Mycobacterium tuberculosis is one of the most important opportunistic pathogens of HIV-infected individuals. Current treatments for tuberculosis are being threatened by the rapid emergence of drug resistance. Our research program has worked to define the mechanisms of action of a leading anti-tuberculosis drug isoniazid (INH) and an important second line drug ethionamide (ETH), with the goal of enabling rational drug design. Using a combination of genetics, biochemistry, X-ray crystallography, electron microscopic and gene analysis approaches, we discovered a common target to be an enoyl reductase of the Fatty Acid Synthase (FAS) type II system responsible for mycolic acid synthesis. The three dimensional structure of InhA was determined, and its enzymatic activity was unexpectedly found be in inhibited by an INH-NAD adduct that bound to the InhA NADH binding pocket. We have also identified a set of contiguous genes iniB, iniA, and iniC, that confer tolerance to INH in mycobacteria overexpressing these genes. We have established InhA as an excellent drug target by demonstrating that InhA-thermal inactivation of a temperature-sensitive mutation in inhA leads to lysis of the mycobacterial cell. Moreover, we have identified novel compounds that inhibit InhA and possess anti-mycobacterial activity. Despite these accomplishments, additional questions remain concerning the molecular events that lead to INH-induced cell lysis, and the mechanisms that confer resistance to this phenomenon. Here we propose to continue our multi-disciplinary approach to study novel resistance mechanisms, including one that is caused by defects in NADH dehydrogenase. We will also focus on defining the molecular events that lead to mycobacterial cell lysis by comparing and contrasting three different mutant strains that cause lysis following inhibition of three different metabolic pathways. Finally, we will characterize the tolerance mediated by the IniA and IniB proteins, and define the mechanisms by which mycobacteria prevent lysis. The knowledge gained by this work will lead to the identification of novel drug targets, strategies to overcome tolerance, and more effective treatments for tuberculosis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI043268-07
Application #
6895541
Study Section
AIDS-associated Opportunistic Infections and Cancer Study Section (AOIC)
Program Officer
Goldman, Robert C
Project Start
1998-07-01
Project End
2009-04-30
Budget Start
2005-05-01
Budget End
2006-04-30
Support Year
7
Fiscal Year
2005
Total Cost
$617,440
Indirect Cost
Name
Albert Einstein College of Medicine
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
110521739
City
Bronx
State
NY
Country
United States
Zip Code
10461
Harbut, Michael B; Yang, Baiyuan; Liu, Renhe et al. (2018) Small Molecules Targeting Mycobacterium tuberculosis Type II NADH Dehydrogenase Exhibit Antimycobacterial Activity. Angew Chem Int Ed Engl 57:3478-3482
Vilcheze, Catherine; Baughn, Anthony D; Tufariello, JoAnn et al. (2011) Novel inhibitors of InhA efficiently kill Mycobacterium tuberculosis under aerobic and anaerobic conditions. Antimicrob Agents Chemother 55:3889-98
Vilcheze, Catherine; Weinrick, Brian; Wong, Ka-Wing et al. (2010) NAD+ auxotrophy is bacteriocidal for the tubercle bacilli. Mol Microbiol 76:365-77
Colangeli, R; Haq, A; Arcus, V L et al. (2009) The multifunctional histone-like protein Lsr2 protects mycobacteria against reactive oxygen intermediates. Proc Natl Acad Sci U S A 106:4414-8
Sridharan, Sudharsan; Wang, Lei; Brown, Alistair K et al. (2007) X-ray crystal structure of Mycobacterium tuberculosis beta-ketoacyl acyl carrier protein synthase II (mtKasB). J Mol Biol 366:469-80