ИСТИНА

While in most bacteria penicillin-binding enzymes D,D-transpeptidases play the major role in the cell wall peptidoglycan cross-linking catalyzing formation of the so-called 4-3 bonds between 4th D-Ala and 3rd meso-diaminopimelic acid (m-Dap) residues of different chains, in Mycobacterium tuberculosis the most of cross-links are formed by L,D-transpeptidases (LdtMt), that catalyze formation of 3-3 cross-links between two 3rd m-Dap residues. L,D-transpeptidases, in contrast to D,D-transpeptidases, are not sensitive to widely used penicillins and cephalosporins [1,2], and are considered as attractive molecular targets searching for novel antitubercular agents. In this work interaction of L,D-transpeptidase 2 from M. tuberculosis with natural peptidoglycan fragment that consists of both glycan (N-acetyl-D-glucosamine [NAG] and N-acetyl muramic acid [NAM]) and tetrapeptide (L-Ala-γ-D-Glu-m-Dap-D-Ala) parts and contain either C-terminal carboxyl groups of γ-D-Glu(COO-) and m-Dap(COO-) residues or their amides, was studied using advanced molecular modeling. The force field parameters were obtained for natural peptidoglycan fragment NAG-NAM-[L-Ala-γ-D-Glu-m-Dap-D-Ala]-NAG that include γ-D-Glu and m-Dap residues in both carboxyl and amide forms: γ-D-Glu(COO-/CONH2) and m-Dap(COO-/CONH2). Energetics of the active site lid opening process that control substrate delivery to the active site was evaluated using the well-tempered metadynamics. The system’s collective variables which describe formation of the enzyme-substrate complex were selected and free energy surface (FES) for the substrate binding in the active site was built. Energetically more favorable formation of enzyme-substrate complex in case of amide substrate was observed compared to the corresponding substrate bearing free carboxyl groups. The results of molecular modeling are supported by the experimental data. This work was supported by the Russian Science Foundation (grant 15-14-00069-P) References: 1. Böth D., Steiner E.M., Stadler D., Lindqvist Y., Schnell R., Schneider G. // Acta Crystallogr. Sect. D Biol. Crystallogr. 2013. V. 69. № 3. P. 432–441. 2. Erdemli S.B., Gupta R., Bishai W.R., Lamichhane G., Amzel L.M., Bianchet M.A. // Structure. 2012. V. 20. № 12. P. 2103–2115.