding has raised concern about the potential for its growth and survival in fermented dairy products. A bioluminescent strain of E. coli O157:H7 was used to determine postprocessing survival in yogurt with live cultures at pH 4.17, 4.39, and 4.47 stored at 4 and 10°C. In addition, survival of E. coli O157:H7 was monitored during the manufacture of Cottage, Colby, Romano, and Feta cheeses. Results indicated survival for 8 and 5 days at 4 and 10°C respectively in yogurt at pH 4.17, 17 and 15 days at 4 and 10°C respectively in yogurt at pH 4.39, and 17days at both 4 and 10°C in yogurt at pH 4.47. E. coli O157:H7 did not survive cooking procedures at 56°C in Cottage cheese. However, the pathogen survived for 27, 30, and 27 days in Colby, Romano, and Feta cheeses respectively. A high correlation of r2 > 0.89 was obtained between counts of bioluminescenct colonies and standard plate count for all yogurt and cheese varieties, indicating that bioluminescence was a sensitive and rapid indicator of cellular viability for E. coli O157:H7. Survival of the pathogen, as indicated by this method, is possible in highly acidic environments even at refrigeration temperatures. This poses a potential hazard should postprocessing contamination occur.

scholarly journals Effect of Dimethyl Sulphoxide on the Expression of Nitrogen Fixation in Bacteria

1977 ◽  
Vol 30 (2) ◽  
pp. 141 ◽  
Author(s):  
Mary L Skotnicki ◽  
Barry G Rolfe
Keyword(s):  
Escherichia Coli ◽  
Nitrogen Fixation ◽  
Energy Metabolism ◽  
Membrane Proteins ◽  
Klebsiella Pneumoniae ◽  
Dimethyl Sulphoxide ◽  
Rhizobium Trifolii ◽  
Nif Genes ◽  
E Coli ◽  
Selection Of

Storage in dimethyl sulphoxide (DMSO) of Escherichia coli K12 hybrids carrying nif+ genes from Klebsiella pneumoniae can result in selection of a defective nitrogen-fixing phenotype. Similar results are obtained with E. coli K12 hybrids containing the nitrogep-fixing capacity from Rhizobium trifolii. DMSO appears to affect particular inner membrane proteins associated with energy metabolism in E. coli K12 and four chromosomal regions (chID, chlG, his and unc) are associated with resistance to DMSO.

Food Poisoning Potential of Artificially Contaminated Vacuum Packaged Sliced Ham in Sandwiches1

1981 ◽  
Vol 44 (6) ◽  
pp. 430-434 ◽  
Author(s):  
JAMES E. STEELE ◽  
MICHAEL E. STILES
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Storage Temperature ◽  
Food Poisoning ◽  
Microbial Competition ◽  
Growth And Survival ◽  
E Coli ◽  
Enteropathogenic Bacteria ◽  
Test Conditions ◽  
And Staphylococcus Aureus

Ham sandwiches inoculated with a mixture of five enteropathogenic bacteria, Bacillus cereus, Clostridium perfringens. Escherichia coli, Salmonella typhimurium and Staphylococcus aureus, were held at 30, 21 and 4 C for up to 24 h. Food poisoning potential was judged by the growth and survival of the inoculated pathogens. Major differences were observed between new and old (30 days of storage at 4 C) ham samples. On new ham, all enteropathogens were able to grow except C. perfringens, whereas on old ham, with high microbial competition. the pathogens survived but did not grow. Severe storage temperature abuse was necessary to develop a food poisoning potential in new ham samples. The safety of old ham was attributed to the competitive microflora that grew in the ham during storage at 4 C for 30 days. Infective pathogens, E. coli and S. typhimurium, either survived or increased in numbers under all test conditions. The safety of vacuum packaged sliced ham for use in sandwiches, in its present market form, was indicated by these studies.

scholarly journals Polyamine-Mediated Resistance of Uropathogenic Escherichia coli to Nitrosative Stress

2006 ◽  
Vol 188 (3) ◽  
pp. 928-933 ◽  
Author(s):  
Jean M. Bower ◽  
Matthew A. Mulvey
Keyword(s):  
Escherichia Coli ◽  
Nitrosative Stress ◽  
Resistance Mutation ◽  
Reactive Nitrogen ◽  
E Coli ◽  
Exogenous Addition ◽  
Reactive Nitrogen Intermediates ◽  
Transposon Mutants ◽  
Increased Sensitivity ◽  
K 12

ABSTRACT During the course of a urinary tract infection, substantial levels of nitric oxide and reactive nitrogen intermediates are generated. We have found that many uropathogenic strains of Escherichia coli display far greater resistance to nitrosative stress than the K-12 reference strain MG1655. By selecting and screening for uropathogenic E. coli transposon mutants that are unable to grow in the presence of acidified nitrite, the cadC gene product was identified as a key facilitator of nitrosative stress resistance. Mutation of cadC, or its transcriptional targets cadA and cadB, results in loss of significant production of the polyamine cadaverine and increased sensitivity to acidified nitrite. Exogenous addition of cadaverine or other polyamines rescues growth of cad mutants under nitrosative stress. In wild-type cells, the concentration of cadaverine produced per cell is substantially increased by exposure to acidified nitrite. The mechanism behind polyamine-mediated rescue from nitrosative stress is unclear, but it is not attributable solely to chemical quenching of reactive nitrogen species or reduction in mutation frequency.

Behavior of Escherichia coli O157:H7 on Damaged Leaves of Spinach, Lettuce, Cilantro, and Parsley Stored at Abusive Temperatures

2010 ◽  
Vol 73 (2) ◽  
pp. 212-220 ◽  
Author(s):  
ROWAIDA K. KHALIL ◽  
JOSEPH F. FRANK
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Leaf Damage ◽  
Escherichia Coli O157 ◽  
Oxidation Reactions ◽  
Leaf Extracts ◽  
Romaine Lettuce ◽  
Growth And Survival ◽  
E Coli ◽  
Additional Information

Recent foodborne illness outbreaks associated with the consumption of leafy green produce indicates a need for additional information on the behavior of pathogenic bacteria on these products. Previous research indicates that pathogen growth and survival is enhanced by leaf damage. The objective of this study was to compare the behavior of Escherichia coli O157:H7 on damaged leaves of baby Romaine lettuce, spinach, cilantro, and parsley stored at three abusive temperatures (8, 12, and 15°C). The damaged portions of leaves were inoculated with approximately 105 CFU E. coli O157:H7 per leaf. The pathogen grew on damaged spinach leaves held for 3 days at 8 and 12°C (P < 0.05), with the population increasing by 1.18 and 2.08 log CFU per leaf, respectively. E. coli O157:H7 did not grow on damaged Romaine leaves at 8 or 12°C, but growth was observed after 8 h of storage at 15°C, with an increase of less than 1.0 log. Growth of E. coli O157:H7 on Romaine lettuce held at 8 or 12°C was enhanced when inocula were suspended in 0.05% ascorbic acid, indicating the possibility of inhibition by oxidation reactions associated with tissue damage. Damaged cilantro and Italian parsley leaves held at 8°C for 4 days did not support the growth of E. coli O157:H7. Behavior of the pathogen in leaf extracts differed from behavior on the damaged tissue. This study provides evidence that the damaged portion of a leafy green is a distinct growth niche that elicits different microbial responses in the various types of leafy greens.

scholarly journals Nitric Oxide in Chemostat-Cultured Escherichia coli Is Sensed by Fnr and Other Global Regulators: Unaltered Methionine Biosynthesis Indicates Lack of S Nitrosation

2006 ◽  
Vol 189 (5) ◽  
pp. 1845-1855 ◽  
Author(s):  
Steven T. Pullan ◽  
Mark D. Gidley ◽  
Richard A. Jones ◽  
Jason Barrett ◽  
Tania M. Stevanin ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Escherichia Coli ◽  
Nitrosative Stress ◽  
Adaptive Responses ◽  
Methionine Biosynthesis ◽  
Transcriptional Responses ◽  
No Donor ◽  
Global Regulators ◽  
E Coli ◽  
Cellular Effects

ABSTRACT We previously elucidated the global transcriptional responses of Escherichia coli to the nitrosating agent S-nitrosoglutathione (GSNO) in both aerobic and anaerobic chemostats, demonstrated the expression of nitric oxide (NO)-protective mechanisms, and obtained evidence of critical thiol nitrosation. The present study was the first to examine the transcriptome of NO-exposed E. coli in a chemostat. Using identical conditions, we compared the GSNO stimulon with the stimulon of NO released from two NO donor compounds {3-[2-hydroxy-1-(1-methyl-ethyl)-2-nitrosohydrazino]-1-propanamine (NOC-5) and 3-(2-hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-propanamine (NOC-7)} simultaneously and demonstrated that there were marked differences in the transcriptional responses to these distinct nitrosative stresses. Exposure to NO did not induce met genes, suggesting that, unlike GSNO, NO does not elicit homocysteine S nitrosation and compensatory increases in methionine biosynthesis. After entry into cells, exogenous methionine provided protection from GSNO-mediated killing but not from NO-mediated killing. Anaerobic exposure to NO led to up-regulation of multiple Fnr-repressed genes and down-regulation of Fnr-activated genes, including nrfA, which encodes cytochrome c nitrite reductase, providing strong evidence that there is NO inactivation of Fnr. Other global regulators apparently affected by NO were IscR, Fur, SoxR, NsrR, and NorR. We tried to identify components of the NorR regulon by performing a microarray comparison of NO-exposed wild-type and norR mutant strains; only norVW, encoding the NO-detoxifying flavorubredoxin and its cognate reductase, were unambiguously identified. Mutation of norV or norR had no effect on E. coli survival in mouse macrophages. Thus, GSNO (a nitrosating agent) and NO have distinct cellular effects; NO more effectively interacts with global regulators that mediate adaptive responses to nitrosative stress but does not affect methionine requirements arising from homocysteine nitrosation.

scholarly journals Widespread Distribution in Pathogenic Bacteria of Di-Iron Proteins That Repair Oxidative and Nitrosative Damage to Iron-Sulfur Centers

2008 ◽  
Vol 190 (6) ◽  
pp. 2004-2013 ◽  
Author(s):  
Tim W. Overton ◽  
Marta C. Justino ◽  
Ying Li ◽  
Joana M. Baptista ◽  
Ana M. P. Melo ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Nitrosative Stress ◽  
Human Pathogens ◽  
Paramagnetic Resonance ◽  
Widespread Distribution ◽  
Iron Proteins ◽  
E Coli

ABSTRACT Expression of two genes of unknown function, Staphylococcus aureus scdA and Neisseria gonorrhoeae dnrN, is induced by exposure to oxidative or nitrosative stress. We show that DnrN and ScdA are di-iron proteins that protect their hosts from damage caused by exposure to nitric oxide and to hydrogen peroxide. Loss of FNR-dependent activation of aniA expression and NsrR-dependent repression of norB and dnrN expression on exposure to NO was restored in the gonococcal parent strain but not in a dnrN mutant, suggesting that DnrN is necessary for the repair of NO damage to the gonococcal transcription factors, FNR and NsrR. Restoration of aconitase activity destroyed by exposure of S. aureus to NO or H2O2 required a functional scdA gene. Electron paramagnetic resonance spectra of recombinant ScdA purified from Escherichia coli confirmed the presence of a di-iron center. The recombinant scdA plasmid, but not recombinant plasmids encoding the complete Escherichia coli sufABCDSE or iscRSUAhscBAfdx operons, complemented repair defects of an E. coli ytfE mutant. Analysis of the protein sequence database revealed the importance of the two proteins based on the widespread distribution of highly conserved homologues in both gram-positive and gram-negative bacteria that are human pathogens. We provide in vivo and in vitro evidence that Fe-S clusters damaged by exposure to NO and H2O2 can be repaired by this new protein family, for which we propose the name repair of iron centers, or RIC, proteins.

scholarly journals Time-Dependent Proteome Alterations under Osmotic Stress during Aerobic and Anaerobic Growth in Escherichia coli

2006 ◽  
Vol 188 (20) ◽  
pp. 7165-7175 ◽  
Author(s):  
Arnim Weber ◽  
Stephanie A. Kögl ◽  
Kirsten Jung
Keyword(s):  
Escherichia Coli ◽  
Osmotic Stress ◽  
Integration Host Factor ◽  
Anaerobic Growth ◽  
Time Dependent ◽  
Transcriptional Level ◽  
Anaerobic Conditions ◽  
Growth And Survival ◽  
E Coli ◽  
Protein Gels

ABSTRACT Escherichia coli lives in the mammalian gastrointestinal tract anaerobically at high osmolarity as well as in the soil aerobically at varying osmolarities. Adaptation to these varying environmental conditions is crucial for growth and survival of E. coli. Two-dimensional protein gels were used to visualize global time-dependent changes (10 to 60 min) in the proteome of cells responding to osmotic stress (0.4 M NaCl or 0.7 M sorbitol) under aerobic or anaerobic conditions. The protein profiles revealed an induction of 12 proteins (Dps, HchA, HdhA, InfB, OsmC, OsmY, ProX, KatE, PspA, TalA, TktB, and TreF) under osmotic stress in an aerobic milieu. Eleven additional proteins (OtsB, YceI, YciE, YciF, YgaU, YjbJ, AcnA, MetL, PoxB, Ssb, and YhbO) were induced by osmotic stress imposed by NaCl. Most of the accumulated proteins were cross-protecting proteins (e.g., OsmY, OsmC, Dps, and KatE) which are regulated at the transcriptional level predominantly by RpoS and other regulators (e.g., integration host factor, OxyR, H-NS, LRP, and FIS). Comparative analysis of the proteome of E. coli grown under aerobic or anaerobic conditions under osmotic stress (NaCl) revealed an overlap of the up-regulated proteins of more than 50%. Ten proteins (PoxB, AcnA, TalA, TktB, KatE, PspA, Ssb, TreF, MetL, and YhbO) were detectable only under aerobic, high-osmolality conditions. Time-dependent alterations of the proteome were monitored, allowing classification of the up-regulated proteins into early, middle, and long-term phases of adaptation. Only a few proteins were found to be down-regulated upon osmotic stress.

scholarly journals A Predicted ABC Transporter, FtsEX, Is Needed for Cell Division in Escherichia coli

2004 ◽  
Vol 186 (3) ◽  
pp. 785-793 ◽  
Author(s):  
Kari L. Schmidt ◽  
Nicholas D. Peterson ◽  
Ryan J. Kustusch ◽  
Mark C. Wissel ◽  
Becky Graham ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Abc Transporter ◽  
Luria Bertani ◽  
Wild Type ◽  
E Coli ◽  
Division Site ◽  
Site Localization ◽  
Free Media ◽  
Mass Increase

ABSTRACT FtsE and FtsX have homology to the ABC transporter superfamily of proteins and appear to be widely conserved among bacteria. Early work implicated FtsEX in cell division in Escherichia coli, but this was subsequently challenged, in part because the division defects in ftsEX mutants are often salt remedial. Strain RG60 has an ftsE::kan null mutation that is polar onto ftsX. RG60 is mildly filamentous when grown in standard Luria-Bertani medium (LB), which contains 1% NaCl, but upon shift to LB with no NaCl growth and division stop. We found that FtsN localizes to potential division sites, albeit poorly, in RG60 grown in LB with 1% NaCl. We also found that in wild-type E. coli both FtsE and FtsX localize to the division site. Localization of FtsX was studied in detail and appeared to require FtsZ, FtsA, and ZipA, but not the downstream division proteins FtsK, FtsQ, FtsL, and FtsI. Consistent with this, in media lacking salt, FtsA and ZipA localized independently of FtsEX, but the downstream proteins did not. Finally, in the absence of salt, cells depleted of FtsEX stopped dividing before any change in growth rate (mass increase) was apparent. We conclude that FtsEX participates directly in the process of cell division and is important for assembly or stability of the septal ring, especially in salt-free media.

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