Modification of α-chain or γ-chain heme pocket polarity by Val(E11) → Thr substitution has different effects on the steric, dynamic, and functional properties of human recombinant hemoglobin. Deoxy derivatives

The dynamic and functional properties of mutant deoxyhemoglobins in which either the β-globin Val⁶⁷(E11) or the α-globin Val⁶²(E11) is replaced by threonine have been investigated through the thermal evolution of the Soret absorption band in the temperature range 300 to 20 K and through the kinetics of CO rebinding after flash photolysis at room temperature. The conformational properties of the modified α chain and β chain distal heine pockets were also studied through x-ray crystallography and molecular modeling. The data obtained with the various techniques consistently indicate that the polar isosteric mutation in the distal side of the α chain heme pocket has a larger effect on the investigated properties than the analogous mutation on the β chain. We attribute the observed differences to the presence of a water molecule in the distal heme pocket of the modified a chains, interacting with the hydroxyl of the threonine side chain. This is indicated by molecular modeling which showed that the water molecule present in the α chain distal heme pocket can bridge by H bonding between Thr⁶²(E11) and His⁵⁸(E7) without introducing any unfavorable steric interactions. Consistent with the dynamic and functional data, the presence of a water molecule in the distal heine pocket of the modified β chains is not observed by x-ray crystallography.