scholarly journals Altered Envelope Structure and Nanomechanical Properties of a C-Terminal Protease A-Deficient Rhizobium leguminosarum

2020 ◽  
Vol 8 (9) ◽  
pp. 1421
Author(s):  
Dong Jun ◽  
Ubong Idem ◽  
Tanya E. S. Dahms
Keyword(s):  
Outer Membrane ◽  
Creep Deformation ◽  
High Performance ◽  
Rhizobium Leguminosarum ◽  
Cell Envelope ◽  
Time Dependent ◽  
Wild Type ◽  
Force Microscopy ◽  
Viscoelastic Parameters ◽  
Transmission Electron

(1) Background: Many factors can impact bacterial mechanical properties, which play an important role in survival and adaptation. This study characterizes the ultrastructural phenotype, elastic and viscoelastic properties of Rhizobium leguminosarum bv. viciae 3841 and the C-terminal protease A (ctpA) null mutant strain predicted to have a compromised cell envelope; (2) Methods: To probe the cell envelope, we used transmission electron microscopy (TEM), high performance liquid chromatography (HPLC), mass spectrometry (MS), atomic force microscopy (AFM) force spectroscopy, and time-dependent AFM creep deformation; (3) Results: TEM images show a compromised and often detached outer membrane for the ctpA mutant. Muropeptide characterization by HPLC and MS showed an increase in peptidoglycan dimeric peptide (GlcNAc-MurNAc-Ala-Glu-meso-DAP-Ala-meso-DAP-Glu-Ala-MurNAc-GlcNAc) for the ctpA mutant, indicative of increased crosslinking. The ctpA mutant had significantly larger spring constants than wild type under all hydrated conditions, attributable to more highly crosslinked peptidoglycan. Time-dependent AFM creep deformation for both the wild type and ctpA mutant was indicative of a viscoelastic cell envelope, with best fit to the four-element Burgers model and generating values for viscoelastic parameters k1, k2, η1, and η2; (4) Conclusions: The viscoelastic response of the ctpA mutant is consistent with both its compromised outer membrane (TEM) and fortified peptidoglycan layer (HPLC/MS).

scholarly journals Mutagenesis of the rpoS gene involved in alteration of outer membrane composition

2019 ◽  
Author(s):  
Sayyed Shahryar Rahpeyma ◽  
Jamshid Raheb
Keyword(s):  
Outer Membrane ◽  
Sigma Factor ◽  
Critical Role ◽  
Membrane Composition ◽  
Wild Type ◽  
Rpos Gene ◽  
Suicide Vector ◽  
Transmission Electron ◽  
Bacterial Morphology ◽  
Suitable Vector

Background and Objectives: rpoS is a bacterial sigma factor of RNA polymerase which is involved in the expression of the genes which control regulons and play a critical role in survival against stresses. Few suitable vectors are available which could be maintained successfully in Flexibacter chinesis cells and could in particular be used as a suicide vector to make mutation in the rpoS gene. The aim of this study was to investigate if rpoS mutagenesis has impact on bacterial morphology in addition to cell division. Materials and Methods: A 0.603 kb BamHI-PstI fragment subclone of pICRPOS38Ω was cloned into linearized pLYLO3. The final construct, pLRPOS38 suicide vector, was introduced into Flexibacter chinesis. Then the cytoplasm of mutant strain and wild-type were investigated by transmission electron microscopy. Results: After successful subcloning of suicide vector into F. chinesis, based on TEM study, it was demonstrated that muta- tion in rpoS gene leads to decomposition of outer membrane of F. chinesis. Conclusion: A suitable vector to make suicide mutation in rpoS was constructed for F. chinesi.

scholarly journals Roles of Internal Cysteines in the Function, Localization, and Reactivity of the TraV Outer Membrane Lipoprotein Encoded by the F Plasmid

2002 ◽  
Vol 184 (11) ◽  
pp. 3126-3129 ◽  
Author(s):  
Robin L. Harris ◽  
Philip M. Silverman
Keyword(s):  
Outer Membrane ◽  
Cell Envelope ◽  
Osmotic Shock ◽  
Wild Type ◽  
F Plasmid ◽  
Outer Membrane Lipoprotein ◽  
Numerous Cell ◽  
Mixed Disulfides ◽  
Membrane Lipoprotein

ABSTRACT We have examined the functional role of two internal cysteine residues of the F-plasmid TraV outer membrane lipoprotein. Each was mutated to a serine separately and together to yield three mutant traV genes: traV C10S, traV C18S, and traV C10S/C18S. All three cysteine mutations complemented a traV mutant for DNA donor activity and for sensitivity to donor-specific bacteriophage; however, when measured by a transduction assay, the donor-specific DNA bacteriophage sensitivities of the traV C18S and, especially, traV C10S/C18S mutant strains were significantly less than those of the traV + and traV C10S strains. Thus, unlike the Agrobacterium tumefaciens T-plasmid-encoded VirB7 outer membrane lipoprotein, TraV does not require either internal cysteine to retain significant biological activity. By Western blot analysis, all three mutant TraV proteins were shown to accumulate in the outer membrane. However, by nonreducing gel electrophoresis, wild-type TraV and especially the TraVC18S mutant were shown to form mixed disulfides with numerous cell envelope proteins. This was not observed with the TraVC10S or TraVC10S/C18S proteins. Thus, it appears that TraV C10 is unusually reactive and that this reactivity is reduced by C18, perhaps by intramolecular oxidation. Finally, whereas the TraVC10S and TraVC18S proteins fractionated primarily with the outer membrane, as did the wild-type protein, the TraVC10S/C18S protein was found in osmotic shock fluid and inner membrane fractions as well as outer membrane fractions. Hence, at least one cysteine is required for the efficient localization of TraV to the outer membrane.

scholarly journals Growth Temperature Regulation of Host-Specific Modifications of Rhizobial Lipo-Chitin Oligosaccharides: The Function of nodX Is Temperature Regulated

2000 ◽  
Vol 13 (8) ◽  
pp. 808-820 ◽  
Author(s):  
Maurien M. A. Olsthoorn ◽  
Ellen Stokvis ◽  
Johan Haverkamp ◽  
Herman P. Spaink ◽  
Jane E. Thomas-Oates
Keyword(s):  
Growth Temperature ◽  
High Performance ◽  
Rhizobium Leguminosarum ◽  
High Performance Liquid Chromatographic ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Temperature Dependent ◽  
Gene Encoding ◽  
Two Temperatures ◽  
Liquid Chromatographic

Lipo-chitin oligosaccharides (LCOs) are usually produced and isolated for structural analysis from bacteria cultured under laboratory rather than field conditions. We have studied the influence of bacterial growth temperature on the LCO structures produced by different Rhizobium leguminosarum strains, using thin-layer chromatographic, high-performance liquid chromatographic, and mass spectrometric analyses. Wild-type R. leguminosarum bv. viciae A1 was shown to produce larger relative amounts of nodX-mediated, acetylated LCOs at 12°C than at 28°C, indicating that the activity of nodX (a gene encoding an LCO O-acetyl transferase) is temperature dependent. Interestingly, symbiotic resistance genes sym1 and sym2 found in primitive pea cultivars are also temperature sensitive, only being active at low temperatures, at which they block nodulation by R. leguminosarum bv. viciae strains lacking nodX. We therefore propose that the gene-for-gene relationship between plant and bacterium has a temperature-sensitive mechanism as an adaptation to environmental conditions. An R. leguminosarum bv. trifolii strain was also shown to produce larger relative amounts of nodX-mediated, acetylated LCOs at 12°C than at 28°C. The major components synthesized by the two strains are produced at both temperatures but in different relative amounts, while some minor components are only produced at one of the two temperatures.

scholarly journals Bean and Pea Plastoglobules Change in Response to Chilling Stress

2021 ◽  
Vol 22 (21) ◽  
pp. 11895
Author(s):  
Joanna Wójtowicz ◽  
Joanna Grzyb ◽  
Joanna Szach ◽  
Radosław Mazur ◽  
Katarzyna B. Gieczewska
Keyword(s):  
Plant Species ◽  
High Performance ◽  
Chilling Stress ◽  
Spatial Proximity ◽  
Force Microscopy ◽  
Stage Of Development ◽  
Atomic Force ◽  
Metabolic Channelling ◽  
Transmission Electron ◽  
Insight Into

Plastoglobules (PGs) might be characterised as microdomains of the thylakoid membrane that serve as a platform to recruit proteins and metabolites in their spatial proximity in order to facilitate metabolic channelling or signal transduction. This study provides new insight into changes in PGs isolated from two plant species with different responses to chilling stress, namely chilling-tolerant pea (Pisum sativum) and chilling-sensitive bean (Phaseolus coccineus). Using multiple analytical methods, such as high-performance liquid chromatography and visualisation techniques including transmission electron microscopy and atomic force microscopy, we determined changes in PGs’ biochemical and biophysical characteristics as a function of chilling stress. Some of the observed alterations occurred in both studied plant species, such as increased particle size and plastoquinone-9 content, while others were more typical of a particular type of response to chilling stress. Additionally, PGs of first green leaves were examined to highlight differences at this stage of development. Observed changes appear to be a dynamic response to the demands of photosynthetic membranes under stress conditions.

Characterization of cell surface properties in agglutinable and nonagglutinable mutants of Pseudomonas putida

1993 ◽  
Vol 39 (8) ◽  
pp. 787-794 ◽  
Author(s):  
C. R. Buell ◽  
R. Whetton ◽  
P. Tari ◽  
A. J. Anderson
Keyword(s):  
Molecular Weight ◽  
Cell Surface ◽  
Pseudomonas Putida ◽  
Outer Membrane ◽  
Cell Envelope ◽  
Surface Glycoprotein ◽  
Chemical Mutagenesis ◽  
Wild Type ◽  
Surface Binding ◽  
Surface Features

Cells of an aggressive, root-colonizing isolate of Pseudomonas putida are agglutinated by a root surface glycoprotein. The agglutination phenotype in P. putida isolate Corvallis is lacking in mutants (Agg−) derived by Tn5 insertion and chemical mutagenesis. Specific mutation in the aggA locus by Tn5 insertion results in loss of agglutinability that is complemented in trans by a wild-type copy of the P. putida aggA locus. We examined the biochemical bases of agglutination in P. putida by comparing cell surface features in Agg+, Agg− mutants, and a genetically restored aggA mutant. No changes in gross cell surface features involving hydrophobic or hydrophilic binding or net negative charge were observed. Three macromolecular features, pili, flagella, and lipopolysaccharide size, did not differ between Agg+ and Agg− mutants. Protein profiles of cell envelope, periplasmic, and outer membrane preparations revealed pleiotropic effects of mutation in agglutination phenotype including alterations of an outer membrane protein of 47 000 molecular weight and periplasmic proteins of 56 000 and 60 000 molecular weight. The protein alterations seen in the aggA::Tn5 Agg− mutant 5123 reverted to wild-type patterns upon introduction of a wild-type copy of the aggA locus. These data suggest agglutinability may be conditioned by more than one proteinaceous component associated with the bacterial envelope layers.Key words: cell surface, binding, recognition.

scholarly journals Extracellular Reduction of Hexavalent Chromium by Cytochromes MtrC and OmcA of Shewanella oneidensis MR-1

2011 ◽  
Vol 77 (12) ◽  
pp. 4035-4041 ◽  
Author(s):  
Sara M. Belchik ◽  
David W. Kennedy ◽  
Alice C. Dohnalkova ◽  
Yuanmin Wang ◽  
Papatya C. Sevinc ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Extracellular Matrix ◽  
Outer Membrane ◽  
Initial Rate ◽  
Mutant Cell ◽  
Shewanella Oneidensis ◽  
Wild Type ◽  
Content Type ◽  
Transmission Electron ◽  
Mutant Cells

ABSTRACTTo characterize the roles of cytochromes MtrC and OmcA ofShewanella oneidensisMR-1 in Cr(VI) reduction, the effects of deleting themtrCand/oromcAgene on Cr(VI) reduction and the cellular locations of reduced Cr(III) precipitates were investigated. Compared to the rate of reduction of Cr(VI) by the wild type (wt), the deletion ofmtrCdecreased the initial rate of Cr(VI) reduction by 43.5%, while the deletion ofomcAor bothmtrCandomcAlowered the rate by 53.4% and 68.9%, respectively. In wt cells, Cr(III) precipitates were detected by transmission electron microscopy in the extracellular matrix between the cells, in association with the outer membrane, and inside the cytoplasm. No extracellular matrix-associated Cr(III) precipitates, however, were found in the cytochrome mutant cell suspension. In mutant cells without either MtrC or OmcA, most Cr(III) precipitates were found in association with the outer membrane, while in mutant cells lacking both MtrC and OmcA, most Cr(III) precipitates were found inside the cytoplasm. Cr(III) precipitates were also detected by scanning election microscopy on the surfaces of the wt and mutants without MtrC or OmcA but not on the mutant cells lacking both MtrC and OmcA, demonstrating that the deletion ofmtrCandomcAdiminishes the extracellular formation of Cr(III) precipitates. Furthermore, purified MtrC and OmcA reduced Cr(VI) with apparentkcatvalues of 1.2 ± 0.2 (mean ± standard deviation) and 10.2 ± 1 s−1andKmvalues of 34.1 ± 4.5 and 41.3 ± 7.9 μM, respectively. Together, these results consistently demonstrate that MtrC and OmcA are the terminal reductases used byS. oneidensisMR-1 for extracellular Cr(VI) reduction where OmcA is a predominant Cr(VI) reductase.

scholarly journals Preparation of a Hybrid Membrane from Whey Protein Fibrils and Activated Carbon to Remove Mercury and Chromium from Water

Membranes ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 386
Author(s):  
Laura Cristina Ramírez-Rodríguez ◽  
Luis Eduardo Díaz Barrera ◽  
María Ximena Quintanilla-Carvajal ◽  
Didilia Ileana Mendoza-Castillo ◽  
Adrián Bonilla-Petriciolet ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Activated Carbon ◽  
Whey Protein ◽  
High Performance ◽  
Membrane Filtration ◽  
Hybrid Membrane ◽  
Force Microscopy ◽  
Adsorption Capacities ◽  
Transmission Electron ◽  
Protein Fibrils

Water contamination by mercury and chromium has a direct effect in human health. A promising technology to remove heavy metals by membrane filtration is the use of hybrid membranes produced with whey protein fibrils (WPF) and activated carbon (AC). In this study, the best conditions to produce WPF by heat treatment were determined to maximize the removal of mercury and chromium from water using a central composed design. The results indicated that the best conditions to prepare WPF were 74 °C, 7 h and 3.8% of whey protein with adsorption capacities of 25 and 18 mg/g and removal efficiencies of 81 and 57% for mercury and chromium, respectively. WPF and AC were used to prepare a hybrid membrane that was characterized using transmission electron microscopy, atomic force microscopy, scanning electron microscopy, Fourier transform infrared spectroscopy and Brunauer–Emmett–Teller surface area measurements. Batch filtration experiments were performed with the hybrid membrane for chromium and mercury removal at 25, 50 and 100 mg/L to determine its adsorption capacities. A high performance of the hybrid membrane was demonstrated removing efficiently mercury and chromium from water, thus supporting more than ten filtration cycles.

Sensitivity of Escherichia coli to Viral Nucleic Acid, XII. Ca2+-or Ba2+-Facilitated Transfection of Cell Envelope Mutants

1977 ◽  
Vol 32 (5-6) ◽  
pp. 429-433 ◽  
Author(s):  
Akira Taketo
Keyword(s):  
Escherichia Coli ◽  
Nucleic Acid ◽  
Outer Membrane ◽  
Cell Envelope ◽  
Nucleoside Transport ◽  
Wild Type ◽  
Viral Nucleic Acid ◽  
E Coli ◽  
The Relationship ◽  
Cellular Competence

Abstract Using various envelope mutants of Escherichia coli, the relationship between cell surface struc­ture and the Ca2+-or Ba2+-dependent competence for transfection was investigated. In contrast with rough strains, smooth bacteria treated with Ca2+ or Ba2+ were incompetent for the trans­fection by ØA RF. In E. coli K12 D21 derivatives, Ca2+-dependent competence remarkably in­ creased by lpsAl mutation and the highest level of competence was attained by further deficiency in glucose units of the LPS. Upon treatment with BaCl2 , strain D21 and its lpsAl mutant became highly competent for ØA RF. The effect of Ba2+ was, however, feeble for lpsAl mutants further deficient in heptose units and/or glucose units. Among different LPS mutants of E. coli B, variation of the Ca2+-or Ba2+-dependent competence was relatively small and even the competence of strain BB12, whose LPS core contained only two KDO units, was nearly equal to that of wild type bacteria. However, the level of cellular competence induced by Ba2+ was not allways parallel to that induced by Ca2+. In mutants deficient in outer membrane protein I, either Ca2+-or Ba2+-dependent competence increased several-fold, whereas in mutants devoid of outer membrane II*, the com­petence decreased considerably. Unlike nucleoside transport, the uptake of DNA was not affected by tsx mutation.

scholarly journals Water Treatment Ability of CuO-ZnO Nanocomposites Synthesized by Laser Ablation and Anodization Techniques

2020 ◽  
Vol 55 (1) ◽  
Author(s):  
R.B. Rashid ◽  
D. H. Hussain ◽  
R. S. Mahmood
Keyword(s):  
Laser Ablation ◽  
High Performance ◽  
Pollutant Removal ◽  
Force Microscopy ◽  
Laser Ablation Method ◽  
Atomic Force ◽  
Transmission Electron ◽  
New Ideas ◽  
Diyala River ◽  
Anodization Method

The article describes new ideas about the differences in the pollutant-removal ability of nanocomposite prepared by two methods. Nanocomposites (CuO-ZnO) can be simply synthesized chemically in aqueous solution by anodization method or physically by a laser ablation method. Therefore, both anodization and laser ablation methods were investigated. UV-visible spectra, atomic force microscopy AFM, and transmission electron microscopy TEM were used for characterizations. Results indicated that nanoparticles prepared by laser ablation had less diameters (74nm) than that prepared by anodization method. TEM images showed that particles produced by laser ablation method were homogeneous and core-shell structures with no aggregation, but that produced by anodization method were aggregated. This result could suggest that the laser ablation method is specifically preferable. The nanoparticles diameters ranged from 10–17 nm, which may confirm that ZnO nanoparticles have successfully shielded the CuO nanocore. Results also indicated that these synthesized nanocomposites have actively removed pollutions from water samples of Diyala River. There is a clear difference between the activity of laser ablation nanocomposite and that of anodization composite. Laser ablation nanocomposite needs less than 60 min to treat 85% of water pollution, but with anodization composite, even after 90 min. the water sample is still polluted (30% treated). Laser ablation nanocomposites give high performance of removing pollutants.

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