THPP target assignment reveals EchA6 as an essential fatty acid shuttle in mycobacteria

Abstract

Phenotypic screens for bactericidal compounds against drug-resistant tuberculosis are beginning to yield novel inhibitors. However, reliable target identification remains challenging. Here, we show that tetrahydropyrazo[1,5-a]pyrimidine-3-carboxamide (THPP) selectively pulls down EchA6 in a stereospecific manner, instead of the previously assigned target Mycobacterium tuberculosis MmpL3. While homologous to mammalian enoyl-coenzyme A (CoA) hydratases, EchA6 is non-catalytic yet essential and binds long-chain acyl-CoAs. THPP inhibitors compete with CoA-binding, suppress mycolic acid synthesis, and are bactericidal in a mouse model of chronic tuberculosis infection. A point mutation, W133A, abrogated THPP-binding and increased both the in vitro minimum inhibitory concentration and the in vivo effective dose 99 in mice. Surprisingly, EchA6 interacts with selected enzymes of fatty acid synthase II (FAS-II) in bacterial two-hybrid assays, suggesting essentiality may be linked to feeding long-chain fatty acids to FAS-II. Finally, our data show that spontaneous resistance-conferring mutations can potentially obscure the actual target or alternative targets of small molecule inhibitors.

Publication DOI: https://doi.org/10.1038/nmicrobiol.2015.6
Divisions: Life & Health Sciences
Life & Health Sciences > Applied Health Research Group
Life & Health Sciences > Cell & Tissue Biomedical Research
PURE Output Type: Article
Published Date: 2016-01-18
Accepted Date: 2015-10-02
Submitted Date: 2015-08-03
Authors: Cox, Jonathan A.G. (ORCID Profile 0000-0001-5208-4056)
Abrahams, Katherine A.
Alemparte, Carlos
Ghidelli-Disse, Sonja
Rullas, Joaquín
Angulo-Barturen, Iñigo
Singh, Albel
Gurcha, Sudagar S.
Nataraj, Vijayashankar
Bethell, Stephen
Remuiñán, Modesto J.
Encinas, Lourdes
Jervis, Peter J.
Cammack, Nicholas C.
Bhatt, Apoorva
Kruse, Ulrich
Bantscheff, Marcus
Fütterer, Klaus
Barros, David
Ballell, Lluis
Drewes, Gerard
Besra, Gurdyal S.

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