Target-based screen against a periplasmic serine protease that regulates intrabacterial pH homeostasis in Mycobacterium tuberculosis.

TitleTarget-based screen against a periplasmic serine protease that regulates intrabacterial pH homeostasis in Mycobacterium tuberculosis.
Publication TypeJournal Article
Year of Publication2015
AuthorsZhao N, Darby CM, Small J, Bachovchin DA, Jiang X, Burns-Huang KE, Botella H, Ehrt S, Boger DL, Anderson ED, Cravatt BF, Speers AE, Fernandez-Vega V, Hodder PS, Eberhart C, Rosen H, Spicer TP, Nathan CF
JournalACS Chem Biol
Volume10
Issue2
Pagination364-71
Date Published2015 Feb 20
ISSN1554-8937
KeywordsBacterial Proteins, Benzoxazines, Gene Expression Regulation, Bacterial, Gene Expression Regulation, Enzymologic, Homeostasis, Hydrogen-Ion Concentration, Molecular Probes, Molecular Structure, Mycobacterium tuberculosis, Periplasm, Serine Proteases, Serine Proteinase Inhibitors
Abstract

Mycobacterium tuberculosis (Mtb) maintains its intrabacterial pH (pHIB) near neutrality in the acidic environment of phagosomes within activated macrophages. A previously reported genetic screen revealed that Mtb loses this ability when the mycobacterial acid resistance protease (marP) gene is disrupted. In the present study, a high throughput screen (HTS) of compounds against the protease domain of MarP identified benzoxazinones as inhibitors of MarP. A potent benzoxazinone, BO43 (6-chloro-2-(2'-methylphenyl)-4H-1,3-benzoxazin-4-one), acylated MarP and lowered Mtb's pHIB and survival during incubation at pH 4.5. BO43 had similar effects on MarP-deficient Mtb, suggesting the existence of additional target(s). Reaction of an alkynyl-benzoxazinone, BO43T, with Mycobacterium bovis variant bacille Calmette-Guérin (BCG) followed by click chemistry with azido-biotin identified both the MarP homologue and the high temperature requirement A1 (HtrA1) homologue, an essential protein. Thus, the chemical probe identified through a target-based screen not only reacted with its intended target in the intact cells but also implicated an additional enzyme that had eluded a genetic screen biased against essential genes.

DOI10.1021/cb500746z
Alternate JournalACS Chem Biol
PubMed ID25457457
PubMed Central IDPMC4340348
Grant ListR01 AI081725 / AI / NIAID NIH HHS / United States
U54 MH084512 / MH / NIMH NIH HHS / United States
MH084512 / MH / NIMH NIH HHS / United States
T32GM07739 / GM / NIGMS NIH HHS / United States
T32 GM007739 / GM / NIGMS NIH HHS / United States

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