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ACS Chem Biol. 2017 Feb 17;12(2):422-434. doi: 10.1021/acschembio.6b01037. Epub 2016 Dec 23.

Rapid Chagas Disease Drug Target Discovery Using Directed Evolution in Drug-Sensitive Yeast.

Author information

1
Department of Pediatrics, University of California, San Diego, School of Medicine , La Jolla, California 92093, United States.
2
Department of Synthetic Biology and Bioenergy, J. Craig Venter Institute , La Jolla, California 92037, United States.
3
Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego , La Jolla, California 92093, United States.
4
Cellular Ultrastructure Laboratory, IOC, FIOCRUZ , Rio de Janeiro, Rio de Janeiro, Brazil 21045-360.
5
Department of Pharmacy, Federal University of Paraná , Curitiba, Paraná, Brazil 80210-170.
6
Department of Chemistry & Biochemistry, University of California, San Diego , La Jolla, California 92093-0340, United States.
7
Department of Biological Sciences, University of Pittsburgh , Pittsburgh, Pennsylvania 15260, United States.

Abstract

Recent advances in cell-based, high-throughput phenotypic screening have identified new chemical compounds that are active against eukaryotic pathogens. A challenge to their future development lies in identifying these compounds' molecular targets and binding modes. In particular, subsequent structure-based chemical optimization and target-based screening require a detailed understanding of the binding event. Here, we use directed evolution and whole-genome sequencing of a drug-sensitive S. cerevisiae strain to identify the yeast ortholog of TcCyp51, lanosterol-14-alpha-demethylase (TcCyp51), as the target of MMV001239, a benzamide compound with activity against Trypanosoma cruzi, the etiological agent of Chagas disease. We show that parasites treated with MMV0001239 phenocopy parasites treated with another TcCyp51 inhibitor, posaconazole, accumulating both lanosterol and eburicol. Direct drug-protein binding of MMV0001239 was confirmed through spectrophotometric binding assays and X-ray crystallography, revealing a binding site shared with other antitrypanosomal compounds that target Cyp51. These studies provide a new probe chemotype for TcCyp51 inhibition.

PMID:
27977118
PMCID:
PMC5649375
DOI:
10.1021/acschembio.6b01037
[Indexed for MEDLINE]
Free PMC Article

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