A natural prodrug activation mechanism in nonribosomal peptide synthesis

Reimer, Daniela; Pos, Klaas M.; Thines, Marco; Grün, Peter; Bode, Helge B.

doi:10.1038/nchembio.688

Cited by 123 publications

(156 citation statements)

References 22 publications

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“…Xenocoumacin 1 is bactericidal against the producer Xenorhabdus nematophila, and X. nematophila has evolved an elegant mechanism to avoid this toxicity; an inactive prodrug, prexenocoumacin, is first biosynthesised by an NRPS, after which the active form is derived by cleavage and transport through the membrane by a dual-function membrane bound protein (Reimer et al, 2011). A further mechanism of resistance to the exported xenocoumacin 1 also exists, where X. nematophila cells that take up the compound can detoxify it through an intracellular cyclisation reaction (Reimer et al, 2011).…”

Section: Insect-associated Bacteria With Antibiotic Potentialmentioning

confidence: 99%

Antibiotics from neglected bacterial sources

Pidot¹,

Coyne²,

Kloß³

et al. 2014

International Journal of Medical Microbiology

119

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Section: Insect-associated Bacteria With Antibiotic Potentialmentioning

confidence: 99%

Antibiotics from neglected bacterial sources

Pidot¹,

Coyne²,

Kloß³

et al. 2014

International Journal of Medical Microbiology

119

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“…The m/z ratios observed in the different extracts are clearly in the range one would expect for the products of the identified gene clusters. Especially interesting is the hybrid PKS-NRPS gene cluster 4, encoding a compound that is most likely activated via proteolytic cleavage, as was shown for the potent antibiotic xenocoumacin from the entomopathogenic bacterium Xenorhabdus nematophila (39) and the still-unknown compound colibactin, a potent virulence factor of pathogenic Escherichia coli strains (51).…”

Section: Discussionmentioning

confidence: 99%

“…Similar NRPS have been shown to be involved in the biosynthesis of piperazines or terphenylquinones from fungi (48,49). The largest gene cluster (cluster 4) encodes an NRPS-PKS hybrid with all structural features of the prodrug activation mechanism previously identified in the biosynthesis of xenocoumacin, zwittermicin, and colibactin (39,(50)(51)(52). Additionally, a putatively incomplete NRPS-PKS hybrid gene cluster (cluster 5 in Fig.…”

Section: Artificial Rearingmentioning

confidence: 99%

“…For this, DSM 16116 was grown with or without the adsorber resin Amberlite XAD16 that binds typical low-molecular-weight natural products like peptides or polyketides (39,46,47), and the cell-free culture supernatant was consecutively used to prepare the larva diet. Adding XAD16-extracted culture supernatant or nonextracted supernatant to the diet reduced the survival time of larvae compared to that of the control larvae.…”

Section: Artificial Rearingmentioning

confidence: 99%

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Low-Molecular-Weight Metabolites Secreted by Paenibacillus larvae as Potential Virulence Factors of American Foulbrood

Schild

Fuchs

Bode

et al. 2014

Appl Environ Microbiol

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The spore-forming bacterium Paenibacillus larvae causes a severe and highly infective bee disease, American foulbrood (AFB). Despite the large economic losses induced by AFB, the virulence factors produced by P. larvae are as yet unknown. To identify such virulence factors, we experimentally infected young, susceptible larvae of the honeybee, Apis mellifera carnica, with different P. larvae isolates. Honeybee larvae were reared in vitro in 24-well plates in the laboratory after isolation from the brood comb. We identified genotype-specific differences in the etiopathology of AFB between the tested isolates of P. larvae, which were revealed by differences in the median lethal times. Furthermore, we confirmed that extracts of P. larvae cultures contain low-molecular-weight compounds, which are toxic to honeybee larvae. Our data indicate that P. larvae secretes metabolites into the medium with a potent honeybee toxic activity pointing to a novel pathogenic factor(s) of P. larvae. Genome mining of P. larvae subsp. larvae BRL-230010 led to the identification of several biosynthesis gene clusters putatively involved in natural product biosynthesis, highlighting the potential of P. larvae to produce such compounds.

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“…However, HPLC/MS-based FA analysis is diffi cult, mainly because of little ionizability and hence little signal strength. Small modifi cations introduced to certain FAs, however, can greatly increase ionizability water gradient ranging from 5 to 95% in 22 min at a fl ow rate of 0.6 ml/min ( 12 ). High resolution mass spectra were obtained from a MALDI LTQ Orbitrap XL (Thermo Fisher Scientifi c, Inc., Waltham, MA) equipped with a laser at 337 nm.…”

mentioning

confidence: 99%

ω-Azido fatty acids as probes to detect fatty acid biosynthesis, degradation, and modification

Pérez

Bode

2014

Journal of Lipid Research

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while decreasing lipophilicity, which in turn increases signal resolution on reversed phase HPLC systems ( 7 ).The method we developed for HPLC/MS-based detection of FA intermediates involves three steps : i ) simple preparation of FAs with a terminal azido group (AFAs) that allows most FA modifi cations to occur; ii ) use of these AFAs as metabolic probes and labeling of the in vivo formed derivatives in the corresponding organism with tetramethoxydibenzoazacyclooctyne (TDAC) ( 1 , Fig. 1A ), a cyclooctyne synthesized originally by Starke, Walther, and Pietzsch ( 8 ,9 ); and iii ) the detection of the clicked compounds by HPLC/MS. Although usually a reporter function such as a fl uorophore is linked to the alkyne reagent, no such modifi cations were needed here, because the formation of an electron-rich triazole ring make it quite susceptible to protonation and thus detection by MS, especially as the molecular mass range of the clicked FAs differs strongly from other lipophilic compounds found in most cells. AFAs are bioorthogonal, do not react with other functional groups other than alkynes, are easily taken up by the cells like other FAs when externally added, and can be used to follow the fate of AFAs in a given organism in real time. Thus AFAs will add to the overall toolbox of lipid analysis in general. Similarily, alkyne-modifi ed cholesterol has been used recently to trace cellular cholesterol metabolism and localization ( 10 ). MATERIALS AND METHODS General experimental proceduresSolvents and reagents were obtained from Sigma-Aldrich (München, Germany). TDAC was synthesized using the procedure described by Starke, Walther, and Pietzsch ( 9 ) FAs are found in all known living organisms, playing a vital role in cell compartmentalization, energy storage, and secondary metabolite production. In bacteria, most FAs are found in the cell membrane as part of the lipid bilayer ( 1 ). The actual FA profi le of a cell can strongly vary depending on environmental and developmental conditions requiring de novo biosynthesis, degradation, and modifi cation of the FAs involved ( 2-5 ). The monitoring of these metabolic pathways is usually conducted by GC-MS of FA methyl esters or other volatile FA derivatives ( 6 ). In spite of its merits, such as high sensibility and direct observability of especially volatile natural products, it is desirable to also have simple and effective methods of FA analysis via HPLC/MS, which today is a widespread tool in analytical chemistry as well. However, HPLC/MS-based FA analysis is diffi cult, mainly because of little ionizability and hence little signal strength. Small modifi cations introduced to certain FAs, however, can greatly increase ionizability

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A natural prodrug activation mechanism in nonribosomal peptide synthesis

Cited by 123 publications

References 22 publications

Antibiotics from neglected bacterial sources

Antibiotics from neglected bacterial sources

Low-Molecular-Weight Metabolites Secreted by Paenibacillus larvae as Potential Virulence Factors of American Foulbrood

ω-Azido fatty acids as probes to detect fatty acid biosynthesis, degradation, and modification

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