Screening For Potential Inhibitors Of Mycobacterium Tuberculosis Isocitrate Lyase: In Silico And In Vitro Approaches
Loading...
Date
2017-11
Authors
Lee, Yie Vern
Journal Title
Journal ISSN
Volume Title
Publisher
Universiti Sains Malaysia
Abstract
Tuberculosis (TB) that is caused by Mycobacterium tuberculosis (MTB) still remains as global epidemic and affects one-third of the world population, regardless of active TB or latent TB infection. The metabolism for active and latent MTB are different, for instance they have different energy generation cycle during active and latent stage. Instead of focusing on the drug targets during the active stage, which already have the effective therapeutic drugs, this study targeted isocitrate lyase (ICL), a potential target for dormant (latent) MTB energy generation cycle (glyoxylate cycle). Studies showed that without ICL, dormant MTB cannot survives within the lung of a murine model. Besides, this enzyme is not found in mammals hence it is an appropriate drug target to eliminate TB. In this study, molecular dynamics (MD) simulations were first performed on the crystal structure of ICL, using AMBER 8 molecular dynamics package. Results from 30 ns of MD simulations showed that C-terminal loop is involved in the access towards the active site. In addition, MM-PBSA calculation showed that the substrate (isocitrate) binding was contributed by electrostatic interaction as well as the hydrophobic interactions. Ensemble conformations obtained from MD simulation were then used for virtual screening to search for the potential ICL inhibitor from Malaysian local plant database (NADI) with the collection of 3,000 compounds. Virtual screening that performed with AutoDock 3.05 has managed to identify 22 potential candidates which are within Lipinski's Rule of Five (with drug-likeliness criteria). Of the 22 candidates, 6 were commercially available as pure compound while the remaining 16 candidates can only be obtained from 12 plants (however, only 10 plants were available during the study). Therefore, 6 pure compounds and 10 crude extracts were used to test for their inhibition activity by enzymatic assay and disc diffusion assay. In enzymatic inhibitory assay, MTB ICL was cloned, expressed and purified whereas disc diffusion assay used a replacing MTB model, M. smegmatis to test against the potential inhibitors suggested from virtual screening.
Description
Keywords
Mycobacterium tuberculosis isocitrate lyase , In silico and in vitro approaches