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Information × Registration Number 0221U106929, 0120U104795 , R & D reports Title Site-directed mutagenesis as a novel strategy to investigate and overcome Mycobacterium tuberculosis antibiotic resistance popup.stage_title Head Yarmoliuk Serhii M., Доктор хімічних наук Registration Date 22-12-2021 Organization Institute of Molecular Biology and Genetics of NAS of Ukraine popup.description2  For the first time we have calculated the binding free energy (ΔG) of wild type and mutant forms of M. tuberculosis MetRS such as I10A, Y12A, E24A, D49A, K54A, D263A, H290A, L293A, F292A with the ligand – methionyl-adenylate using umbrella sampling algorithm. According to the results of metadynamics we have selected two amino acid residues – Glu24 and Leu293, for experimental site-directed mutagenesis. Recombinant M. tuberculosis LeuRS and MetRS were obtained in sufficient amount. We have built artificial neural network (ANN) models for the search of potential M. tuberculosis MetRS and LeuRS inhibitors. The ROC score for LeuRS ANN models was in the range 0.91-0.95. The ROC score for MetRS ANN models was in the range 0,91 – 0,97. Among the 100 compounds selected by machine lerning approach, as a result of biochemical screening, we have identified 17 novel MetRS inhibitors and 15 novel LeuRS inhibitors which inhibit enzymes activity by more than 50% at the concentration of 100 µM. The optimization of inhibitors with dual-targeted activity toward M. tuberculosis MetRS and LeuRS among 2-(quinolin-2-ylsulfanyl)-acetamide derivatives was performed. Among the 10 synthesized analogs of this chemical class, 8 MetRS inhibitors which inhibit the enzyme in the range from 29% to 88% at the concentrantion 50 µM and 10 LeuRS inhibitors which inhibit the enzyme in the range from 26% to 94% at the concentrantion 50 µM were identified. It was found that the compound 2-[2-(6-chloro-4-methyl-quinolin-2-ylsulfanyl)-acetylamino]-5-phenyl-thiophene-3-carboxylic acid amide exhibits inhibitory activity simultaneously toward mycobacterial MetRS and LeuRS with IC50 values of 33±16 µM and 23,9±2,5 µM, respectively. Product Description popup.authors popup.nrat_date 2021-12-22 Close