scholarly journals Interaction and localization studies of enteropathogenic Escherichia coli type IV bundle-forming pilus outer membrane components

Microbiology ◽  
2006 ◽  
Vol 152 (8) ◽  
pp. 2405-2420 ◽  
Author(s):  
Anu Daniel ◽  
Aparna Singh ◽  
Lynette J. Crowther ◽  
Paula J. Fernandes ◽  
Wiebke Schreiber ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Cytoplasmic Membrane ◽  
Complete Characterization ◽  
Type Iv Pili ◽  
Amino Terminus ◽  
Type Iv ◽  
Membrane Components ◽  
Nucleotide Binding Proteins

Typical enteropathogenic Escherichia coli strains express an established virulence factor belonging to the type IV pili family, called the bundle-forming pilus (BFP). BFP are present on the cell surface as bundled filamentous appendages, and are assembled and retracted by proteins encoded by the bfp operon. These proteins assemble to form a molecular machine. The BFP machine may be conceptually divided into three components: the cytoplasmic membrane (CM) subassembly, which is composed of CM proteins and cytoplasmic nucleotide-binding proteins; the outer membrane (OM) subassembly and the pilus itself. The authors have previously characterized the CM subassembly and the pilus. In this study, a more complete characterization of the OM subassembly was carried out using a combination of biochemical, biophysical and genetic approaches. It is reported that targeting of BfpG to the OM was influenced by the secretin BfpB. BfpG and BfpU interacted with the amino terminus of BfpB. BfpU had a complex cellular distribution pattern and, along with BfpB and BfpG, was part of the OM subassembly.

scholarly journals Structural Characterization of Outer Membrane Components of the Type IV Pili System in Pathogenic Neisseria

PLoS ONE ◽  
2011 ◽  
Vol 6 (1) ◽  
pp. e16624 ◽  
Author(s):  
Samta Jain ◽  
Katarzyna B. Mościcka ◽  
Martine P. Bos ◽  
Emilia Pachulec ◽  
Marc C. A. Stuart ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Structural Characterization ◽  
Type Iv Pili ◽  
Type Iv ◽  
Membrane Components

scholarly journals Cryo-ET Characterization of Novel Cellular Extrusions in Escherichia coli Induced by the Major Subunit Protein of Type IV Pili, PilA, from Pseudomonas aeruginosa

2021 ◽  
Vol 27 (S1) ◽  
pp. 280-282
Author(s):  
Juan Sanchez ◽  
Daniel Parrell ◽  
Alba Gonzalez-Rivera ◽  
Nicoleta Ploscariu ◽  
Katrina Forest ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Type Iv Pili ◽  
Subunit Protein ◽  
Type Iv

scholarly journals Effects of bfp Mutations on Biogenesis of Functional Enteropathogenic Escherichia coli Type IV Pili

2000 ◽  
Vol 182 (9) ◽  
pp. 2498-2506 ◽  
Author(s):  
Ravi P. Anantha ◽  
Kelly D. Stone ◽  
Michael S. Donnenberg
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Signal Sequence ◽  
Outer Membrane Proteins ◽  
Laboratory Strain ◽  
Type Iv Pili ◽  
Host Cells ◽  
Wild Type ◽  
Type Iv ◽  
The Individual

ABSTRACT Enteropathogenic Escherichia coli expresses a type IV fimbria known as the bundle-forming pilus (BFP) that is required for autoaggregation and localized adherence (LA) to host cells. A cluster of 14 genes is sufficient to reconstitute BFP biogenesis in a laboratory strain of E. coli. We have undertaken a systematic mutagenesis of the individual genes to determine the effect of each mutation on BFP biogenesis and LA. Here we report the construction and analysis of nonpolar mutations in six genes of thebfp cluster, bfpG, bfpB,bfpC, bfpD, bfpP, andbfpH, as well as the further analysis of a previously described bfpA mutant strain that is unable to express bundlin, the pilin protein. We found that mutations inbfpB, which encodes an outer membrane protein;bfpD, which encodes a putative nucleotide-binding protein; and bfpG and bfpC, which do not have sequence homologues in other type IV pilus systems, do not affect prebundlin expression or processing but block both BFP biogenesis and LA. The mutation in bfpP, the prepilin peptidase gene, does not affect prebundlin expression but blocks signal sequence cleavage of prebundlin, BFP biogenesis, and LA. The mutation in bfpH, which is predicted to encode a lytic transglycosylase, has no effect on prebundlin expression, prebundlin processing, BFP biogenesis, or LA. For each mutant for which altered phenotypes were detected, complementation with a plasmid containing the corresponding wild-type allele restored the wild-type phenotypes. We also found that association of prebundlin or bundlin with sucrose density flotation gradient fractions containing both inner and outer membrane proteins does not require any accessory proteins. These studies indicate that many bfp gene products are required for biogenesis of functional type IV pili but that mutations in the individual genes do not lead to the identification of new phases of pilus assembly.

scholarly journals The High-Molecular-Weight Cytochrome c Cyc2 of Acidithiobacillus ferrooxidans Is an Outer Membrane Protein

2002 ◽  
Vol 184 (1) ◽  
pp. 313-317 ◽  
Author(s):  
Andrés Yarzábal ◽  
Gaël Brasseur ◽  
Jeanine Ratouchniak ◽  
Karen Lund ◽  
Danielle Lemesle-Meunier ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Outer Membrane ◽  
High Molecular Weight ◽  
Acidithiobacillus Ferrooxidans ◽  
Ferrous Iron ◽  
Membrane Fraction ◽  
Cytoplasmic Membrane ◽  
Respiratory Pathway ◽  
Membrane Components ◽  
Membrane Level

ABSTRACT A high-molecular-weight c-type cytochrome, Cyc2, and a putative 22-kDa c-type cytochrome were detected in the membrane fraction released during spheroplast formation from Acidithiobacillus ferrooxidans. This fraction was enriched in outer membrane components and devoid of cytoplasmic membrane markers. The genetics, as well as the subcellular localization of Cyc2 at the outer membrane level, therefore make it a prime candidate for the initial electron acceptor in the respiratory pathway between ferrous iron and oxygen.

scholarly journals Ib-M6 Antimicrobial Peptide: Antibacterial Activity against Clinical Isolates of Escherichia coli and Molecular Docking

Antibiotics ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 79 ◽  
Author(s):  
J. M. Flórez-Castillo ◽  
P. Rondón-Villareal ◽  
J. L. Ropero-Vega ◽  
S. Y. Mendoza-Espinel ◽  
J. A. Moreno-Amézquita ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Activity ◽  
Molecular Docking ◽  
Outer Membrane ◽  
Docking Simulations ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Membrane Components ◽  
K 12 ◽  
Susceptibility Assay

The Ib-M6 peptide has antibacterial activity against non-pathogenic Escherichia coli K-12 strain. The first part of this study determines the antibacterial activity of Ib-M6 against fourteen pathogenic strains of E. coli O157:H7. Susceptibility assay showed that Ib-M6 had values of Minimum Inhibitory Concentration (MIC) lower than streptomycin, used as a reference antibiotic. Moreover, to predict the possible interaction between Ib-M6 and outer membrane components of E. coli, we used molecular docking simulations where FhuA protein and its complex with Lipopolysaccharide (LPS–FhuA) were used as targets of the peptide. FhuA/Ib-M6 complexes had energy values between −39.5 and −40.5 Rosetta Energy Units (REU) and only one hydrogen bond. In contrast, complexes between LPS–FhuA and Ib-M6 displayed energy values between −25.6 and −40.6 REU, and the presence of five possible hydrogen bonds. Hence, the antimicrobial activity of Ib-M6 peptide shown in the experimental assays could be caused by its interaction with the outer membrane of E. coli.

scholarly journals Characterization of type IV pili in the life cycle of the predator bacterium Bdellovibrio

Microbiology ◽  
2010 ◽  
Vol 156 (4) ◽  
pp. 1040-1051 ◽  
Author(s):  
Khaled K. Mahmoud ◽  
Susan F. Koval
Keyword(s):  
Life Cycle ◽  
Insoluble Fraction ◽  
Type Iv Pili ◽  
Host Cells ◽  
Prey Cell ◽  
Gram Negative ◽  
Type Iv ◽  
Genes Encoding ◽  
Attack Phase

Bdellovibrio and like organisms (BALOs) are obligate prokaryotic predators of other Gram-negative bacteria. Bdellovibrio bacteriovorus is the most studied organism among BALOs. It has a periplasmic life cycle with two major stages: a motile, non-replicative stage spent searching for prey (the attack phase) and a stage spent inside the periplasm of the Gram-negative prey cell (the growth phase) after forming an osmotically stable body termed the bdelloplast. Within Bdellovibrio, there are also strains exhibiting an epibiotic life cycle. The genome sequence of the type strain B. bacteriovorus HD100T revealed the presence of multiple dispersed pil genes encoding type IV pili. Type IV pili in other bacteria are involved in adherence to and invasion of host cells and therefore can be considered to play a role in invasion of prey cells by Bdellovibrio. In this study, genes involved in producing type IV pili were identified in the periplasmic strain B. bacteriovorus 109J and an epibiotic Bdellovibrio sp. strain JSS. The presence of fibres on attack-phase cells was confirmed by examining negative stains of cells fixed with 10 % buffered formalin. Fibres were at the non-flagellated pole on approximately 25 % of attack-phase cells. To confirm that these fibres were type IV pili, a truncated form of PilA lacking the first 35 amino acids was designed to facilitate purification of the protein. The truncated PilA fused to a His-tag was overexpressed in Escherichia coli BL21(DE3) plysS. The fusion protein, accumulated in the insoluble fraction, was purified under denaturing conditions and used to produce polyclonal antisera. Immunoelectron microscopy showed that polar fibres present on the cell surface of the predator were composed of PilA, the major subunit of type IV pili. Immunofluorescence microscopy showed the presence of pilin on attack-phase cells of B. bacteriovorus 109J during attachment to prey cells and just after penetration, inside the bdelloplast. Antibodies against PilA delayed and inhibited predation in co-cultures of Bdellovibrio. This study confirms that type IV pili play a role in invasion of prey cells by Bdellovibrio.

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