α-Hemoglobin Stabilizing Protein (AHSP) Markedly Decreases the Redox Potential and Reactivity of α-Subunits of Human HbA with Hydrogen Peroxide

Abstract

Background: ␣-Hemoglobin stabilizing protein (AHSP) modifies the redox properties of bound ␣-subunits. Results: Isolated hemoglobin subunits exhibit significantly different redox properties compared with HbA. A significant decrease in the reduction potential of ␣-subunits bound to AHSP results in preferential binding of ferric ␣. Conclusion: AHSP⅐␣-subunit complexes do not participate in ferric-ferryl heme redox cycling. Significance: AHSP binding to ␣-subunits inhibits subunit pseudoperoxidase activity.

Post-translational Transformation of Methionine to Aspartate Is Catalyzed by Heme Iron and Driven by Peroxide

“…The apparent differences in redox reactivity correlate with our previously reported studies indicating that ␣ subunits form fewer transient ferryl intermediates and accumulate much smaller amounts of detectable protein radicals when exposed to H 2 O 2 in solution (22). This difference appears to be due to rapid autoreduction rates of ferryl ␣ chains, which, in turn, have been attributed in part to the proximity of ␣Tyr-42 to the heme iron atom.…”

“…A signal was observed at g ϭ 6 and represented high spin ferric heme, and a second signal was observed at g ϭ 2 and represented the tyrosine free radical as described in previous publications (22,37). Both the ferryl heme and its associated protein cation radical induce a wide variety of oxidative reactions affecting both the protein itself and nearby molecules due to their high midpoint redox potentials (E 1/2 0 ϳ1.0 V) (38).…”

“…1B) data point plotted. Rate constants with standard deviations were obtained by averaging data from at least three different experiments and fitting each time course to a singleexponent expression with an offset as described previously (22 (24). Optical absorbance experiments were performed using either an Agilent Cary 100 or an Agilent 8453 UV-visible light optical absorbance spectrophotometer (Agilent).…”

“…This is reflected by the accumulation of ferryl intermediates in the ␤ subunit of oxidized Hb (22). To follow-up these studies, we examined the positional equivalent Met(E11) mutations in HbA ␤V67M and ␣V62M, respectively.…”

Iron accelerates hemoglobin oxidation increasing mortality in vascular diseased guinea pigs following transfusion of stored blood

“…Autoxidation experiments were carried out as described previously (35). Hb (100 μM) was incubated in phosphate buffered saline (pH 7.4), and visible spectra (400-700 nm) were recorded at every 10 minutes for a 4-hour incubation at 37°C.…”