scholarly journals Dual avatars of E. coli grxB encoded Glutaredoxin 2 perform ascorbate recycling and ion channel activities

2021 ◽  
Author(s):  
Sreeshma Nellootil Sreekumar ◽  
Bhaba Krishna Das ◽  
Rahul Raina ◽  
Neethu Puthumadathil ◽  
Sonakshi Udinia ◽  
...  
Keyword(s):  
Ion Channel ◽  
Structural Data ◽  
Bilayer Membrane ◽  
Soluble Form ◽  
Gram Negative Bacteria ◽  
Human Pathogens ◽  
E Coli ◽  
Ascorbate Recycling ◽  
First Time

AbstractGlutaredoxins (Grxs) are single-domain redox enzymes of the thioredoxin superfamily, and primarily function as glutathione (GSH) dependent disulphide reductases. Whereas, the E. coli Glutaredoxin 2 (EcGrx2) encoded by grxB has two conserved GST-fold domains, it still lacks a classical Grx-like functions. In this study, we show for the first time, that EcGrx2 exists in both soluble and membrane integrated forms. The soluble form associates with a previously unidentified GSH dependent dehydroascrobate (DHA) reductase, and the membrane integrated form possesses ion channel activities. Using enzyme kinetic data and structural data we unequivocally demonstrate that EcGrx2 recycles ascorbate (AsA) from DHA. This ability to recycle AsA is inhibited by Zinc (Zn2+). We also show that both wildtype and the E. coli grxB deletion mutant can be rescued from H2O2-induced oxidative stress using ascorbate as an antioxidant, which otherwise is only known as a carbon source in bacteria. Moreover, the grxB- mutant is susceptible to intracellular killing by ROS producing macrophages. We further discovered that EcGrx2 integrates into the native E. coli membrane and show that the purified soluble protein readily inserts into artificial lipid bilayer membrane and conducts ions in vitro. Our data demonstrates a highly conserved functional similarity among EcGrx2-orthologs and highlights that the utilization and subsequent recycling of ascorbate as an antioxidant by grxB harbouring gram-negative bacteria, including human pathogens, may provide a survival advantage under hostile oxidative environments.

scholarly journals Interaction of Antimicrobial Peptide Temporin L with Lipopolysaccharide In Vitro and in Experimental Rat Models of Septic Shock Caused by Gram-Negative Bacteria

2006 ◽  
Vol 50 (7) ◽  
pp. 2478-2486 ◽  
Author(s):  
Andrea Giacometti ◽  
Oscar Cirioni ◽  
Roberto Ghiselli ◽  
Federico Mocchegiani ◽  
Fiorenza Orlando ◽  
...  
Keyword(s):  
Septic Shock ◽  
Antimicrobial Peptide ◽  
Gram Negative Bacteria ◽  
Amphibian Skin ◽  
Necrosis Factor Alpha ◽  
Gram Negative ◽  
Rat Models ◽  
E Coli ◽  
Tnf Α

ABSTRACT Sepsis remains a major cause of morbidity and mortality in hospitalized patients, despite intense efforts to improve survival. The primary lead for septic shock results from activation of host effector cells by endotoxin, the lipopolysaccharide (LPS) associated with cell membranes of gram-negative bacteria. For these reasons, the quest for compounds with antiendotoxin properties is actively pursued. We investigated the efficacy of the amphibian skin antimicrobial peptide temporin L in binding Escherichia coli LPS in vitro and counteracting its effects in vivo. Temporin L strongly bound to purified E. coli LPS and lipid A in vitro, as proven by fluorescent displacement assay, and readily penetrated into E. coli LPS monolayers. Furthermore, the killing activity of temporin L against E. coli was progressively inhibited by increasing concentrations of LPS added to the medium, further confirming the peptide's affinity for endotoxin. Antimicrobial assays showed that temporin L interacted synergistically with the clinically used β-lactam antibiotics piperacillin and imipenem. Therefore, we characterized the activity of temporin L when combined with imipenem and piperacillin in the prevention of lethality in two rat models of septic shock, measuring bacterial growth in blood and intra-abdominal fluid, endotoxin and tumor necrosis factor alpha (TNF-α) concentrations in plasma, and lethality. With respect to controls and single-drug treatments, the simultaneous administration of temporin L and β-lactams produced the highest antimicrobial activities and the strongest reduction in plasma endotoxin and TNF-α levels, resulting in the highest survival rates.

Antibacterial Function of Chromium Nanoparticles Against K. Pneumonia, E. coli and P. typhus

2020 ◽  
Author(s):  
Giriraj Tailor ◽  
Jyoti Chaudhary ◽  
Ajit Joshi ◽  
Deepshikha Verma ◽  
Osahon Michael
Keyword(s):  
Antibacterial Activity ◽  
Decomposition Method ◽  
Clinical Importance ◽  
Gram Negative Bacteria ◽  
Human Pathogens ◽  
Zone Of Inhibition ◽  
Plate Method ◽  
E Coli ◽  
Thermal Decomposition Method ◽  
Chromium Nanoparticles

Abstract The bioactive chromium nanoparticles were synthesized by calcination followed by thermal decomposition method. The antibacterial activity of chromium nanoparticles diffused in Dimethyl sulphoxide (DMSO). The antibacterial activity of chromium nanoparticles carried out against significant human pathogens (gram negative bacteria) viz, K. pneumonia, E. coli and P. typhus using agar diffusion cup plate method at 100 µg/ml concentration. The highest zone of inhibition was observed (12.0 mm) against K. pneumonia and lowest zone of inhibition (7.0 mm) E. coli. Thus, the outcomes of these studies suggest that synthesized chromium nanoparticles are of clinical importance.

Synthesis and in vitro Biological Activity of New 4,6-Disubstituted 3(2H)-Pyridazinone-acetohydrazide Derivatives

2012 ◽  
Vol 67 (5-6) ◽  
pp. 257-265
Author(s):  
Murat Sukuroglu ◽  
Tijen Onkol ◽  
Fatma Kaynak Onurdağ ◽  
Gulsen Akalın ◽  
M. Fethi Şahin
Keyword(s):  
Biological Activity ◽  
Antimicrobial Activities ◽  
Cytotoxic Activities ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
E Coli ◽  
Preliminary Results ◽  
H Nmr ◽  
Pyridazinone Derivatives

New 3(2H)-pyridazinone derivatives containing a N’-benzyliden-acetohydrazide moiety at position 2 were synthesized. The structures of these newly synthesized compounds were confi rmed by IR, 1H NMR, and MS data. These compounds were tested for their antibacterial, antifungal, antimycobacterial, and cytotoxic activities. The compounds 2-[4-(4-chlorophenyl)- 6-(morpholin-4-yl)-3-oxo-(2H)-pyridazin-2-yl]-N’-(4-tert-butylbenzyliden)acetohydrazide and 2-[4-(4-chlorophenyl)-6-(morpholin-4-yl)-3-oxo-(2H)-pyridazin-2-yl]-N’-(4-chlorobenzyliden) acetohydrazide exhibited activity against both Gram-positive and Gram-negative bacteria. Most of the compounds were active against E. coli ATCC 35218. The preliminary results of this study revealed that some target compounds exhibited promising antimicrobial activities

scholarly journals Membrane Protein Complex ExbB4-ExbD1-TonB1from Escherichia coli Demonstrates Conformational Plasticity

2015 ◽  
Vol 197 (11) ◽  
pp. 1873-1885 ◽  
Author(s):  
Aleksandr Sverzhinsky ◽  
Jacqueline W. Chung ◽  
Justin C. Deme ◽  
Lucien Fabre ◽  
Kristian T. Levey ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Iron Acquisition ◽  
Proton Translocation ◽  
Three Dimensional ◽  
Structural Data ◽  
Structural Features ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Content Type

ABSTRACTIron acquisition at the outer membrane (OM) of Gram-negative bacteria is powered by the proton motive force (PMF) of the cytoplasmic membrane (CM), harnessed by the CM-embedded complex of ExbB, ExbD, and TonB. Its stoichiometry, ensemble structural features, and mechanism of action are unknown. By panning combinatorial phage libraries, periplasmic regions of dimerization between ExbD and TonB were predicted. Using overexpression of full-length His6-taggedexbB-exbDand S-taggedtonB, we purified detergent-solubilized complexes of ExbB-ExbD-TonB fromEscherichia coli. Protein-detergent complexes of ∼230 kDa with a hydrodynamic radius of ∼6.0 nm were similar to previously purified ExbB4-ExbD2complexes. Significantly, they differed in electronegativity by native agarose gel electrophoresis. The stoichiometry was determined to be ExbB4-ExbD1-TonB1. Single-particle electron microscopy agrees with this stoichiometry. Two-dimensional averaging supported the phage display predictions, showing two forms of ExbD-TonB periplasmic heterodimerization: extensive and distal. Three-dimensional (3D) particle classification showed three representative conformations of ExbB4-ExbD1-TonB1. Based on our structural data, we propose a model in which ExbD shuttles a proton across the CM via an ExbB interprotein rearrangement. Proton translocation would be coupled to ExbD-mediated collapse of extended TonB in complex with ligand-loaded receptors in the OM, followed by repositioning of TonB through extensive dimerization with ExbD. Here we present the first report for purification of the ExbB-ExbD-TonB complex, molar ratios within the complex (4:1:1), and structural biology that provides insights into 3D organization.IMPORTANCEReceptors in the OM of Gram-negative bacteria allow entry of iron-bound siderophores that are necessary for pathogenicity. Numerous iron-acquisition strategies rely upon a ubiquitous and unique protein for energization: TonB. Complexed with ExbB and ExbD, the Ton system links the PMF to OM transport. Blocking iron uptake by targeting a vital nanomachine holds promise in therapeutics. Despite much research, the stoichiometry, structural arrangement, and molecular mechanism of the CM-embedded ExbB-ExbD-TonB complex remain unreported. Here we demonstratein vitroevidence of ExbB4-ExbD1-TonB1complexes. Using 3D EM, we reconstructed the complex in three conformational states that show variable ExbD-TonB heterodimerization. Our structural observations form the basis of a model for TonB-mediated iron acquisition.

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 Trends in the Susceptibility of Clinically Important Resistant Bacteria to Tigecycline: Results from the TigecyclineIn VitroSurveillance in Taiwan Study, 2006 to 2010

2011 ◽  
Vol 56 (3) ◽  
pp. 1452-1457 ◽  
Author(s):  
Yen-Hsu Chen ◽  
Po-Liang Lu ◽  
Cheng-Hua Huang ◽  
Chun-Hsing Liao ◽  
Chin-Te Lu ◽  
...  
Keyword(s):  
Resistant Bacteria ◽  
Gram Negative Bacteria ◽  
Drug Resistant ◽  
Content Type ◽  
E Coli ◽  
Drug Resistant Bacteria ◽  
Vancomycin Resistant ◽  
Susceptibility Rate ◽  
Important Drug

ABSTRACTThe TigecyclineIn VitroSurveillance in Taiwan (TIST) study, a nationwide, prospective surveillance during 2006 to 2010, collected a total of 7,793 clinical isolates, including methicillin-resistantStaphylococcus aureus(MRSA) (n= 1,834), penicillin-resistantStreptococcus pneumoniae(PRSP) (n= 423), vancomycin-resistant enterococci (VRE) (n= 219), extended-spectrum β-lactamase (ESBL)-producingEscherichia coli(n= 1,141), ESBL-producingKlebsiella pneumoniae(n= 1,330),Acinetobacter baumannii(n= 1,645), andStenotrophomonas maltophilia(n= 903), from different specimens from 20 different hospitals in Taiwan. MICs of tigecycline were determined following the criteria of the U.S. Food and Drug Administration (FDA) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST-2011). Among drug-resistant Gram-positive pathogens, all of the PRSP isolates were susceptible to tigecycline (MIC90, 0.03 μg/ml), and only one MRSA isolate (MIC90, 0.5 μg/ml) and three VRE isolates (MIC90, 0.125 μg/ml) were nonsusceptible to tigecycline. Among the Gram-negative bacteria, the tigecycline susceptibility rates were 99.65% for ESBL-producingE. coli(MIC90, 0.5 μg/ml) and 96.32% for ESBL-producingK. pneumoniae(MIC90, 2 μg/ml) when interpreted by FDA criteria but were 98.7% and 85.8%, respectively, when interpreted by EUCAST-2011 criteria. The susceptibility rate forA. baumannii(MIC90, 4 μg/ml) decreased from 80.9% in 2006 to 55.3% in 2009 but increased to 73.4% in 2010. A bimodal MIC distribution was found among carbapenem-susceptibleA. baumanniiisolates, and a unimodal MIC distribution was found among carbapenem-nonsusceptibleA. baumanniiisolates. In Taiwan, tigecycline continues to have excellentin vitroactivity against several major clinically important drug-resistant bacteria, with the exception ofA. baumannii.

scholarly journals A Campylobacter jejuni znuA Orthologue Is Essential for Growth in Low-Zinc Environments and Chick Colonization

2008 ◽  
Vol 191 (5) ◽  
pp. 1631-1640 ◽  
Author(s):  
Lindsay M. Davis ◽  
Tsutomu Kakuda ◽  
Victor J. DiRita
Keyword(s):  
Campylobacter Jejuni ◽  
Transport System ◽  
The United States ◽  
Gram Negative Bacteria ◽  
Limited Growth ◽  
E Coli ◽  
Poultry Products ◽  
Abc Transport ◽  
Bacterial Gastroenteritis

ABSTRACT Campylobacter jejuni infection is a leading cause of bacterial gastroenteritis in the United States and is acquired primarily through the ingestion of contaminated poultry products. Here, we describe the C. jejuni orthologue of ZnuA in other gram-negative bacteria. ZnuA (Cj0143c) is the periplasmic component of a putative zinc ABC transport system and is encoded on a zinc-dependent operon with Cj0142c and Cj0141c, which encode the other two likely components of the transport system of C. jejuni. Transcription of these genes is zinc dependent. A mutant lacking Cj0143c is growth deficient in zinc-limiting media, as well as in the chick gastrointestinal tract. The protein is glycosylated at asparagine 28, but this modification is dispensable for zinc-limited growth and chick colonization. Affinity-purified FLAG-tagged Cj0143c binds zinc in vitro. Based on our findings and on its homology to E. coli ZnuA, we conclude that Cj0143c encodes the C. jejuni orthologue of ZnuA.

scholarly journals A Novel Cd(II) Isophthalate Complex with Triethanolamine: Crystal Structure, Fluorescence and Antimicrobial Activity

2021 ◽  
Vol 68 (2) ◽  
pp. 466-474
Author(s):  
Zuhal Yolcu ◽  
Sinem Yurtcan ◽  
Meryem Çıtlakoğlu
Keyword(s):  
Antimicrobial Activity ◽  
Room Temperature ◽  
Diffusion Method ◽  
Gram Negative Bacteria ◽  
Single Crystal Diffraction ◽  
X Ray ◽  
Ft Ir ◽  
First Time ◽  
Trigonal Prismatic

A mixed ligand Cd(II) complex [Cd(IsoPht)(TEA)H2O]·3H2O was synthesized for the first time by using isophthalic acid (H2IsoPht) and tetradentate triethanolamine (TEA) and characterized by X-ray single-crystal diffraction, FT-IR, and thermogravimetric analysis (TGA). This novel complex crystallizes in the triclinic system with P-1 space group and distorted monocapped trigonal prismatic geometry. The Cd(II) has seven coordinates with bidentate IsoPht, a TEA in the tetradentate mode, and an aqua ligand. The fluorescence properties of the Cd(II) complex and TEA ligand were investigated at room temperature. The present Cd(II) complex was also tested for its antimicrobial activity by in vitro agar diffusion method against some Gram-positive and Gram-negative bacteria and a fungus.

Researches on the intestinal protozoa of monkeys and man

Parasitology ◽  
1943 ◽  
Vol 35 (3) ◽  
pp. 134-158 ◽  
Author(s):  
Clifford Dobell
Keyword(s):  
Life History ◽  
Special Reference ◽  
Entamoeba Histolytica ◽  
Cyst Wall ◽  
Equatorial Plate ◽  
Intestinal Protozoa ◽  
E Coli ◽  
Additional Information ◽  
First Time

1. This memoir amplifies my accounts ofEndolimax nanapublished in 1919 and 1933. The complete life-history, as observable in cultures, is now described—special attention being devoted to the nuclear structure at all stages.2. Additional information is also given about methods of cultivation, viability of the cystsin vitro, the ‘nucleal reaction’, natural parasites (Sphaerita, etc.), and other details.3. The life-history has been found to comprise trophic, precystic, cystic, and metacystic stages, comparable with those described inEntamoeba histolytica(1928) andE. coli(1938).4. The trophic amoeba is redescribed, with special reference to the polymorphism of its karyosome.5. Division of the trophic amoeba (by binary fission) has been studied, and the mitosis of its nucleus is now first described in detail. It is characterized by the formation of an intranuclear spindle, with well-developed centrioles and centrodesmus. The granular chromosomes are 10 in number, and arranged in a ring on the equatorial plate.6. Precystic amoebae are smaller than trophic forms —the reduction in size being effected during the last two divisions before encystation.7. The cyst is formed by a precystic amoeba in the usual way. Inside the cyst, the nucleus undergoes two successive mitotic divisions similar to those seen in trophic forms. The ripe cyst is therefore 4-micleate, though supernucleate specimens—which are viable— occur occasionally. Glycogen is normally present in young (1- and 2-nucleate) cysts, but not in those which are mature.8. The process of excystation is described for the first time, and closely resembles that seen inE. histolytica. The entire protoplasmic contents emerge as a single 4-nucleate amoeba through a minute pore in the cyst wall.9. Metacystic development—not previously studied —consists in simple division, by successive cytoplasmic bipartitions without division of the nuclei, into 4 uninucleate amoebulae. These grow into trophic amoebae once more.10. The whole life-history is thus asexual, with no gametes, conjugation, or autogamy, at any stage.

Population Reductions of Gram-Negative Pathogens Following Treatments with Nisin and Chelators under Various Conditions

1995 ◽  
Vol 58 (9) ◽  
pp. 977-983 ◽  
Author(s):  
CATHERINE N. CUTTER ◽  
GREGORY R. SIRAGUSA
Keyword(s):  
Divalent Cations ◽  
Escherichia Coli O157 ◽  
Gram Negative Bacteria ◽  
Bacterial Populations ◽  
Gram Negative ◽  
E Coli ◽  
Transmission Electron ◽  
Colony Forming Units ◽  
Citrate Phosphate

When used in combination with chelating agents (EDTA, EGTA, citrate, phosphate), the bacteriocin nisin is effective for reducing populations of gram-negative bacteria in vitro. This study examined parameters (buffers, temperature presence of divalent cations) that affect nisin inhibition of Escherichia coli O157:H7 and Salmonella typhimurium. Approximately 7 log10 colony-forming units (CFU) per ml of E. coli and S. typhimurium were treated in PBS or MOPS buffers containing 50 μg/ml of purified nisin, alone or in combination with 500 mM lactate, 100 mM citrate, 50 mM EDTA, and 1% (wt/vol) sodium hexametaphosphate (pH 7.0) at 37°C for 60 min or 5°C for 30 min. Surviving bacterial populations were compared to untreated controls (buffers without nisin). Data indicated that treatments with nisin in buffers resulted in reductions of 4.30 and 2.30 log10 CFU/ml of E. coli and S. typhimurium, respectively, as compared to untreated controls. Population reductions ranging from 2.29 to 5.49 log10 CFU/ml were observed when cells were treated with nisin and chelator combinations at either 37°C for 60 min or 5°C for 30 min. The addition of magnesium and calcium to buffers with nisin decreased inhibition. Data obtained from spectrophotometric experiments indicated that treatments were causing the release of cellular constituents. However, transmission electron microscopy (TEM) analyses were inconclusive, since cellular membranes did not appear to be disrupted.

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