Effect of ionizing radiation on haemoglobin : the oxy-derivative of haemoglobin Iwate

Abstract

It is well established that exposure of oxyhaemoglobin to ionizing radiation results in remarkably selective electron addition to the (FeO 2 ) unit, giving a novel species, (FeO 2 ) - , in which the extra electron is largely localized on iron and dioxygen. This work has now been extended to haemoglobin (Hb.) Iwate. The haemoglobin M. Iwate used is a mutant haemoglobin having only Fe(III) α -chains by oxy β-chains ( α 2 Met β 2 oxy ). The haem iron atoms in the α -chains are coordinated in the fifth site by a proximal tyrosine in place of histidine. This unit is a high-spin Fe(III) with axial symmetry and prominent electron spin resonance (ESR) features in the g = 6 and g = 2 regions. On exposure to 60 Co γ -rays at 77 K, efficient electron addition occurred at both types of iron centre, giving Fe(II) and (FeO 2 ) - units. The former was monitored by the decrease of the g = 6 feature for Fe(III) and the latter by the growth of g -features at 2.254 (g x ), 2.149 (g y ) and 1.967 (g z ). These values are close to those for the FeO 2 - centre formed in the β-chains of normal oxyhaemoglobin. On annealing above 77 K, two changes occurred : first there was a small but clear increase in g x and g y , followed by a marked reduction in g x and g y giving g-values close to those for the centre formed directly in the α -chains of the normal protein. Finally, this intermediate species gave a centre having g x = 2.310, g y = 2.180 and g z = 1.935. These values are typical of low-spin Fe(III) haemoglobin and are assigned to the protonated complex, Fe(III)O 2 H. Ultimately at ca . room temperature, this was converted into the high-spin, met -form, with a gain in the g = 6 feature. These results established that the β-chain centre in Hb. Iwate behave in the same way as isolated β-chains. They also confirm that electron addition to the oxy-units is facile, even in the presence of Fe(III) units in each tetramer. The results also confirm that electron capture to give (FeO 2 ) - units is not followed by internal electron-transfer to give Fe(II) from the Fe(III) centres in the α -chains.