Capture of activated dioxygen intermediates at the copper-active site of a lytic polysaccharide monooxygenase

Metalloproteins perform a diverse array of redox-related reactions facilitated by the increased chemical functionality afforded by their metallocofactors. Lytic polysaccharide monooxygenases (LPMOs) are a class of copper-dependent enzymes that are responsible for the breakdown of recalcitrant polysaccharides via oxidative cleavage at the glycosidic bond. The activated copper-oxygen intermediates and their mechanism of formation remains to be established. Neutron protein crystallography which permits direct visualization of protonation states was used to investigate the initial steps of oxygen activation directly following active site copper reduction in Neurospora crassa LPMO9D. Herein, we cryo-trap an activated dioxygen intermediate in a mixture of superoxo and hydroperoxo states, and we identify the conserved second coordination shell residue His157 as the proton donor. Density functional theory (DFT) calculations indicate that both active site states are stable. The hydroperoxo formed is potentially an intermediate in the mechanism of hydrogen peroxide formation in the absence of substrate. We establish that the N-terminal amino group of the copper coordinating His1 remains doubly protonated directly following molecular oxygen reduction by copper. Aided by mining minima free energy calculations we establish His157 conformational flexibility in solution that is abolished by steric hindrance in the crystal. A neutron crystal structure of NcLPMO9D at low pH supports occlusion of the active site which prevents protonation of His157 at acidic conditions.

Plasma Disinfection: Cold Atmospheric Pressure Plasma Attenuates SARS-CoV-2 Spike Protein Binding to ACE2 Protein and the RNA

Cold atmospheric pressure (CAP) plasma has a profound effect on protein-protein interactions. In this work, we have highlighted the deactivation of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Spike protein by CAP plasma treatment. Complete deactivation of spike protein binding to the human ACE2 protein was observed within exposure time of 5 minutes and which is correlated to the higher concentration of hydrogen peroxide formation due to the interaction with the reactive oxygen species present in the plasma. On the other hand, it is established that CAP plasma is also capable of degrading RNA of SARS-CoV-2 virus which is also linked to hydrogen peroxide concentration. Results of this work could be useful in designing plasma-based disinfection systems over those based on environmentally hazardous chemicals-based disinfections.

DETERMINATION OF GLUCOSE OXIDASE ACTIVITY BY SPECTROPHOTOMETRIC METHOD

В статье рассматривается универсальная, чувствительная, быстрая и воспроизводимая методика определения активности глюкозооксидазы, основанная на окислении пероксидом водорода йодида калия в присутствии молибдата аммония и фотометрировании образующегося синего комплекса «йод-крахмал». Построен калибровочный график для определения концентрации пероксида водорода в реакционной смеси. Проведен анализ образования пероксида водорода в реакции окисления глюкозы глюкозооксидазой при варьировании начальной концентрации глюкозы. The article developed a universal, sensitive, fast and reproducible method for determining glucose oxidase activity, based on the oxidation of potassium iodide by hydrogen peroxide in the presence of ammonium molybdate and photometry of the resulting blue iodine-starch complex. A calibration graph is constructed to determine the concentration of hydrogen peroxide in the reaction mixture. Analysis of hydrogen peroxide formation in glucose oxidation reaction with glucose oxidase at variation of initial glucose concentration was performed.

Hydrogen Production via Pd-TiO2 Photocatalytic Water Splitting under Near-UV and Visible Light: Analysis of the Reaction Mechanism

Photocatalytic hydrogen production via water splitting using a noble metal on a TiO2 is a technology that has developed rapidly over the past few years. Specifically, palladium doped TiO2 irradiated with near-UV or alternatively with visible light has shown promising results. With this end in mind, strategically designed experiments were developed in the Photo-CREC Water-II (PCW-II) Reactor using a 0.25 wt% Pd-TiO2 under near-UV and visible light, and ethanol as an organic scavenger. Acetaldehyde, carbon monoxide, carbon dioxide, methane, ethane, ethylene, and hydrogen peroxide together with hydrogen were the main chemical species observed. A Langmuir adsorption isotherm was also established for hydrogen peroxide. On this basis, it is shown that pH variations, hydrogen peroxide formation/adsorption, and the production of various redox chemical species provide an excellent carbon element balance, as well as OH• and H• radicals balances. Under near-UV irradiation, 108 cm3 STP of H2 is produced after 6 h, reaching an 99.8% elemental carbon balance and 98.2% OH• and H• and radical balance. It is also proven that a similar reaction network can be considered adequate for the photoreduced Pd-TiO2 photocatalyst yielding 29 cm3 STP of H2 with 97.5% carbon and the 99.2% OH•–H• radical balance closures. It is shown on this basis that a proposed “series-parallel” reaction network describes the water splitting reaction using the mesoporous Pd-TiO2 and ethanol as organic scavenger.