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Depsipeptide Intermediates Interrogate Proposed Biosynthesis of Cereulide, the Emetic Toxin of Bacillus cereus.

ABSTRACT: Cereulide and isocereulides A-G are biosynthesized as emetic toxins by Bacillus cereus via a non-ribosomal peptide synthetase (NRPS) called Ces. Although a thiotemplate mechanisms involving cyclo-trimerization of ready-made D-O-Leu-D-Ala-L-O-Val-L-Val via a thioesterase (TE) domain is proposed for cereulide biosynthesis, the exact mechanism is far from being understood. UPLC-TOF MS analysis of B. cereus strains in combination with (13)C-labeling experiments now revealed tetra-, octa-, and dodecapeptides of a different sequence, namely (L-O-Val-L-Val-D-O-Leu-D-Ala)1-3, as intermediates of cereulide biosynthesis. Surprisingly, also di-, hexa-, and decadepsipeptides were identified which, together with the structures of the previously reported isocereulides E, F, and G, do not correlate to the currently proposed mechanism for cereulide biosynthesis and violate the canonical NRPS biosynthetic logic. UPLC-TOF MS metabolite analysis and bioinformatic gene cluster analysis highlighted dipeptides rather than single amino or hydroxy acids as the basic modules in tetradepsipeptide assembly and proposed the CesA C-terminal C* domain and the CesB C-terminal TE domain to function as a cooperative esterification and depsipeptide elongation center repeatedly recruiting the action of the C* domain to oligomerize tetradepsipeptides prior to the release of cereulide from the TE domain by macrocyclization.

SUBMITTER: Marxen S

PROVIDER: S-EPMC4445039 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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Depsipeptide Intermediates Interrogate Proposed Biosynthesis of Cereulide, the Emetic Toxin of Bacillus cereus.

Marxen Sandra S Stark Timo D TD Rütschle Andrea A Lücking Genia G Frenzel Elrike E Scherer Siegfried S Ehling-Schulz Monika M Hofmann Thomas T

Scientific reports 20150527

Cereulide and isocereulides A-G are biosynthesized as emetic toxins by Bacillus cereus via a non-ribosomal peptide synthetase (NRPS) called Ces. Although a thiotemplate mechanisms involving cyclo-trimerization of ready-made D-O-Leu-D-Ala-L-O-Val-L-Val via a thioesterase (TE) domain is proposed for cereulide biosynthesis, the exact mechanism is far from being understood. UPLC-TOF MS analysis of B. cereus strains in combination with (13)C-labeling experiments now revealed tetra-, octa-, and dodeca ...[more]

PMID: 26013201

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Project description:BackgroundCereulide, a depsipeptide structurally related to valinomycin, is responsible for the emetic type of gastrointestinal disease caused by Bacillus cereus. Recently, it has been shown that this toxin is produced by a nonribosomal peptide synthetase (NRPS), but its exact genetic organization and biochemical synthesis is unknown.ResultsThe complete sequence of the cereulide synthetase (ces) gene cluster, which encodes the enzymatic machinery required for the biosynthesis of cereulide, was dissected. The 24 kb ces gene cluster comprises 7 CDSs and includes, besides the typical NRPS genes like a phosphopantetheinyl transferase and two CDSs encoding enzyme modules for the activation and incorporation of monomers in the growing peptide chain, a CDS encoding a putative hydrolase in the upstream region and an ABC transporter in the downstream part. The enzyme modules responsible for incorporation of the hydroxyl acids showed an unusual structure while the modules responsible for the activation of the amino acids Ala and Val showed the typical domain organization of NRPS. The ces gene locus is flanked by genetic regions with high homology to virulence plasmids of B. cereus, Bacillus thuringiensis and Bacillus anthracis. PFGE and Southern hybridization showed that the ces genes are restricted to emetic B. cereus and indeed located on a 208 kb megaplasmid, which has high similarities to pXO1-like plasmids.ConclusionThe ces gene cluster that is located on a pXO1-like virulence plasmid represents, beside the insecticidal and the anthrax toxins, a third type of B. cereus group toxins encoded on megaplasmids. The ces genes are restricted to emetic toxin producers, but pXO1-like plasmids are also present in emetic-like strains. These data might indicate the presence of an ancient plasmid in B. cereus which has acquired different virulence genes over time. Due to the unusual structure of the hydroxyl acid incorporating enzyme modules of Ces, substantial biochemical efforts will be required to dissect the complete biochemical pathway of cereulide synthesis.

Dataset Information

Depsipeptide Intermediates Interrogate Proposed Biosynthesis of Cereulide, the Emetic Toxin of Bacillus cereus.

Publications

Depsipeptide Intermediates Interrogate Proposed Biosynthesis of Cereulide, the Emetic Toxin of Bacillus cereus.

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