The major Vibrio cholerae autoinducer and its role in virulence factor production

Department of Chemistry, Princeton University, Princeton, New Jersey, United States
Nature (Impact Factor: 42.35). 01/2008; 450(7171):883-6. DOI: 10.1038/nature06284
Source: PubMed

ABSTRACT Vibrio cholerae, the causative agent of the human disease cholera, uses cell-to-cell communication to control pathogenicity and biofilm formation. This process, known as quorum sensing, relies on the secretion and detection of signalling molecules called autoinducers. At low cell density V. cholerae activates the expression of virulence factors and forms biofilms. At high cell density the accumulation of two quorum-sensing autoinducers represses these traits. These two autoinducers, cholerae autoinducer-1 (CAI-1) and autoinducer-2 (AI-2), function synergistically to control gene regulation, although CAI-1 is the stronger of the two signals. V. cholerae AI-2 is the furanosyl borate diester (2S,4S)-2-methyl-2,3,3,4-tetrahydroxytetrahydrofuran borate. Here we describe the purification of CAI-1 and identify the molecule as (S)-3-hydroxytridecan-4-one, a new type of bacterial autoinducer. We provide a synthetic route to both the R and S isomers of CAI-1 as well as simple homologues, and we evaluate their relative activities. Synthetic (S)-3-hydroxytridecan-4-one functions as effectively as natural CAI-1 in repressing production of the canonical virulence factor TCP (toxin co-regulated pilus). These findings suggest that CAI-1 could be used as a therapy to prevent cholera infection and, furthermore, that strategies to manipulate bacterial quorum sensing hold promise in the clinical arena.

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    ABSTRACT: Abstract Cholix toxin from V. cholerae, is the third member of the diphtheria toxin group of mono-ADP-ribosyltransferase bacterial toxins. It shares structural and functional properties with P. aeruginosa exotoxin A and C. diphtheriae diphtheria toxin. Cholix toxin is an important model for the development of antivirulence approaches and therapeutics against these toxins from pathogenic bacteria. Herein, we have used the high-resolution X-ray structure of full-length cholix complexed with NAD(+) to describe the properties of the NAD(+)-binding pocket at the residue level, including the role of crystallographic water molecules in the NAD(+) substrate interaction. The full length apo cholix structure is used to describe the putative NAD(+) binding site(s) and to correlate biochemical with crystallographic data to study the stoichiometry and orientation of bound NAD(+) molecules. We quantitatively describe the NAD(+) substrate interactions on a residue basis for the main 22 pocket residues in cholixf, a glycerol and 5 contact water molecules as part of the recognition surface by the substrate according to the conditions of crystallization. In addition, the dynamic properties of an in silico version of the catalytic domain were investigated in order to understand the lack of electronic density for one of the main flexible loops (R-loop) in the pocket of X-ray complexes. Implications for a rational drug design approach for mART toxins are derived.
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    ABSTRACT: The purpose of the present study was to analyze the composition of marine bacterial communities around the world, and to investigate bacterial isolates regarding the production of antibiotics. This included molecular analyses of marine bacterioplankton, as well as culture-based studies of marine bacterial isolates with antagonistic activity. The work was based on samples collected during the Galathea 3 and LOMROG-II marine research expeditions that have explored many different oceanic regions worldwide. A molecular survey of marine bacterioplankton at 24 worldwide stations investigated the abundance of major bacterial groups, potential biogeographical patterns, and their relation to environmental parameters. The original aim was to determine whether the composition of the total microbiota correlates with the occurrence of culturable bioactive bacteria. No such correlation was found. Quantitative community analyses showed latitudinal patterns in bacterial distribution, revealing significantly different relative abundances of Bacteroidetes, unclassified Bacteria and Vibrio between warmer and colder oceans. Absolute cell numbers of most bacterial groups were positively correlated with nutrient concentrations in warmer oceans, and negatively with oxygen saturation in colder oceans. The finding of differing communities in warmer and colder oceans underlined the presence of biogeographical patterns among marine bacteria and the influence of environmental parameters on bacterial distribution. Studies of antagonistic isolates focused on six bioactive Vibrionaceae isolated during Galathea 3. The six strains were identified as Vibrio coralliilyticus (two strains), V. neptunius (two strains), V. nigripulchritudo (one strain), and Photobacterium halotolerans (one strain) by sequencing of housekeeping genes. Chemical metabolite profiling underlined genetic relationships by showing highly similar production of secondary metabolites for each species. Two known antibiotics were purified; andrimid from V. coralliilyticus and holomycin from P. halotolerans. In addition, two novel cyclic peptides from P. halotolerans and a novel siderophore-like compound from V. nigripulchritudo were isolated. All three compounds interfere with quorum sensing in S. aureus. During LOMROG-II further seventeen strains with antagonistic activity were isolated, affiliating with the Actinobacteria (8 strains), Pseudoalteromonas (4 strains), the Vibrionaceae (3 strains), and Psychrobacter (2 strains). Seven of the eight bioactive Actinobacteria, being isolated from different sources throughout the Arctic Ocean, were related to Arthrobacter davidanieli. Its broad antibiotic spectrum was likely based on production of the known arthrobacilin antibiotics. The eighth actinomycete, tentatively identified as Brevibacterium sp., produces a potentially novel antimicrobial compound. Most studies of antagonistic marine bacteria have been conducted with the aim of isolating novel antimicrobials with potential clinical applications. However, little is known about production and role of these compounds in the natural environment. This thesis took one step in this direction and demonstrated that V. coralliilyticus S2052 produced its antibiotic andrimid when grown with chitin as the sole carbon source. Whilst the strain produced an array of secondary metabolites in laboratory media, it focused on andrimid production with chitin. This indicates that the antibiotic is likely produced in the natural habitat and may serve an ecophysiological function. The finding that two related strains from public culture collections do not produce andrimid and have different biosynthetic temperature optima suggested that V. coralliilyticus may comprise different subspecies with different niches. In summary, the present study shows biogeographical patterns of marine bacterioplankton on a global scale. In addition, the thesis work has demonstrated that marine Vibrionaceae and polar Actinobacteria are a resource of antibacterial compounds and may have potential for future natural product discovery.
    04/2011, Degree: PhD, Supervisor: Lone Gram