scholarly journals Outer-membrane siderophore receptors of heterotrophic oceanic bacteria

2004 ◽  
Vol 49 (2) ◽  
pp. 579-587 ◽  
Author(s):  
Evelyn Armstrong ◽  
Julie Granger ◽  
Elizabeth L. Mann ◽  
Neil M. Price
Keyword(s):  
Outer Membrane ◽  
Siderophore Receptors

scholarly journals Thiocillin and Micrococcin Exploit the Ferrioxamine Receptor of Pseudomonas aeruginosa for Uptake

2020 ◽  
Author(s):  
Derek C. K. Chan ◽  
Lori L. Burrows
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Outer Membrane ◽  
Inhibitory Concentration ◽  
Iron Chelator ◽  
Inhibit Protein Synthesis ◽  
Gram Negative ◽  
Siderophore Receptors ◽  
Ferrioxamine Receptor ◽  
Gain Access

ABSTRACTThiopeptides are a class of Gram-positive antibiotics that inhibit protein synthesis. They have been underutilized as therapeutics due to solubility issues, poor bioavailability, and lack of activity against Gram-negative pathogens. We discovered recently that a member of this family, thiostrepton, has activity against Pseudomonas aeruginosa and Acinetobacter baumannii under iron-limiting conditions. Thiostrepton uses pyoverdine siderophore receptors to cross the outer membrane, and combining thiostrepton with an iron chelator yielded remarkable synergy, significantly reducing the minimal inhibitory concentration. These results led to the hypothesis that other thiopeptides could also inhibit growth by using siderophore receptors to gain access to the cell. Here, we screened six thiopeptides for synergy with the iron chelator deferasirox against P. aeruginosa and a mutant lacking the pyoverdine receptors FpvA and FpvB. Our findings suggest that thiopeptides such as thiocillin cross the outer membrane using FoxA, the ferrioxamine siderophore receptor. Other structurally related thiopeptides did not inhibit growth of P. aeruginosa, but had greater potency against methicillin-resistant Staphylococcus aureus than thiostrepton and related thiopeptides. These results suggest that thiopeptide structures have evolved with considerations for target affinity and entry into cells.

scholarly journals Role of Receptor Proteins for Enterobactin and 2,3-Dihydroxybenzoylserine in Virulence of Salmonella enterica

2003 ◽  
Vol 71 (12) ◽  
pp. 6953-6961 ◽  
Author(s):  
W. Rabsch ◽  
U. Methner ◽  
W. Voigt ◽  
H. Tschäpe ◽  
R. Reissbrodt ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Parental Strain ◽  
Salmonella Enterica ◽  
Growth Promotion ◽  
Outer Membrane Proteins ◽  
Breakdown Product ◽  
Triple Mutant ◽  
Siderophore Receptors ◽  
Systemic Spread ◽  
Serovar Typhimurium

ABSTRACT Single, double, and triple mutants of an enterobactin-deficient mutant strain of Salmonella enterica serovar Typhimurium were constructed that were defective in the expression of the iron-regulated outer membrane proteins (IROMPs) FepA, IroN, and Cir, which are proposed to function as catecholate receptors. Uptake of naturally occurring and chemically synthesized catecholate molecules by these mutants was assessed in standard growth promotion assays. Unique patterns of uptake were identified for each IROMP; specifically, FepA and IroN were confirmed to be required for transport of enterobactin, and all three proteins were shown to function as receptors for the enterobactin breakdown product 2,3-dihydroxybenzoylserine. The fepA, iroN, and cir alleles were transduced to enterobactin-proficient strains of S. enterica serovar Typhimurium and S. enterica serovar Enteritidis, and the resulting phenotypes were confirmed by analysis of outer membrane protein profiles, by sensitivity to KP-736, a catecholate-cephalosporin conjugate, and by growth promotion tests on egg white agar. Intragastric infections of mice with the S. enterica serovar Typhimurium strains indicated that the parental strain and the fepA iroN double mutant were similarly virulent but that the fepA iroN cir triple mutant was significantly attenuated. Moreover, in mixed infections, the fepA iroN mutant showed similar cecal colonization and invasion of the liver to the parental strain, while the triple mutant showed significantly reduced cecal colonization and no measurable spread to the liver. Infections of 4-day-old chicks with S. enterica serovar Enteritidis strains also indicated that mutation of the fepA iroN genes did not significantly reduce cecal colonization and systemic spread compared with those of the parental strain. The results indicate that, while enterobactin uptake is not essential for the virulence of S. enterica serovars in mouse and chicken infection models, the ability to take up 2,3-dihydroxybenzoylserine via any of the three catecholate siderophore receptors appears to play an important role, since the S. enterica serovar Typhimurium triple mutant was significantly attenuated in the mouse model. Salmochelins appear not to be involved in the virulence of S. enterica.

scholarly journals Thiocillin and micrococcin exploit the ferrioxamine receptor of Pseudomonas aeruginosa for uptake

2021 ◽  
Author(s):  
Derek C K Chan ◽  
Lori L Burrows
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antimicrobial Activity ◽  
Outer Membrane ◽  
Susceptibility Testing ◽  
Gram Negative Bacteria ◽  
Null Mutant ◽  
Translation Inhibition ◽  
Gram Positive Bacteria ◽  
Siderophore Receptors ◽  
Ferrioxamine Receptor

Abstract Background Thiopeptides are a class of antibiotics that are active against Gram-positive bacteria and inhibit translation. They were considered inactive against Gram-negative bacteria due to their inability to cross the outer membrane. However, we discovered previously that a member of this class, thiostrepton (TS), has activity against Pseudomonas aeruginosa and Acinetobacter baumannii under iron-limiting conditions. TS hijacks the pyoverdine siderophore receptors of P. aeruginosa to cross the outer membrane and synergizes with iron chelators. Objectives To test other thiopeptides for antimicrobial activity against P. aeruginosa and determine their mechanism of uptake, action and spectrum of activity. Methods Eight thiopeptides were screened in chequerboard assays against a mutant of P. aeruginosa PA14 lacking both pyoverdine receptors. Thiopeptides that retain activity against a pyoverdine receptor-null mutant may use alternative siderophore receptors for entry. Susceptibility testing against siderophore receptor mutants was used to determine thiopeptide mechanism of uptake. Results The thiopeptides thiocillin (TC) and micrococcin (MC) use the ferrioxamine siderophore receptor (FoxA) for uptake and inhibit the growth of P. aeruginosa at low micromolar concentrations. The activity of TC required the TonB-ExbBD system used to energize siderophore uptake. TC acted through its canonical mechanism of action of translation inhibition. Conclusions Multiple thiopeptides have antimicrobial activity against P. aeruginosa, countering the historical assumption that they cannot cross the outer membrane. These results demonstrate the potential for thiopeptides to act as antipseudomonal antibiotics.

Molecular Structure of the Outermost Cell Wall Component of Spirillum Serpens

1977 ◽  
Vol 35 ◽  
pp. 342-343
Author(s):  
Wah Chiu ◽  
David Grano
Keyword(s):  
Molecular Structure ◽  
Cell Wall ◽  
Outer Membrane ◽  
Gel Electrophoresis ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Cell Wall Component ◽  
Protein Components ◽  
Exposure Technique ◽  
Structural Aspects

The periodic structure external to the outer membrane of Spirillum serpens VHA has been isolated by similar procedures to those used by Buckmire and Murray (1). From SDS gel electrophoresis, we have found that the isolated fragments contain several protein components, and that the crystalline structure is composed of a glycoprotein component with a molecular weight of ∽ 140,000 daltons (2). Under an electron microscopic examination, we have visualized the hexagonally-packed glycoprotein subunits, as well as the bilayer profile of the outer membrane. In this paper, we will discuss some structural aspects of the crystalline glycoproteins, based on computer-reconstructed images of the external cell wall fragments.The specimens were prepared for electron microscopy in two ways: negatively stained with 1% PTA, and maintained in a frozen-hydrated state (3). The micrographs were taken with a JEM-100B electron microscope with a field emission gun. The minimum exposure technique was essential for imaging the frozen- hydrated specimens.

Cryo-Electron Microscopy and Correlation Averaging of Frozen-Hydrated, Polymorphic 2D Crystals of the Mitochondrial Outer Membrane Channel

1990 ◽  
Vol 48 (1) ◽  
pp. 100-101
Author(s):  
Xiao-Wei Guo
Keyword(s):  
Outer Membrane ◽  
Hexagonal Lattice ◽  
Mitochondrial Outer Membrane ◽  
Rectangular Lattice ◽  
Electron Microscopic ◽  
Cryo Electron Microscopy ◽  
Voltage Dependent ◽  
Outer Membrane Channel ◽  
2D Crystals ◽  
Vdac Channel

Voltage-dependent, anion-selective channels (VDAC) are formed in the mitochondrial outer membrane (mitOM) by a 30-kDa polypeptide. These channels form ordered 2D arrays when mitOMs from Neurospora crassa are treated with soluble phospholipase A2. We obtain low-dose electron microscopic images of unstained specimens of VDAC crystals preserved in vitreous ice, using a Philips EM420 equipped with a Gatan cryo-transfer stage. We then use correlation analysis to compute average projections of the channel crystals. The procedure involves Fourier-filtration of a region within a crystal field to obtain a preliminary average that is subsequently cross-correlated with the entire crystal. Subregions are windowed from the crystal image at coordinates of peaks in the cross-correlation function (CCF, see Figures 1 and 2) and summed to form averages (Figure 3).The VDAC channel forms several different types of crystalline arrays in mitOMs. The polymorph first observed during phospholipase treatment is a parallelogram array (a=13 run, b=11.5 run, θ==109°) containing 6 water-filled pores per unit cell. Figure 1 shows the CCF of a sub-field of such an “oblique” array used to compute the correlation average of Figure 3A. With increased phospholipase treatment, other polymorphs are observed, often co-existing within the same crystal. For example, two distinct (but closely related) types of lattices occur in the field corresponding to the CCF of Figure 2: a “contracted” version of the parallelogram lattice (a=13 run, b=10 run, θ=99°), and a near-rectangular lattice (a=8.5 run, b=5 nm). The pattern of maxima in this CCF suggests that a third, near-hexagonal lattice (a=4.5 nm) may also be present. The correlation averages of Figures 3B-D were computed from polycrystalline fields, using peak coordinates in regions of CCFs corresponding to each of the three lattice types.

Electrophysiological Characterization of Transport Across Outer Membrane Channels from Gram-Negative Bacteria in Their Native Environment

2019 ◽  
Author(s):  
Jiajun Wang ◽  
Rémi Terrasse ◽  
Jayesh Arun Bafna ◽  
Lorraine Benier ◽  
Mathias Winterhalter
Keyword(s):  
Outer Membrane ◽  
Lipid Membrane ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Multi Drug Resistance ◽  
Gram Negative Bacteria ◽  
Membrane Channels ◽  
Gram Negative ◽  
Electrophysiological Characterization

Multi-drug resistance in Gram-negative bacteria is often associated with low permeability of the outer membrane. To investigate the role of membrane channels in the uptake of antibiotics, we extract, purify and reconstitute them into artificial planar membranes. To avoid this time-consuming procedure, here we show a robust approach using fusion of native outer membrane vesicles (OMV) into planar lipid bilayer which moreover allows also to some extend the characterization of membrane protein channels in their native environment. Two major membrane channels from <i>Escherichia coli</i>, OmpF and OmpC, were overexpressed from the host and the corresponding OMVs were collected. Each OMV fusion revealed surprisingly single or only few channel activities. The asymmetry of the OMV´s translates after fusion into the lipid membrane with the LPS dominantly present at the side of OMV addition. Compared to conventional reconstitution methods, the channels fused from OMVs containing LPS have similar conductance but a much broader distribution. The addition of Enrofloxacin on the LPS side yields somewhat higher association (<i>k<sub>on</sub></i>) and lower dissociation (<i>k<sub>off</sub></i>) rates compared to LPS-free reconstitution. We conclude that using outer membrane vesicles is a fast and easy approach for functional and structural studies of membrane channels in the native membrane.

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