Electron transfer parameters for Methemoglobin formation in mutant Hemoglobin α-chains

Hemoglobin mediated transport of dioxygen (O2) critically depends on the stability of the reduced (Fe2+) form of the Heme cofactors. Some protein mutations stabilize oxidized (Fe3+) state (Methemoglobin, Hb M) causing methemoglobinemia and can be lethal above 30 %. Majority of the analyses of factors influencing Hb oxidation are retrospective and give insights only for inner sphere mutations of Heme (His58, His87). Herein, we report the first all atom MD simulations on redox states and calculations of the Marcus ET parameters for the α-chain Hb oxidation and reduction rates for Hb M. The Hb (wild type), and most of the studied α-chain variants maintain globin structure except the Hb M Iwate (H87Y). Using linear response approximation we calculated average energy gaps (<ΔE>), total (λ), protein (λprot), solvent (λsolv) reorganization energies, and redox potentials (E°), and oxidation free energies (ΔG°). The total λ ranges from 0.685 – 0.730 eV in agreement with literature on Hb and similar Heme proteins. The mutants forming Hb M tend to lower the E° and thus stabilize the oxidized (Fe3+) state (e.g. the Hb Miyagi variant with K61E mutation). Solvent reorganization (λsolv 73 – 96 %) makes major contributions to λ, while protein reorganization (λprot) accounts for 27 – 30 % except for the Miyagi and J-Buda variants (λprot ∼ 4 %). Analysis of Heme-solvent H-bonding interactions among variants provide insights into the role of Lys61 residue in stabilizing Fe2+ state and ET parameters. The ET parameters provide valuable insights into the Hb oxidation to Hb M in agreement with the experimental data. Thus our methodology explains the effect of mutations on the structure, stability and Hb oxidation, and has potential for the prediction of methemoglobinemia.

A Probable Mechanism for Recurring Outbreak of Respiratory Syndrome During Autumn Season in Ahvaz City, Iran

Background: Recurring annual asthma or asthma-like outbreak in Ahvaz City, Iran, has become a serious public health problem. A toxic gas, a chemical compound, or an allergen with unknown nature and mechanism leads to severe asthma outbreaks each year. Bronchospasm, cough, dyspnea, chest tightness, and wheezing are the main symptoms of these attacks. Oxygen and anti-asthma therapy are effective treatments in outbreak management. Based on our findings, it is hypothesized that vast amounts of ammonium nitrate used in agriculture underlay high concentrations of nitrate. This substance is transported by PM10 and is the main cause of the respiratory outbreak. Methods: The concentration of nitrate was determined by the ion chromatography method in dust samples during 2015-2016. Using spectroscopic methods, the effect of nitrate ions on hemoglobin oxidation and methemoglobin formation was studied. Results: Our results indicate that the concentration of ammonium nitrate in dust collected from indoor spaces is five times higher than the levels allowed amount in airborne pollutants. Our findings also indicate that this concentration of nitrate in the bloodstream can instantly increase methemoglobin percent to 18% with symptomatic consequences. Conclusion: The exact mechanism proposed by this study is that inhaled nitrate or nitric acid finally increases the nitrate in the circulation of affected individuals. Hemoglobin oxidation and methemoglobin formation are the subsequent events leading to methemoglobinemia with asthma-like reactions seen in Ahvaz respiratory syndrome. Restricting or even prohibiting ammonium nitrate application in agriculture seems to be an urgent measure to stop Ahvaz's recurring respiratory syndrome.

Hemoglobin Oxidation in Stored Blood Accelerates Hemolysis and Oxidative Injury to Red Blood Cells

Introduction Maintaining blood supply is a challenge in blood banks. Red blood cells (RBCs) stored at 4°C experience issues of biochemical changes due to metabolism of cells, leading to changes collectively referred to as “storage lesions.” Oxidation of the red cell membrane, leading to lysis, contributes to these storage lesions. Methods Blood bags with CPD-SAGM stored at 4°C for 28 days were withdrawn aseptically on days 1, 14, and 28. Hematology analyzer was used to investigate RBC indices. Hemoglobin oxidation was studied through spectrophotometric scan of spectral change. RBC lysis was studied with the help of Drabkin's assay, and morphological changes were observed by light and scan electron microscopy. Results RBCs show progressive changes in morphology echinocytes and spherocytes on day 28. There was 0.85% RBC lysis, an approximately 20% decrease in percentage oxyhemoglobin, and a 14% increase in methemoglobin formation, which shows hemoglobin oxidation on day 28. Conclusions Oxidative damage to RBC, with an increase in storage time was observed in the present study. The observed morphological changes to RBC during the course of increased time shows that there is progressive damage to RBC membrane and a decrease in hemoglobin concentration; percentage RBC lysis is probably due to free hemoglobin and iron.

Maltose-mediated long-term stabilization of freeze- and spray- dried forms of bovine and porcine hemoglobin

Slaughterhouse blood represents a valuable source of hemoglobin, which can be used in the production of heme-iron based supplements for the prevention/treatment of iron-deficiency anemia. In order to obtain a stable solid-state formulation, the effect of maltose addition (30 %) on the stability and storage of bovine and porcine hemoglobin in powders obtained by spray and freeze-drying (without maltose: Hb; with maltose: HbM) were investigated. Differential scanning calorimetry of spray- and freeze-dried powders indicated satisfying quality of the formulation prepared with maltose on dissolving back into solution. After two-year storage at room temperature (20?5?C) in solid forms, protected from moisture and light, rehydrated spray- and freeze-dried HbM were red, while Hb were brown. Dynamic light scattering showed the presence of native hemoglobin monomers in rehydrated spray- and freeze-dried HbM, but their agglomerates in Hb samples. UV?Vis spectrophotometry confirmed an absence of significant hemoglobin denaturation and methemoglobin formation in HbM freeze-dried powders. In spray-dried HbM, an increased level of methemoglobin was detected. The results confirmed the stabilizing effect of maltose, and suggested its use in the production of long-term stable solid-state formulations of hemoglobin, along with drying processes optimization.