Loss of allosteric behaviour in recombinant hemoglobin α2β29 r)F8)His→Ala: Restoration upon addition of strong effectors

Abstract

In the stereochemical model proposed by Perutz [1], the Fe-His(F8) bond plays a significant role in the allosteric transition in hemoglobin and the resulting cooperativity in ligand binding. When this bond is ruptured, there is a loss in the transmission of the information concerning ligand binding; examples are Hb(NO) 4 in the presence of inositol hexakisphosphate (IHP), or nickel substituted Hb hybrids which, despite being liganded, exhibit deoxy-like properties. To study the effects of the loss of the iron proximal histidine bond, we have engineered the α 2 β 2(F8)H92A recombinant Hb. The replacement of the highly conserved proximal histidine F8 residue by an alanine results in a low affinity for the heme group and a loss of the allosteric properties; kinetics of CO recombination after photodissociation show only the rapid bimolecular phase, characteristic of the high affinity R-state. However, a significant amount of deoxy (T-state) kinetics are observed after addition of external effectors such as IHP. The iron-histidine bond is apparently crucial for the heme-heme interaction, but the allosteric equilibrium may still be influenced by external constraints.