The flavin-dependent sugar oxidoreductase pyranose dehydrogenase (PDH) from the plant litter-degrading fungus Agaricus meleagris oxidizes D-glucose (GLC) efficiently at positions C2 and C3. The closely related pyranose 2-oxidase (P2O) from Trametes multicolor oxidizes GLC only at position C2. Consequently, the electron output per molecule GLC is twofold for PDH compared to P2O making it a promising catalyst for bioelectrochemistry or for introducing novel carbonyl functionalities into sugars. The aim of this study was to rationalize the mechanism of GLC dioxidation by employing molecular dynamics simulations of GLC–PDH interactions. Shape complementarity through nonpolar van der Waals interactions was identified as the main driving force for GLC binding. Together with a very diverse hydrogen-bonding pattern, this has the potential to explain the experimentally observed promiscuity of PDH towards different sugars. Based on geometrical analysis, we propose a similar reaction mechanism as in P2O involving a general base proton abstraction, stabilization of the transition state, an alkoxide intermediate, through interaction with a protonated catalytic histidine followed by a hydride transfer to the flavin N5 atom. Our data suggest that the presence of the two potential catalytic bases His-512 and His-556 increases the versatility of the enzyme, by employing the most suitably oriented base depending on the substrate and its orientation in the active site. Our findings corroborate and rationalize the experimentally observed dioxidation of GLC by PDH and its promiscuity towards different sugars.
COBISS.SI-ID: 5218330
The outbreak of avian influenza A (H5N1) virus has raised a global concern for both the animal as well as human health. Besides vaccination, that may not achieve full protection in certain groups of patients, inhibiting neuraminidase or the transmembrane protein M2 represents the main measure of controlling the disease. Due to alarming emergence of influenza virus strains resistant to the currently available drugs, development of new neuraminidase N1 inhibitors is of utmost importance. The present paper provides an overview of the recent advances in the design of new antiviral drugs against avian influenza. It also reports findings in binding free energy calculations for nine neuraminidase N1 inhibitors (oseltamivir, zanamivir, and peramivir -carboxylate, -phosphonate, and -sulfonate) using the Linear Interaction Energy method. Molecular dynamics simulations of these inhibitors were performed in a free and two bound states – the so called ˝open˝ and ˝closed˝ conformations of neuraminidase N1. Obtained results successfully reproduce the experimental binding affinities of the already known neuraminidase N1 inhibitors, i.e. peramivir being a stronger binder than zanamivir that is in turn stronger binder than oseltamivir, or phosphonate inhibitors being stronger binders than their carboxylate analogues. In addition, the newly proposed sulfonate inhibitors are predicted to be the strongest binders – a fact to be confirmed by their chemical synthesis and a subsequent test of their biological activity. Finally, contributions of individual inhibitor moieties to the overall binding affinity are explicitly evaluated to assist further drug development towards inhibition of the H5N1 avian influenza A virus.
COBISS.SI-ID: 5396250
ProBiS is a new method to identify the binding site of protein through local structural alignment against the nonredundant Protein Data Bank (PDB), which may result in unique findings compared to the energy-based, geometry-based, and sequence-based predictors. In this work, binding sites of Hemagglutinin (HA), which is an important target for drugs and vaccines in influenza treatment, have been revisited by ProBiS. For the first time, the identification of conserved binding sites by local structural alignment across all subtypes and strains of HA available in PDB is presented. ProBiS finds three distinctive conserved sites on HA's structure (named Site 1, Site 2, and Site 3). Compared to other predictors, ProBiS is the only one that accurately defines the receptor binding site (Site 1). Apart from that, Site 2, which is located slightly above the TBHQ binding site, is proposed as a potential novel conserved target for membrane fusion inhibitor. Lastly, Site 3, located around Helix A at the stem domain and recently targeted by cross-reactive antibodies, is predicted to be conserved in the latest H7N9 China 2013 strain as well. The further exploration of these three sites provides valuable insight in optimizing the influenza drug and vaccine development.
COBISS.SI-ID: 5298970