Targeting DnaN for tuberculosis therapy using novel griselimycins

Kling, Angela; Lukat, Peer; Almeida, Deepak; Bauer, Armin; Fontaine, Evelyne; Sordello, Sylvie; Zaburannyi, Nestor; Herrmann, Jennifer; Wenzel, Silke C.; König, Claudia; Ammerman, Nicole C.; Barrio, María Belén Pascual; Borchers, Kai; Bordon-Pallier, Florence; Brönstrup, Mark; Courtemanche, Gilles; Gerlitz, Martin; Geslin, Michel; Hammann, Peter; Heinz, Dirk W.; Hoffmann, Holger; Klieber, Sylvie; Kohlmann, Markus; Kurz, Michael; Lair, Christine; Matter, Hans; Nuermberger, Eric L.; Tyagi, Sandeep; Fraisse, Laurent; Grosset, J; Lagrange, Sophie; Müller, Rolf

doi:10.1126/science.aaa4690

Cited by 274 publications

(294 citation statements)

References 66 publications

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“…It might be worth noting that most nonribosomal antimicrobial peptides, such as polymyxin, fusaricidin, and daptomycin, are "lipopeptides" which contain a mixture of charged residues and aliphatic tails and act by disrupting cell membrane or cell wall function (52,53). In contrast, our peptides contain only aliphatic groups and are reminiscent of natural products that inhibit intracellular targets, such as griselimycin, which targets DnaN of Mycobacterium tuberculosis (54). Whether the peptomers can cross the bacterial membrane remains to be tested.…”

Section: Discussionmentioning

confidence: 99%

Synthetic Cyclic Peptomers as Type III Secretion System Inhibitors

Lam

Schwochert

Lao

et al. 2017

Antimicrob Agents Chemother

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Antibiotic-resistant bacteria are an emerging threat to global public health. New classes of antibiotics and tools for antimicrobial discovery are urgently needed. Type III secretion systems (T3SS), which are required by dozens of Gramnegative bacteria for virulence but largely absent from nonpathogenic bacteria, are promising virulence blocker targets. The ability of mammalian cells to recognize the presence of a functional T3SS and trigger NF-B activation provides a rapid and sensitive method for identifying chemical inhibitors of T3SS activity. In this study, we generated a HEK293 stable cell line expressing green fluorescent protein (GFP) driven by a promoter containing NF-B enhancer elements to serve as a readout of T3SS function. We identified a family of synthetic cyclic peptide-peptoid hybrid molecules (peptomers) that exhibited dose-dependent inhibition of T3SS effector secretion in Yersinia pseudotuberculosis and Pseudomonas aeruginosa without affecting bacterial growth or motility. Among these inhibitors, EpD-3=N, EpD-1,2N, EpD-1,3=N, EpD-1,2,3=N, and EpD-1,2,4=N exhibited strong inhibitory effects on translocation of the Yersinia YopM effector protein into mammalian cells (Ͼ40% translocation inhibition at 7.5 M) and showed no toxicity to mammalian cells at 240 M. In addition, EpD-3=N and EpD-1,2,4=N reduced the rounding of HeLa cells caused by the activity of Yersinia effector proteins that target the actin cytoskeleton. In summary, we have discovered a family of novel cyclic peptomers that inhibit the injectisome T3SS but not the flagellar T3SS.

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Section: Discussionmentioning

confidence: 99%

Synthetic Cyclic Peptomers as Type III Secretion System Inhibitors

Lam

Schwochert

Lao

et al. 2017

Antimicrob Agents Chemother

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“…20 Of these mechanisms, target modification uniquely correlates an antibiotic to its mode of action. The antibiotics novobiocin (gyrase B), 21 platensin (FabB/F), 22 and griselimycin (DnaN), 23 for instance, represent a few examples in which target-duplicated resistance genes are co-clustered with BGCs (Supporting Information Figure 1). With the notion in mind that antibiotic-producing bacteria often duplicate and mutate genes encoding targeted proteins to confer resistance, we reasoned that identifying putative resistance genes within BGCs would provide insight to the molecular targets of BGC chemical products prior to their isolation and structure elucidation.…”

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confidence: 99%

Identification of Thiotetronic Acid Antibiotic Biosynthetic Pathways by Target-directed Genome Mining

et al. 2015

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Recent genome sequencing efforts have led to the rapid accumulation of uncharacterized or “orphaned” secondary metabolic biosynthesis gene clusters (BGCs) in public databases. This increase in DNA-sequenced big data has given rise to significant challenges in the applied field of natural product genome mining, including (i) how to prioritize the characterization of orphan BGCs, and (ii) how to rapidly connect genes to biosynthesized small molecules. Here we show that by correlating putative antibiotic resistance genes that encode target-modified proteins with orphan BGCs, we predict the biological function of pathway specific small molecules before they have been revealed in a process we call target-directed genome mining. By querying the pan-genome of 86 Salinispora bacterial genomes for duplicated house-keeping genes co-localized with natural product BGCs, we prioritized an orphan polyketide synthase-nonribosomal peptide synthetase hybrid BGC (tlm) with a putative fatty acid synthase resistance gene. We employed a new synthetic double-stranded DNA-mediated cloning strategy based on transformation-associated recombination to efficiently capture tlm and the related ttm BGCs directly from genomic DNA and to heterologously express them in Streptomyces hosts. We show the production of a group of unusual thiotetronic acid natural products, including the well-known fatty acid synthase inhibitor thiolactomycin that was first described over 30 years ago, yet never at the genetic level in regards to biosynthesis and auto-resistance. This finding not only validates the target-directed genome mining strategy for the discovery of antibiotic producing gene clusters without a priori knowledge of the molecule synthesized, but also paves the way for the investigation of novel enzymology involved in thiotetronic acid natural product biosynthesis.

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“…5 Critically, these drugs, drug candidates, and screening hits have also been used to identify a number of new TB drug targets. These include the AtpE subunit of ATP synthase (the target of bedaquiline 6 ); the decaprenylphosphoryl-β- d -ribose 2-epimerase, DprE1 (the target of PBTZ169 7 and other pharmacophores 8 ); the trehalose monomycolate transporter, MmpL3 (the target of indolcarboxamides 9 and multiple other pharmacophores 10,11 ); QcrB, a component of the cytochrome bc 1 – aa 3 complex (the target of Q203 12 and other pharmacophores 13 ); DnaN, the target of griselimycin; 14 and FadD32, the target of diarylcoumarins. 15 By virtue of their novel mechanisms of action, drugs that are active against these targets have the potential to offer new therapeutic options for the treatment of DS- as well as DR-TB.…”

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confidence: 99%

The Inosine Monophosphate Dehydrogenase, GuaB2, Is a Vulnerable New Bactericidal Drug Target for Tuberculosis

et al. 2016

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VCC234718, a molecule with growth inhibitory activity against Mycobacterium tuberculosis (Mtb), was identified by phenotypic screening of a 15344-compound library. Sequencing of a VCC234718-resistant mutant identified a Y487C substitution in the inosine monophosphate dehydrogenase, GuaB2, which was subsequently validated to be the primary molecular target of VCC234718 in Mtb. VCC234718 inhibits Mtb GuaB2 with a Ki of 100 nM and is uncompetitive with respect to IMP and NAD+. This compound binds at the NAD+ site, after IMP has bound, and makes direct interactions with IMP; therefore, the inhibitor is by definition uncompetitive. VCC234718 forms strong pi interactions with the Y487 residue side chain from the adjacent protomer in the tetramer, explaining the resistance-conferring mutation. In addition to sensitizing Mtb to VCC234718, depletion of GuaB2 was bactericidal in Mtb in vitro and in macrophages. When supplied at a high concentration (≥125 μM), guanine alleviated the toxicity of VCC234718 treatment or GuaB2 depletion via purine salvage. However, transcriptional silencing of guaB2 prevented Mtb from establishing an infection in mice, confirming that Mtb has limited access to guanine in this animal model. Together, these data provide compelling validation of GuaB2 as a new tuberculosis drug target.

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Targeting DnaN for tuberculosis therapy using novel griselimycins

Cited by 274 publications

References 66 publications

Synthetic Cyclic Peptomers as Type III Secretion System Inhibitors

Synthetic Cyclic Peptomers as Type III Secretion System Inhibitors

Identification of Thiotetronic Acid Antibiotic Biosynthetic Pathways by Target-directed Genome Mining

The Inosine Monophosphate Dehydrogenase, GuaB2, Is a Vulnerable New Bactericidal Drug Target for Tuberculosis

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