Biophysical Society Thematic Meeting - November 16-20, 2015

Biophysics in the Understanding, Diagnosis, and Treatment of Infectious Diseases Speaker Abstracts

Identifying Vulnerable Steps in the CoA Biosynthesis Pathway of M. Tuberculosis Joanna Evans 1 , Hyungjin Eoh 2 , Carolina Trujillo 2 , Sabine Ehrt 2 , Dirk Schnappinger 2 , Helena Boshoff 3 , Clifton Barry III 3 , Kyu Rhee 2 , Valerie Mizrahi 1 . 1 University of Cape Town, Observatory, Cape Town, South Africa, 2 Weill Cornell Medical College, New York, NY, USA, 3 NIAID, Bethesda, MD, USA. Enzymes in the pantothenate and coenzyme A (CoA) biosynthesis pathways have attracted considerable interest as potential targets for the development of drugs against a number of human pathogens, including M. tuberculosis . However, although potent inhibitors have been developed against pantothenate synthase (PanC), pantothenate kinase (PanK), these have failed to translate into compounds with significant whole-cell activity. In addition to issues of permeability, metabolism and efflux, such target-led approaches to TB drug discovery are confounded by a lack of understanding of target vulnerability. In this talk, I will describe the combined genetic, physiologic and metabolomic approach we have taken to identify vulnerable steps in the pantothenate and CoA biosynthesis pathway in M. tuberculosis . The impact of target depletion on the viability of M. tuberculosis has been assessed using a set of conditional mutants in various steps in the pathway. While transcriptional silencing of panB , panC or coaE was bacteriostatic, coaBC silencing was apparently bactericidal in M. tuberculosis in vitro, as deduced by CFU enumeration. CoaBC was similarly shown to be required for growth and persistence of M. tuberculosis in mice, based on quantification of organ bacillary loads. However, taking advantage of the fact that M. tuberculosis is capable of CoaBC bypass through CoA salvage, we showed that coaBC silencing results in a ‘non-growing but metabolically active’ (NGMA) state from which non-culturable bacilli can be partially and transiently rescued by CoA salvage. The response of M. tuberculosis to CoA depletion is being further explored by metabolomic analyses which have elucidated similarities and differences in the way in which the organism adapts metabolically to depletion of different targets in the biosynthetic pathway. These findings have significant implications for TB drug discovery, which will be discussed.

22