The Effects on Oxygen Affinity and Gelation of Hemoglobin S Crosslinked by Reaction with Methyl Acetimidate

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Abstract
The contribution of the high MW Hb (HMW Hb) to the antisickling effect produced by treatment of sickle cell erythrocytes with methyl acetimidate (MAI) was investigated. Erythrocytes obtained from sickle cell anemia and normal individuals were incubated with varying concentration of MAI. The presence of intermolecularly crosslinked HMW Hb was detected by gel filtration of dialyzed hemolysates obtained from the incubated cells. HMW Hb has an increased oxygen affinity and a decreased Hill constant. Intermolecular crosslinking per se had no additional effect on Hb oxygen affinity other than that due to Hb amino group modification. Modified deoxy Hb S of normal MW has a higher minimum gel concentration (MGC) than control deoxy Hb S. Deoxy HMW Hb did not gel at intracellular Hb concentrations. The incracellular concentration of deoxy Hb S that gels necessarily decreases as the HMW Hb concentration increases. This excluded volume effect of the non-gelling Hb would be expected to increase the MGC of completely deoxygenated hemolysate obtained from the treated cells. The HMW Hb contributes to the inhibition of sickling resulting from treatment of sickle erythrocytes with MAI by increasing both MGC and oxygen affinity of the modified Hb.