Kinetic and Equilibrium Properties of Hemoglobin Kansas

Abstract

Abstract The reactions of hemoglobin Kansas (102 β Asn → Thr) have been examined by kinetic and equilibrium methods. The behavior of the T state toward oxygen, derived from oxygen binding and oxygen pulse experiments, appears similar to that of hemoglobin A. The behavior of the R tetramer is difficult to separate from that of the dimer (the dissociation constant, K4,2, for oxyhemoglobin Kansas is about 70 µm as against 1.5 µm for hemoglobin A), but correlation of kinetic and equilibrium data suggests that the R tetramer of hemoglobin Kansas has a lower affinity for oxygen than that of hemoglobin A. Carbon monoxide binding is slower than for hemoglobin A, chiefly because of a lower rate of binding by the β chains. Comparisons between carbon monoxide binding and release of 8-hydroxypyrene-1,3,6-trisulfonic acid, and reaction with p-hydroxymercuribenzoate suggest that a conformation change (T → R transition) occurs on ligand binding as with hemoglobin A. Detailed examination of the dissociation reactions from the liganded dimers of oxygen and of carbon monoxide shows abnormally rapid dissociation from the β chain. The hemoglobin-haptoglobin reaction shows that deoxyhemoglobin Kansas is too slightly dissociated to give a measurable reaction. This result is consistent with an abnormally low ligand affinity of the R form. It is suggested that the differences between hemoglobin Kansas and hemoglobin A are due, at least in part, to differences between the R forms, and that an R → T transition occurs normally in hemoglobin Kansas. The allosteric effector 2,3-diphosphoglycerate lowers the affinity of hemoglobin Kansas for oxygen, but the effect is much smaller than for hemoglobin A when measured at high concentrations of hemoglobin. The data are most consistent with the assumption that the binding constant for 2,3-diphosphoglycerate at the primary site on tetrameric, liganded hemoglobin Kansas is only slightly lower than that for the unliganded form.