Research Projects

Tuberculosis (TB), caused by the highly infectious and persistent pathogen Mycobacterium tuberculosis (Mtb), continues to engulf over one-third of the world’s human population, with an estimated 2.61 million active cases in our country in the year 2024. The severity in global incidence, fatality and rapid surge in counts appeals for innovative scientific solutions aligning with national missions to ensure elimination of TB on a priority basis. One key challenge in eradicating this disease lies in the bacterium’s capability to survive for years at the compromise of the host’s clearance, coupled to multiple, and extensive drug-resistance and co-infections, presenting a difficult situation to treat this infection with the current pool of antibiotics. Incidentally, Mtb retains its pathogenic and metabolic activity by acquisition of essential nutrients from the host such as lipids (fatty acids and cholesterol). Recently, it has been found that cholesterol catabolism by non-heme iron co-factors is the most crucial aspect during the chronic phase of Mtb infection that helps to retain its virulence and pathogenesis. Hence, identifying the weaknesses in the bacterium’s catabolism pathways and dysregulating its nutrient utilization opens-up an indirect approach to potential drug discovery. In this project, we therefore intend to have molecular level understanding of iron-dependent two-component Rieske oxygenase KshAB (3-Ketosteroid 9α-hydroxylase) catalyzed cholesterol catabolism, particularly the regioselective mono-hydroxylation of steroid derivative, 1,4-androstadiene-3,17-dione (ADD), in aerobic conditions that holds promise to identify key intermediates in cholesterol catabolism and screening the factors responsible for its catalytic efficiency. We believe, identification of the substrate binding site in KshAB would provide solid foundation towards design of receptor-enzyme based novel inhibitors to effectively treat tuberculosis. We will employ state-of-the-art computational approaches like density functional theory (DFT), quantum mechanics/molecular mechanics (QM/MM), molecular dynamics and machine learning techniques to elucidate the electronic structure of the enzymatic intermediates, identify the factors that control the crucial C-H hydroxylation step by KshAB, and create a database of potential inhibitors based on their reactivity principles. To provide proof-of-principle, we also aim to carry out collaborative experiments and compare relative kinetics of C-H oxidation of the identified novel inhibitor and the ADD substrate with model bio-inspired iron-complexes. We believe our studies would bridge the gap in fundamental understanding of the reactivity of KsAB in cholesterol catabolism and provide leads for biological assays of tuberculosis inhibition. Indirectly, our study would also prove insightful for reactivity, regioselectivity, and potential application of metalloenzymes in organic synthesis of hydroxysteroids.