Initial Steps of Colicin E1 Import across the Outer Membrane of
            <i>Escherichia coli</i>

Masi, Muriel; Vuong, Phu; Humbard, Matthew A.; Malone, Karen E.; Misra, Rajeev

doi:10.1128/jb.01448-06

Cited by 33 publications

(33 citation statements)

References 54 publications

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“…The finding in the experiments reported here that direct binding of the colicin T domain can occur in vivo without prior interaction with BtuB, as detected by the protection of sensitive bacteria by T domain peptides from killing by colicin E1, contradicts the conclusion of Masi et al (29). They could detect in vivo cross-linking of E1 to TolC by dithiobis(succinimidyl propionate) (DSP) in cells with a functional BtuB colicin receptor but not in btuB mutant cells.…”

Section: Discussioncontrasting

confidence: 56%

“…Binding of some somewhat-shorter E1 T domain peptides was also detected by similar occlusion of TolC channels in planar bilayers and by coelution of those peptides with TolC on a sizing column (28). Mutations of a glycine residue known to face the inside of the TolC channel have been shown to reduce the efficiency of killing by colicin E1, implying that passage of the colicin through the channel was obstructed by the bulkier mutant amino acids (29). However, the nature of the interaction of colicin E1 with TolC and the mechanism of its passage through the periplasm, via translocation using TolC, are presently poorly understood.…”

Section: Importancementioning

confidence: 99%

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The Colicin E1 TolC Box: Identification of a Domain Required for Colicin E1 Cytotoxicity and TolC Binding

Jakes

2017

J Bacteriol

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Colicins are protein toxins made by Escherichia coli to kill related bacteria that compete for scarce resources. All colicins must cross the target cell outer membrane in order to reach their intracellular targets. Normally, the first step in the intoxication process is the tight binding of the colicin to an outer membrane receptor protein via its central receptor-binding domain. It is shown here that for one colicin, E1, that step, although it greatly increases the efficiency of killing, is not absolutely necessary. For colicin E1, the second step, translocation, relies on the outer membrane/transperiplasmic protein TolC. The normal role of TolC in bacteria is as an essential component of a family of tripartite drug and toxin exporters, but for colicin E1, it is essential for its import. Colicin E1 and some N-terminal translocation domain peptides had been shown previously to bind in vitro to TolC and occlude channels made by TolC in planar lipid bilayer membranes. Here, a set of increasingly shorter colicin E1 translocation domain peptides was shown to bind to Escherichia coli in vivo and protect them from subsequent challenge by colicin E1. A segment of only 21 residues, the "TolC box," was thereby defined; that segment is essential for colicin E1 cytotoxicity and for binding of translocation domain peptides to TolC. IMPORTANCEThe Escherichia coli outer membrane/transperiplasmic protein TolC is normally an essential component of the bacterium's tripartite drug and toxin export machinery. The protein toxin colicin E1 instead uses TolC for its import into the cells that it kills, thereby subverting its normal role. Increasingly shorter constructs of the colicin's N-terminal translocation domain were used to define an essential 21-residue segment that is required for both colicin cytotoxicity and for binding of the colicin's translocation domain to bacteria, in order to protect them from subsequent challenge by active colicin E1. Thus, an essential TolC binding sequence of colicin E1 was identified and may ultimately lead to the development of drugs to block the bacterial drug export pathway.KEYWORDS TolC, colicins, drug efflux, membrane translocation, toxins E scherichia coli competes for scarce resources by making plasmid-encoded protein toxins called colicins, which efficiently kill closely related bacteria. All of the few dozen or so colicins that have been identified kill their targets by one of a few basic mechanisms: (i) making an ion-permeable channel in the inner membrane of the target cell, which depolarizes and kills the cell (1); (ii) enzymatically cleaving its rRNA, tRNA, or DNA in the cytoplasm (2, 3); or (iii) degrading peptidoglycan precursors in the periplasm (4, 5). Regardless of their ultimate killing mechanism, all colicins must cross at least the outer membrane in order to reach their targets. The way one particular colicin, E1, makes that transit is the subject of this study.Colicins have a well-defined domain structure, with the killing (catalytic or channelforming) domain at the ...

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Section: Discussioncontrasting

confidence: 56%

Section: Importancementioning

confidence: 99%

The Colicin E1 TolC Box: Identification of a Domain Required for Colicin E1 Cytotoxicity and TolC Binding

Jakes

2017

J Bacteriol

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“…Moreover, in agreement with earlier observations OmpT has been shown to cleave several receptor-bound colicins and thus to improve the survival of target bacteria exposed to colicins or other antimicrobial peptides (23)(24)(25).…”

supporting

confidence: 75%

FtsH-dependent Processing of RNase Colicins D and E3 Means That Only the Cytotoxic Domains Are Imported into the Cytoplasm

Chauleau¹,

Mora²,

Serba

et al. 2011

Journal of Biological Chemistry

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It has long been suggested that the import of nuclease colicins requires protein processing; however it had never been formally demonstrated. Here we show that two RNase colicins, E3 and D, which appropriate two different translocation machineries to cross the outer membrane (BtuB/Tol and FepA/TonB, respectively), undergo a processing step inside the cell that is essential to their killing action. We have detected the presence of the C-terminal catalytic domains of these colicins in the cytoplasm of target bacteria. The same processed forms were identified in both colicin-sensitive cells and in cells immune to colicin because of the expression of the cognate immunity protein. We demonstrate that the inner membrane protease FtsH is necessary for the processing of colicins D and E3 during their import. We also show that the signal peptidase LepB interacts directly with the central domain of colicin D in vitro and that it is a specific but not a catalytic requirement for in vivo processing of colicin D. The interaction of colicin D with LepB may ensure a stable association with the inner membrane that in turn allows the colicin recognition by FtsH. We have also shown that the outer membrane protease OmpT is responsible for alternative and distinct endoproteolytic cleavages of colicins D and E3 in vitro, presumably reflecting its known role in the bacterial defense against antimicrobial peptides. Even though the OmpT-catalyzed in vitro cleavage also liberates the catalytic domain from colicins D and E3, it is not involved in the processing of nuclease colicins during their import into the cytoplasm.Colicins are antibacterial toxins of Escherichia coli that are released into the extracellular medium in response to environmental stress conditions. Colicin D is an RNase that cleaves the anticodon loop of all four isoaccepting tRNA Arg (1). Colicin E3 cleaves 16 S ribosomal RNA (2). Both colicins provoke cell death by inactivating the protein biosynthetic machinery. Colicin producer cells are protected against both endogenous and exogenous toxin molecules by the constitutive expression of a cognate immunity (Imm) 3 protein, which forms a tight heterodimer complex with the nuclease domain of the colicin (3, 4).Colicin E3, like most colicins, has structurally identifiable N-terminal, central, and C-terminal domains. The first two domains are required for translocation and receptor binding, and they "hijack" certain functions of the target cell (i.e. the BtuB receptor and the Tol system) during colicin import. The C-terminal domain carries the cell-killing RNase function (5, 6). The colicin D protein has an unusual tripartite organization. The N-terminal domain is required for both the binding of the colicin to the high affinity, iron siderophore receptor FepA and for its subsequent translocation across the outer membrane. The 280-residue central domain is essential for uptake (and thus for cell killing), and it is also involved in the formation of the colicin D-ImmD protein complex (7). The passage of colicin D through th...

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“…The double-layer sensitivity assay was performed as described previously (Davies & Reeves, 1975;Masi et al, 2007). In brief, colicin-producing strains (Table 1) were grown at 37 uC for 16 h and streaked across 20 ml LB agar plates with cotton swabs.…”

Section: Methodsmentioning

confidence: 99%

Relationship between O-antigen chain length and resistance to colicin E2 in Shigella flexneri

2014

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The Shigella flexneri polysaccharide co-polymerase class 1a (PCP1a) protein, WzzB SF , regulates LPS O-antigen (Oag) chain length to confer short (S)-type Oag chains of~10-17 Oag repeat units (RUs). The S-type Oag chains affect Shigella flexneri virulence as they influence IcsAmediated actin-based motility. However, they do not confer resistance to complement; this is conferred by the very-long (VL)-type Oag chains determined by WzzB pHS2 . Colicins are bacterial proteins produced by some Escherichia coli strains to kill related strains. While the presence of Oag chains has been shown to shield outer-membrane proteins from colicins, the impact of Oag chain length against colicins is unknown. In this study, initial testing indicated that a Shigella flexneri Y wzz : : kan r mutant was more sensitive to colicin E2 compared with the WT strain. Plasmids encoding Wzz mutant and WT PCP1a proteins conferring different Oag modal chain lengths were then expressed in the mutant background, and tested against purified colicin E2. Analysis of swab and spot sensitivity assays showed that strains expressing either S-type or long (L)-type Oag chains (16-28 Oag RUs) conferred greater resistance to colicin E2 compared with strains having very-short-type (2-8 Oag RUs), intermediate-short-type (8-14 Oag RUs) or VLtype (.80 Oag RUs) Oag chains. These results suggest a novel role for LPS Oag chain length control that may have evolved due to selection pressure from colicins in the environment.

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Initial Steps of Colicin E1 Import across the Outer Membrane of Escherichia coli

Cited by 33 publications

References 54 publications

The Colicin E1 TolC Box: Identification of a Domain Required for Colicin E1 Cytotoxicity and TolC Binding

The Colicin E1 TolC Box: Identification of a Domain Required for Colicin E1 Cytotoxicity and TolC Binding

FtsH-dependent Processing of RNase Colicins D and E3 Means That Only the Cytotoxic Domains Are Imported into the Cytoplasm

Relationship between O-antigen chain length and resistance to colicin E2 in Shigella flexneri

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