1.65 Å resolution structure of the AraC-family transcriptional activator ToxT from<i>Vibrio cholerae</i>

Li, Jiaqin; Wehmeyer, Graham T; Lovell, Scott; Battaile, Kevin P.; Egan, Susan M.

doi:10.1107/s2053230x1601298x

Cited by 13 publications

(18 citation statements)

References 41 publications

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“…ToxT has previously been shown to undergo proteolysis when V. cholerae is shifted to conditions that do not support virulence (i.e., from 30°C to 37°C, or when the pH increases from pH 6.5 to pH 8), but the specific proteases involved in this process are unknown (55). ToxT appears to be specifically cleaved in a recently structured region of the protein that lies in the N-terminal domain of the protein between amino acids 100 and 109 (56,57). Moreover, it has also been shown that the addition of the UFA linoleic acid to ToxT prevents its proteolysis (56), possibly by promoting the "closed" conformation of ToxT that is unable to bind to DNA or to activate gene expression (18).…”

Section: Discussionmentioning

confidence: 99%

The Fatty Acid Regulator FadR Influences the Expression of the Virulence Cascade in the El Tor Biotype of Vibrio cholerae by Modulating the Levels of ToxT via Two Different Mechanisms

et al. 2017

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FadR is a master regulator of fatty acid (FA) metabolism that coordinates the pathways of FA degradation and biosynthesis in enteric bacteria. We show here that a ΔfadR mutation in the El Tor biotype of Vibrio cholerae prevents the expression of the virulence cascade by influencing both the transcription and the posttranslational regulation of the master virulence regulator ToxT. FadR is a transcriptional regulator that represses the expression of genes involved in FA degradation, activates the expression of genes involved in unsaturated FA (UFA) biosynthesis, and also activates the expression of two operons involved in saturated FA (SFA) biosynthesis. Since FadR does not bind directly to the toxT promoter, we determined whether the regulation of any of its target genes indirectly influenced ToxT. This was accomplished by individually inserting a double point mutation into the FadRbinding site in the promoter of each target gene, thereby preventing their activation or repression. Although preventing FadR-mediated activation of fabA, which encodes the enzyme that carries out the first step in UFA biosynthesis, did not significantly influence either the transcription or the translation of ToxT, it reduced its levels and prevented virulence gene expression. In the mutant strain unable to carry out FadRmediated activation of fabA, expressing fabA ectopically restored the levels of ToxT and virulence gene expression. Taken together, the results presented here indicate that V. cholerae FadR influences the virulence cascade in the El Tor biotype by modulating the levels of ToxT via two different mechanisms.IMPORTANCE Fatty acids (FAs) play important roles in membrane lipid homeostasis and energy metabolism in all organisms. In Vibrio cholerae, the causative agent of the acute intestinal disease cholera, they also influence virulence by binding into an N-terminal pocket of the master virulence regulator, ToxT, and modulating its activity. FadR is a transcription factor that coordinately controls the pathways of FA degradation and biosynthesis in enteric bacteria. This study identifies a new link between FA metabolism and virulence in the El Tor biotype by showing that FadR influences both the transcription and posttranslational regulation of the master virulence regulator ToxT by two distinct mechanisms.

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Section: Discussionmentioning

confidence: 99%

The Fatty Acid Regulator FadR Influences the Expression of the Virulence Cascade in the El Tor Biotype of Vibrio cholerae by Modulating the Levels of ToxT via Two Different Mechanisms

et al. 2017

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“…The RD domain is very similar to that of the AraC but without the N-terminal loop. Instead, Li et al ( 2016 ) revealed a small α-helix (D101 to E110) in proximity of the RD to DBD linker. Mutational analysis in ToxT of Met103, Arg105 or Asn106 showed a threefold activation increase of the ctxA promoter, pointing out the important role of this small α-helix (Childers et al, 2007 ).…”

Section: The One-component System Regulatorsmentioning

confidence: 99%

Functional Mechanism of the Efflux Pumps Transcription Regulators From Pseudomonas aeruginosa Based on 3D Structures

Issa

Phan

Broutin

2018

Front. Mol. Biosci.

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Bacterial antibiotic resistance is a worldwide health problem that deserves important research attention in order to develop new therapeutic strategies. Recently, the World Health Organization (WHO) classified Pseudomonas aeruginosa as one of the priority bacteria for which new antibiotics are urgently needed. In this opportunistic pathogen, antibiotics efflux is one of the most prevalent mechanisms where the drug is efficiently expulsed through the cell-wall. This resistance mechanism is highly correlated to the expression level of efflux pumps of the resistance-nodulation-cell division (RND) family, which is finely tuned by gene regulators. Thus, it is worthwhile considering the efflux pump regulators of P. aeruginosa as promising therapeutical targets alternative. Several families of regulators have been identified, including activators and repressors that control the genetic expression of the pumps in response to an extracellular signal, such as the presence of the antibiotic or other environmental modifications. In this review, based on different crystallographic structures solved from archetypal bacteria, we will first focus on the molecular mechanism of the regulator families involved in the RND efflux pump expression in P. aeruginosa, which are TetR, LysR, MarR, AraC, and the two-components system (TCS). Finally, the regulators of known structure from P. aeruginosa will be presented.

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“…The X-ray structure of ToxT revealed a 16-carbon monounsaturated fatty acid cis -palmitoleate bound in a pocket within the N-terminal domain2026, the homologous location as the arabinose-binding site within AraC27, a related protein for which the family is named. The fatty acid bridges the interface between the N-terminal dimerization domain and C-terminal DNA-binding domain of ToxT, suggesting a potential mechanism for fatty acid-mediated inhibition (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…This results in an “open” state, freeing the C-terminal domain to bind DNA and/or allowing the N-terminal domain to dimerize20. It has been suggested that this rearrangement may involve the flexible N-terminal loop between residues 108–11326. Therefore, ToxT is thought to exist in “closed” (inactive) and “open” (active) states20.…”

Section: Resultsmentioning

confidence: 99%

“…Therefore, in order to inhibit the activity of ToxT, our approach in this work was to design small molecules based on the conformation of the ToxT ligand found in the X-ray crystal structure, cis -palmitoleic acid2026. As predicted, these “pre-folded” small molecules bind much more tightly than UFAs, which show no activity at the tested concentrations.…”

Section: Resultsmentioning

confidence: 99%

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A new class of inhibitors of the AraC family virulence regulator Vibrio cholerae ToxT

Woodbrey

Onyango

Pellegrini

et al. 2017

Sci Rep

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Vibrio cholerae is responsible for the diarrheal disease cholera that infects millions of people worldwide. While vaccines protecting against cholera exist, and oral rehydration therapy is an effective treatment method, the disease will remain a global health threat until long-term solutions such as improved sanitation and access to clean water become widely available. Because of this, there is a pressing need for potent therapeutics that can either mitigate cholera symptoms, or act prophylactically to prevent the virulent effects of a cholera infection. Here we report the design, synthesis, and characterization of a set of compounds that bind and inhibit ToxT, the transcription factor that directly regulates the two primary V. cholerae virulence factors. Using the folded structure of the monounsaturated fatty acid observed in the X-ray structure of ToxT as a template, we designed ten novel compounds that inhibit the virulence cascade to a greater degree than any known inhibitor. Our findings provide a structural and functional basis for the development of viable antivirulence therapeutics that combat cholera and, potentially, other forms of bacterial pathogenic disease.

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1.65 Å resolution structure of the AraC-family transcriptional activator ToxT fromVibrio cholerae

Cited by 13 publications

References 41 publications

The Fatty Acid Regulator FadR Influences the Expression of the Virulence Cascade in the El Tor Biotype of Vibrio cholerae by Modulating the Levels of ToxT via Two Different Mechanisms

The Fatty Acid Regulator FadR Influences the Expression of the Virulence Cascade in the El Tor Biotype of Vibrio cholerae by Modulating the Levels of ToxT via Two Different Mechanisms

Functional Mechanism of the Efflux Pumps Transcription Regulators From Pseudomonas aeruginosa Based on 3D Structures

A new class of inhibitors of the AraC family virulence regulator Vibrio cholerae ToxT

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