Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a diverse group of... more Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a diverse group of biologically active bacterial molecules. Due to the conserved genomic arrangement of many of the genes involved in their synthesis, these secondary metabolite biosynthetic pathways can be predicted from genome sequence data. To date, however, despite the myriad of sequenced genomes covering many branches of the bacterial phylogenetic tree, such an analysis for a broader group of bacteria like anaerobes has not been attempted. We investigated a collection of 211 complete and published genomes, focusing on anaerobic bacteria, whose potential to encode RiPPs is relatively unknown. We showed that the presence of RiPP-genes is widespread among anaerobic representatives of the phyla Actinobacteria, Proteobacteria and Firmicutes and that, collectively, anaerobes possess the ability to synthesize a broad variety of different RiPP classes. More than 25% of anaerobes are capable of producing RiPP...
Angewandte Chemie (International ed. in English), Jan 21, 2014
Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial produce... more Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial producer of solvents, revealed the presence of several cryptic gene clusters for secondary metabolite biosynthesis. To unearth its metabolic potential, a C. beijerinckii strain was cultured under various conditions, which led to the discovery of a deep purple pigment. This novel metabolite, named clostrubin (1), was isolated and its structure was fully elucidated. The pentacyclic polyphenol features a benzo[a]tetraphene ring topology that is unprecedented for natural products. Stable-isotope labeling experiments showed that 1 is an aromatic polyketide that folds in a noncanonical manner to form the unusual perifused ring system. In addition to being the first reported polyketide from an anaerobic bacterium, 1 is a potent antibiotic with pronounced activity against various pathogenic bacteria, such as MRSA, VRE, and mycobacteria, with minimum inhibitory concentrations (MIC) of 0.12-0.97 μM.
A total of 211 complete and published genomes from anaerobic bacteria are analysed for the presen... more A total of 211 complete and published genomes from anaerobic bacteria are analysed for the presence of secondary metabolite biosynthesis gene clusters, in particular those tentatively coding for polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS). We investigate the distribution of these gene clusters according to bacterial phylogeny and, if known, correlate these to the type of metabolic pathways they encode. The potential of anaerobes as secondary metabolite producers is highlighted.
Around 25% of vegetable food is lost worldwide because of infectious plant diseases, including mi... more Around 25% of vegetable food is lost worldwide because of infectious plant diseases, including microbe-induced decay of harvested crops. In wet seasons and under humid storage conditions, potato tubers are readily infected and decomposed by anaerobic bacteria (Clostridium puniceum). We found that these anaerobic plant pathogens harbor a gene locus (type II polyketide synthase) to produce unusual polyketide metabolites (clostrubins) with dual functions. The clostrubins, which act as antibiotics against other microbial plant pathogens, enable the anaerobic bacteria to survive an oxygen-rich plant environment.
The Clostridium perfringens tetracycline resistance protein, TetA(P), is an inner-membrane protei... more The Clostridium perfringens tetracycline resistance protein, TetA(P), is an inner-membrane protein that mediates the active efflux of tetracycline from the bacterial cell. This protein comprises 420 aa and is predicted to have 12 transmembrane domains (TMDs). Comparison of the TetA(P) amino acid sequence to that of several members of the major facilitator superfamily (MFS) identified a variant copy of the conserved Motif A. This region consists of the sequence E59xPxxxxxDxxxRK72 and is located within the putative loop joining TMDs 2 and 3 in the predicted structural model of the TetA(P) protein. To study the functional importance of the conserved residues, site-directed mutagenesis was used to construct 17 point mutations that were then analysed for their effect on tetracycline resistance and their ability to produce an immunoreactive TetA(P) protein. Changes to the conserved Phe-58 residue were tolerated, whereas three independent substitutions of Pro-61 abolished tetracycline resi...
... A Blueprint for the Synthesis of Mycolactones Sacha J. Pidot , Nicholas J. Tobias , and Tim S... more ... A Blueprint for the Synthesis of Mycolactones Sacha J. Pidot , Nicholas J. Tobias , and Tim Stinear(*ü) ... 2004) . Other studies have identified mycobacterial isolates from diseased fish from both Chesapeake bay in the United States and the Mediterranean Sea (Rhodes et al. ...
Your browser version may not work well with NCBI's web applications. More information here..... more Your browser version may not work well with NCBI's web applications. More information here... ...
Mycolactone A/B is a lipophilic macrocyclic polyketide that is the primary virulence factor produ... more Mycolactone A/B is a lipophilic macrocyclic polyketide that is the primary virulence factor produced by Mycobacterium ulcerans, a human pathogen and the causative agent of Buruli ulcer. In M. ulcerans strain Agy99 the mycolactone polyketide synthase (PKS) locus spans a 120 kb region of a 174 kb megaplasmid. Here we have identified promoter regions of this PKS locus using GFP reporter assays, in silico analysis, primer extension, and site-directed mutagenesis. Transcription of the large PKS genes mlsA1 (51 kb), mlsA2 (7 kb) and mlsB (42 kb) is driven by a novel and powerful SigA-like promoter sequence situated 533 bp upstream of both the mlsA1 and mlsB initiation codons, which is also functional in Escherichia coli, Mycobacterium smegmatis and Mycobacterium marinum. Promoter regions were also identified upstream of the putative mycolactone accessory genes mup045 and mup053. We transformed M. ulcerans with a GFP-reporter plasmid under the control of the mls promoter to produce a highl...
Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a diverse group of... more Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a diverse group of biologically active bacterial molecules. Due to the conserved genomic arrangement of many of the genes involved in their synthesis, these secondary metabolite biosynthetic pathways can be predicted from genome sequence data. To date, however, despite the myriad of sequenced genomes covering many branches of the bacterial phylogenetic tree, such an analysis for a broader group of bacteria like anaerobes has not been attempted. We investigated a collection of 211 complete and published genomes, focusing on anaerobic bacteria, whose potential to encode RiPPs is relatively unknown. We showed that the presence of RiPP-genes is widespread among anaerobic representatives of the phyla Actinobacteria, Proteobacteria and Firmicutes and that, collectively, anaerobes possess the ability to synthesize a broad variety of different RiPP classes. More than 25% of anaerobes are capable of producing RiPP...
Angewandte Chemie (International ed. in English), Jan 21, 2014
Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial produce... more Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial producer of solvents, revealed the presence of several cryptic gene clusters for secondary metabolite biosynthesis. To unearth its metabolic potential, a C. beijerinckii strain was cultured under various conditions, which led to the discovery of a deep purple pigment. This novel metabolite, named clostrubin (1), was isolated and its structure was fully elucidated. The pentacyclic polyphenol features a benzo[a]tetraphene ring topology that is unprecedented for natural products. Stable-isotope labeling experiments showed that 1 is an aromatic polyketide that folds in a noncanonical manner to form the unusual perifused ring system. In addition to being the first reported polyketide from an anaerobic bacterium, 1 is a potent antibiotic with pronounced activity against various pathogenic bacteria, such as MRSA, VRE, and mycobacteria, with minimum inhibitory concentrations (MIC) of 0.12-0.97 μM.
A total of 211 complete and published genomes from anaerobic bacteria are analysed for the presen... more A total of 211 complete and published genomes from anaerobic bacteria are analysed for the presence of secondary metabolite biosynthesis gene clusters, in particular those tentatively coding for polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS). We investigate the distribution of these gene clusters according to bacterial phylogeny and, if known, correlate these to the type of metabolic pathways they encode. The potential of anaerobes as secondary metabolite producers is highlighted.
Around 25% of vegetable food is lost worldwide because of infectious plant diseases, including mi... more Around 25% of vegetable food is lost worldwide because of infectious plant diseases, including microbe-induced decay of harvested crops. In wet seasons and under humid storage conditions, potato tubers are readily infected and decomposed by anaerobic bacteria (Clostridium puniceum). We found that these anaerobic plant pathogens harbor a gene locus (type II polyketide synthase) to produce unusual polyketide metabolites (clostrubins) with dual functions. The clostrubins, which act as antibiotics against other microbial plant pathogens, enable the anaerobic bacteria to survive an oxygen-rich plant environment.
The Clostridium perfringens tetracycline resistance protein, TetA(P), is an inner-membrane protei... more The Clostridium perfringens tetracycline resistance protein, TetA(P), is an inner-membrane protein that mediates the active efflux of tetracycline from the bacterial cell. This protein comprises 420 aa and is predicted to have 12 transmembrane domains (TMDs). Comparison of the TetA(P) amino acid sequence to that of several members of the major facilitator superfamily (MFS) identified a variant copy of the conserved Motif A. This region consists of the sequence E59xPxxxxxDxxxRK72 and is located within the putative loop joining TMDs 2 and 3 in the predicted structural model of the TetA(P) protein. To study the functional importance of the conserved residues, site-directed mutagenesis was used to construct 17 point mutations that were then analysed for their effect on tetracycline resistance and their ability to produce an immunoreactive TetA(P) protein. Changes to the conserved Phe-58 residue were tolerated, whereas three independent substitutions of Pro-61 abolished tetracycline resi...
... A Blueprint for the Synthesis of Mycolactones Sacha J. Pidot , Nicholas J. Tobias , and Tim S... more ... A Blueprint for the Synthesis of Mycolactones Sacha J. Pidot , Nicholas J. Tobias , and Tim Stinear(*ü) ... 2004) . Other studies have identified mycobacterial isolates from diseased fish from both Chesapeake bay in the United States and the Mediterranean Sea (Rhodes et al. ...
Your browser version may not work well with NCBI's web applications. More information here..... more Your browser version may not work well with NCBI's web applications. More information here... ...
Mycolactone A/B is a lipophilic macrocyclic polyketide that is the primary virulence factor produ... more Mycolactone A/B is a lipophilic macrocyclic polyketide that is the primary virulence factor produced by Mycobacterium ulcerans, a human pathogen and the causative agent of Buruli ulcer. In M. ulcerans strain Agy99 the mycolactone polyketide synthase (PKS) locus spans a 120 kb region of a 174 kb megaplasmid. Here we have identified promoter regions of this PKS locus using GFP reporter assays, in silico analysis, primer extension, and site-directed mutagenesis. Transcription of the large PKS genes mlsA1 (51 kb), mlsA2 (7 kb) and mlsB (42 kb) is driven by a novel and powerful SigA-like promoter sequence situated 533 bp upstream of both the mlsA1 and mlsB initiation codons, which is also functional in Escherichia coli, Mycobacterium smegmatis and Mycobacterium marinum. Promoter regions were also identified upstream of the putative mycolactone accessory genes mup045 and mup053. We transformed M. ulcerans with a GFP-reporter plasmid under the control of the mls promoter to produce a highl...
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