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J Biol Chem. 2018 Feb 23;293(8):2755-2769. doi: 10.1074/jbc.RA117.000760. Epub 2018 Jan 4.

Cyclipostins and cyclophostin analogs inhibit the antigen 85C from Mycobacterium tuberculosis both in vitro and in vivo.

Author information

1
From the Institut de Recherche en Infectiologie de Montpellier (IRIM), Université de Montpellier, CNRS UMR9004, 34293 Montpellier, France.
2
Aix-Marseille Université, CNRS, EIPL, IMM FR3479, 13009 Marseille, France.
3
Aix-Marseille Université, CNRS, LISM, IMM FR3479, 13009 Marseille, France.
4
Aix Marseille Université, CNRS, INSERM, Institut Paoli-Calmettes, CRCM, Marseille Protéomique, 13009 Marseille, France, and.
5
the Department of Chemistry and Biochemistry, University of Missouri, St. Louis, Missouri 63121.
6
From the Institut de Recherche en Infectiologie de Montpellier (IRIM), Université de Montpellier, CNRS UMR9004, 34293 Montpellier, France, mickael.blaise@irim.cnrs.fr.
7
From the Institut de Recherche en Infectiologie de Montpellier (IRIM), Université de Montpellier, CNRS UMR9004, 34293 Montpellier, France, laurent.kremer@irim.cnrs.fr.
8
INSERM, IRIM, 34293 Montpellier, France.

Abstract

An increasing prevalence of cases of drug-resistant tuberculosis requires the development of more efficacious chemotherapies. We previously reported the discovery of a new class of cyclipostins and cyclophostin (CyC) analogs exhibiting potent activity against Mycobacterium tuberculosis both in vitro and in infected macrophages. Competitive labeling/enrichment assays combined with MS have identified several serine or cysteine enzymes in lipid and cell wall metabolism as putative targets of these CyC compounds. These targets included members of the antigen 85 (Ag85) complex (i.e. Ag85A, Ag85B, and Ag85C), responsible for biosynthesis of trehalose dimycolate and mycolylation of arabinogalactan. Herein, we used biochemical and structural approaches to validate the Ag85 complex as a pharmacological target of the CyC analogs. We found that CyC, CyC, and CyC17 bind covalently to the catalytic Ser124 residue in Ag85C; inhibit mycolyltransferase activity (i.e. the transfer of a fatty acid molecule onto trehalose); and reduce triacylglycerol synthase activity, a property previously attributed to Ag85A. Supporting these results, an X-ray structure of Ag85C in complex with CyC disclosed that this inhibitor occupies Ag85C's substrate-binding pocket. Importantly, metabolic labeling of M. tuberculosis cultures revealed that the CyC compounds impair both trehalose dimycolate synthesis and mycolylation of arabinogalactan. Overall, our study provides compelling evidence that CyC analogs can inhibit the activity of the Ag85 complex in vitro and in mycobacteria, opening the door to a new strategy for inhibiting Ag85. The high-resolution crystal structure obtained will further guide the rational optimization of new CyC scaffolds with greater specificity and potency against M. tuberculosis.

KEYWORDS:

Ag85 complex; Mycobacterium tuberculosis; cell wall; crystal structure; cyclipostins; cyclophostin; inhibition mechanism; trehalose dimycolate; trehalose monomycolate; triacylglycerol

PMID:
29301937
PMCID:
PMC5827452
[Available on 2019-02-23]
DOI:
10.1074/jbc.RA117.000760

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