Biotin biosynthesis is essential for survival and persistence of Mycobacterium tuberculosis (Mtb) in vivo. The aminotransferase BioA, which catalyzes the antepenultimate step in the biotin pathway, has been established as a promising target due to its vulnerability to chemical inhibition. We performed high-throughput screening (HTS) employing a fluorescence displacement assay and identified a diverse set of potent inhibitors including many diversity-oriented synthesis (DOS) scaffolds. To efficiently select only hits targeting biotin biosynthesis, we then deployed a whole-cell counterscreen in biotin-free and biotin-containing medium against wild-type Mtb and in parallel with isogenic bioA Mtb strains that possess differential levels of BioA expression. This counterscreen proved crucial to filter out compounds whose whole-cell activity was off target as well as identify hits with weak, but measurable whole-cell activity in BioA-depleted strains. Several of the most promising hits were cocrystallized with BioA to provide a framework for future structure-based drug design efforts.
Bibliographical noteFunding Information:
This work was funded in part by grants from the NIH (R03 MH096537 to C.C.A. and R01AI091790 to D.S.) and by the NIH-MLPCN program (1 U54 HG005032-1 awarded to S.L.S.). We also gratefully acknowledge support from the University of Minnesota Bighley Graduate Fellowship (to R.D.) and resources from the University of Minnesota Supercomputing Institute.
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