SUMMARY Tuberculosis (TB) is the second leading cause of human mortality from infectious diseases worldwide. The WHO reported 1.3 million deaths and 8.6 million new cases of TB in 2012. Mycobacterium tuberculosis (M. tuberculosis), the infectious bacteria that causes TB, is encapsulated by a thick and robust cell wall. The innermost segment of the cell wall is comprised of peptidoglycan, a layer that is required for survival and growth of the pathogen. Enzymes that catalyse biosynthesis of the peptidoglycan are essential and are therefore attractive targets for discovery of novel antibiotics as humans lack similar enzymes making it possible to selectively target bacteria only. In this paper, we have reviewed the structures and functions of enzymes GlmS, GlmM, GlmU, MurA, MurB, MurC, MurD, MurE and MurF from M. tuberculosis that are involved in peptidoglycan biosynthesis. In addition, we report homology modelled 3D structures of those key enzymes from M. tuberculosis of which the structures are still unknown. We demonstrated that natural substrates can be successfully docked into the active sites of the GlmS and GlmU respectively. It is therefore expected that the models and the data provided herein will facilitate translational research to develop new drugs to treat TB.

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NRH:quinone reductase (QR2) is present in the retinas of embryonic and post-hatched (PH) chicks. 5-Methoxycarbonylamino-N-acetyltryptamine (5-MCA-NAT) is a QR2 ligand that increases cAMP levels in developing retinas, but it does not affect cAMP levels in CHO-QR2 cells. The dopamine quinone reductase activity of QR2 retrieves dopamine, which increases cAMP levels in developing retinas. The objective of the present study was to investigate whether 5-MCA-NAT increases endogenous dopamine levels in retinas from chick embryos and post-hatched chicks. Endogenous dopamine was measured by enzyme-linked immunosorbent assay (ELISA). 5-MCA-NAT increased retinal endogenous dopamine levels at all developmental stages studied and in PH chicks (-logEC50=11.62±0.34 M). This effect was inhibited by non-selective antagonists of receptors and melatonin binding sites N-acetyl-2-benzyltryptamine (luzindole, 5 μM), but it was not inhibited by the Mel1b melatonin receptor antagonist 4-phenyl-2-propionamidotetralin (4-P-PDOT, 10 nM). The QR2 cosubstrate, N-methyl-dihydronicotinamide (NMH) (-logEC50=6.74±0.26 M), increased endogenous dopamine levels in controls and in retinas stimulated with 5-MCA-NAT (3 nM). The QR2 inhibitor benzo[e]pyrene inhibited endogenous dopamine levels in both control (-logIC50=7.4±0.28 M) and NMH-stimulated (at 100 nM and 1 μM benzo[e]pyrene concentrations) retinas. Theoretical studies using Molegro Virtual Docking software corroborated these experimental results. We conclude that 5-MCA-NAT increases the level of endogenous dopamine via QR2. We suggest that this enzyme triggers double reduction of the dopamine quinone, recovering dopamine in retinal development.

The GlcNAcstatin is a potent inhibitor of O-glycoprotein 2-acetamino-2-deoxy-β-D-glucopyranosidase, which has been related with type II diabetes and neurodegenerative disorders. Herein, hybrid quantum mechanics/molecular mechanics, molecular dynamics simulations, and potential of mean force were employed to study the interactions established between GlcNAcstatin and a bacterial O-GlcNAcase enzyme from Clostridium perfringens. The results reveal that the imidazole nitrogen atom of GlcNAcstatin has shown a better interaction with the active site of Clostridium perfringens in its protonated form, which is compatible with a substrate-assisted reaction mechanism involving two conserved aspartate residues (297 and 298). Furthermore, the quantum mechanics/molecular mechanics-molecular dynamics simulations appointed a strong interaction between Asp401, Asp298, and Asp297 residues and the GlcNAcstatin inhibitor, which is in accordance with experimental data. Lastly, these results may contribute to understand the molecular mechanism of inhibition of Clostridium perfringens by GlcNAcstatin inhibitor and, consequently, this study might be useful to design new molecules with more interesting inhibitory activity.

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