Formation of Superoxide in the Autoxidation of the Isolated alpha and beta Chains of Human Hemoglobin and Its Involvement in Hemichrome Precipitation

Abstract

The purpose of the work reported here is to look at 0; production during autoxidation df the isolated a and fi chains of human hemoglobin. The formation of superoxide by isolated chains, in fact, may be involved in pathological phenomena observed in red blood cell diseases where excess of either a or fi chains is produced because of inherited defects of synthesis (thalassemus).The autoxidation of the oxygenated chains of human hemoglobin is followed by the transformation of the oxidized molecule (high-spin Fe3+) into a species absorbing as a low-spin Fe3+ compound, that is a hemichrome, which tends to precipitate. The overall process may be described by the following reactions :Oxychains -% Ferric chains % Hemichrome % Precipitate.Depending on conditions the different steps proceed with different relative rates and therefore may overlap to a greater or lesser extent.The results obtained indicate that superoxide is produced during autoxidation of the isolated a and B chains of human hemoglobin. This conclusion has been reached on the basis of an experimental approach based on cooxidation of epinephrine to adrenochrome during 0; production and inhibition of this effects by superoxide dismutase. In addition the rate of precipitation is, in all cases, very much reduced upon addition of either superoxide dismutase or catalase. These facts seem to imply very strongly that oxygen radicals are involved also in the precipitation following oxidation and hemichrome formation in the chains. In particular the effect of superoxide dismutase implies the involvement of 0,. Thus production of 0; ,could well be one of the important events leading to alteration of the red cell membrane and consequently to reduced life span of thalassemic erythrocytes.

Insights into Hemoglobin Assembly through in Vivo Mutagenesis of α-Hemoglobin Stabilizing Protein

“…Fully assembled HbA is very stable in comparison with isolated ␣ and ␤ subunits in their heme (Fe 2ϩ )-or hemin (Fe 3ϩ )-containing (holo) and heme-lacking (apo) forms (4,5). Free ␣ subunits, especially the Fe 3ϩ (ferric) form, tend to catalyze the production of reactive oxygen species (ROS), denature, and form damaging precipitates (4,6,7).…”

The Role of Alpha-Hemoglobin Stabilizing Protein in Redox Chemistry, Denaturation, and Hemoglobin Assembly

“…We have been unable to find homologous protein or DNA sequences in the other vertebrates, most of which have nucleated red cells. Because free a H chains are unstable (14,56,92), an early hypothesis was that AHSP acts as a molecular chaperone for free a chains prior to their incorporation into HbA (38,62). In support of this idea, it has been shown that: (a) AHSP associates with a H chains in solution, but not with b H chains or HbA; (b) HbA forms as b H chains are titrated into solutions containing AHSP:a H -chain duplexes; and (c) AHSP does not remain bound to HbA following the titration (62).…”

Oxidative damage in trout erythrocyte in response to “in vitro” copper exposure

“…Fridovich, 1972; Brunori et al, 1975) and thereby of products such as H 2 O 2 or hydroxyl radical, 8 which can derive from O 2 .-itself. 9 We have observed that the presence of copper sulfate does not influence the level of ROS during 1 10 hr of HbI or HbIV incubation (Figs.3-4).…”