Binding of carbon monoxide to mutant α chains of hemoglobin M Iwate; evidence for distal imidazole ligation

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Abstract
The optical contribution of the .beta. chains to the spectrum of Hb M Iwate (.alpha.87his.fwdarw. tyr)2.beta.2A was subtracted with the aid of a computer so that the spectrum of ferric .alpha.-chains was obtained. Tyrosinate binding to the heme is suggested from spectral resemblance to ferric heme phenolate in dimethyl sulfoxide. The slow reduction of the abnormal ferric .alpha.-chains in Hb M Iwate by dithionite was studied spectrophotometrically both in the presence and absence of CO. The rate of reduction was dependent on the ligation state of normal .beta.-chains. The CO-ligated form of the reduced .alpha.-chain bears strong spectral resemblance to the CO-ligated form of the reduced .beta.-chains suggesting similar structures for the heme-ligand complex. A model compound with similar optical properties to the CO-ligated protein can be prepared in dimethyl sulfoxide from hemin chloride, imidazole and CO using chromous acetate as the heme reductant. Substitution of phenolate for imidazole produces a spectral entity so different from that observed in the protein as to rule out tyrosinate ligation to ferrous heme of the .alpha. chains when CO is bound.