Structural Basis for the Recognition of Mycolic Acid Precursors by KasA, a Condensing Enzyme and Drug Target from Mycobacterium Tuberculosis

Schiebel, J.; Kapilashrami, Kanishk; Fekete, Ágnes; Bommineni, Gopal R.; Schaefer, C.M.; Mueller, Martin J.; Tonge, Peter J.; Kisker, Caroline

doi:10.1074/jbc.m113.511436

Cited by 36 publications

(50 citation statements)

References 65 publications

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“…4b ). This sandwich of PEG and GSK3011724A enables the large acyl cavity to be filled, occupying the volume of the branched phospholipid chains previously observed within this pocket 25 .…”

Section: Resultsmentioning

confidence: 94%

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Identification of KasA as the cellular target of an anti-tubercular scaffold

et al. 2016

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Phenotypic screens for bactericidal compounds are starting to yield promising hits against tuberculosis. In this regard, whole-genome sequencing of spontaneous resistant mutants generated against an indazole sulfonamide (GSK3011724A) identifies several specific single-nucleotide polymorphisms in the essential Mycobacterium tuberculosis β-ketoacyl synthase (kas) A gene. Here, this genomic-based target assignment is confirmed by biochemical assays, chemical proteomics and structural resolution of a KasA-GSK3011724A complex by X-ray crystallography. Finally, M. tuberculosis GSK3011724A-resistant mutants increase the in vitro minimum inhibitory concentration and the in vivo 99% effective dose in mice, establishing in vitro and in vivo target engagement. Surprisingly, the lack of target engagement of the related β-ketoacyl synthases (FabH and KasB) suggests a different mode of inhibition when compared with other Kas inhibitors of fatty acid biosynthesis in bacteria. These results clearly identify KasA as the biological target of GSK3011724A and validate this enzyme for further drug discovery efforts against tuberculosis.

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“…4b ). This sandwich of PEG and GSK3011724A enables the large acyl cavity to be filled, occupying the volume of the branched phospholipid chains previously observed within this pocket 25 .…”

Section: Resultsmentioning

confidence: 94%

“…Curious to understand the molecular details of GSK3011724A binding to KasA, a 2.13 Å co-crystal structure of the dimeric KasA-GSK3011724A complex was solved ( Table 5 ). All literature inhibitors to date, such as TLM, reside in the malonyl substrate pocket close to the catalytic Cys171 residue 25 26 ( Fig. 4a ).…”

Section: Resultsmentioning

confidence: 99%

Identification of KasA as the cellular target of an anti-tubercular scaffold

et al. 2016

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“…While these analogs showed subtle changes in binding affinity to mtFabB, the length of the alkyl substituent modulated the drug:target residence time (t R = 1/k off ) with optimal t R observed for TLM 3 binding to the C171Q mtFabB acyl-enzyme mimic. 33, 52 While the MIC values of these compounds against M. tuberculosis were either similar to the MIC for TLM (3 μg/mL) or larger, the ethyl analog TLM 3 showed activity against a TLM resistant strain of H37Rv, 29 with an MIC value of 2.5 μg/mL. In addition, these analogs did not show improvement in whole cell activity against Y. pestis , B. pseudomallei and S. aureus (Table 3 and Table S2).…”

Section: Resultsmentioning

confidence: 99%

“…32 Subsequently, interligand NOE NMR studies suggested that substituents at the C4 and the C3 positions of TLM (Figure 1) would strengthen interactions with the target enzyme, and subsequently it was demonstrated that modifications to these positions on the thiolactone core resulted in improved binding to the M. tuberculosis FabB (mtFabB; KasA) in vitro . 33, 52, 53 Here, we describe the synthesis, in vitro inhibition and in vivo efficacy of enantiopure ( 5R )-TLM analogs. These compounds were evaluated for activity against mtFabB, the S. aureus FabF enzyme (saFabF) and clinically relevant bacteria such as Mycobacterium tuberculosis , methicillin sensitive S. aureus (MSSA), MRSA, Francisella tularensis , Klebsiella pneumoniae , Burkholderia pseudomallei and Yersinia pestis .…”

Section: Introductionmentioning

confidence: 99%

Thiolactomycin-Based Inhibitors of Bacterial β-Ketoacyl-ACP Synthases with in Vivo Activity

Bommineni¹,

Kapilashrami²,

Cummings

et al. 2016

J. Med. Chem.

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β-Ketoacyl-ACP synthases (KAS) are key enzymes involved in the type II bacterial fatty acid biosynthesis (FASII) pathway and are putative targets for antibacterial discovery. Several natural product KAS inhibitors have previously been reported, including thiolactomycin (TLM), which is produced by Nocardia spp. Here we describe the synthesis and characterization of optically pure 5R-thiolactomycin (TLM) analogs that show improved whole cell activity against bacterial strains including methicillin resistant Staphylococcus aureus (MRSA) and priority pathogens such as Francisella tularensis and Burkholderia pseudomallei. In addition, we identify TLM analogs with in vivo efficacy against MRSA and Klebsiella pneumoniae in animal models of infection.

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“…M. smegmatis, a fast-growing saprophytic relative of Mtb, is a potential expression host for Mtb genes overcoming many of the restrictions of E. coli (36)(37)(38)(39). Unexpectedly, however, only nine proteins of H37Rv represented by 27 structures were produced in M. smegmatis (37,(40)(41)(42)(43)(44)(45)(46). Clearly the faster growth and production rates of E. coli ensure that it remains the most popular expression host.…”

Section: Technical Aspects Of Mtb Structural Analysismentioning

confidence: 99%

Assessing the progress of Mycobacterium tuberculosis H37Rv structural genomics

et al. 2015

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Tuberculosis threatens human health nowhere more than in developing countries with large malnourished and/or immune-compromised (e.g. HIV infected) populations. The etiological agent, Mycobacterium tuberculosis (Mtb), is highly infectious and current interventions demonstrate limited ability to control the epidemic in particular of drug resistant Mtb strains. New drugs and vaccines are thus urgently required. Structural biologists are critical to the TB research community. By identifying potential drug targets and solving their three dimensional structures they open new avenues of identifying potential inhibitors complementing the screening of novel compounds and the investigation of Mtb's molecular physiology by pharmaceutical companies and academic researchers. Much effort has gone into structurally elucidating the Mtb proteome though much remains to be done with progress primarily limited by technological constraints. We review the currently available data for Mtb H37Rv to extract the lessons they have taught us. KeywordsMycobacterium tuberculosis, structural genomics, drug target AbbreviationsAll abbreviations in the manuscript are standard in the field.

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Structural Basis for the Recognition of Mycolic Acid Precursors by KasA, a Condensing Enzyme and Drug Target from Mycobacterium Tuberculosis

Cited by 36 publications

References 65 publications

Identification of KasA as the cellular target of an anti-tubercular scaffold

Identification of KasA as the cellular target of an anti-tubercular scaffold

Thiolactomycin-Based Inhibitors of Bacterial β-Ketoacyl-ACP Synthases with in Vivo Activity

Assessing the progress of Mycobacterium tuberculosis H37Rv structural genomics

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