Functional Properties of Hemoglobin Kempsey

Functional studies have been performed on hemoglobin Kempsey (&32gg asp-tasn), a variant associated with familial erythrocytosis.In most experiments, the properties of hemoglobin Kempsey were compared to those of hemoglobin A, isolated from the same chromatography column.Hemoglobin Kempsey had high oxygen affinity (psO at pH 7.2 was 10% that of hemoglobin A) and nearly absent subunit cooperativity.Its alkaline Bohr effect was less than one-half normal.The addition of organic phosphates (2,3-diphosphoglycerate and inositol hexaphosphate) resulted in a lowering of oxygen affinity and an increase in subunit cooperativity.The dissociation of oxygen from partially saturated intermediates of hemoglobin Kempsey was slow and monophasic with no wavelength dependency indicative of subunit heterogeneity.The combination of carbon monoxide with deoxyhemoglobin Kempsey in a solution of low ionic strength was explained best by the presence of three hemoglobin species in equilibrium: one slowly reacting form which was probably hemoglobin in the T state and comprised 10% of the total; and two rapidly reacting forms, hemoglobin dimer and tetramerit hemoglobin in the R state.The dimer-tetramer equilibrium is shifted to favor the tetramer by inositol hexaphosphate which also converts the rapidly reacting deoxytetramer to a slowly reacting form.The existence of a substantial proportion of deoxyhemoglobin Kempsey in the form of a dimer was supported by the following experiments.(a) The change in absorbance (430 nm) following the addition of inositol hexaphosphate to deoxyhemoglobin Kempsey was dependent upon hemoglobin concentration, and indicated a tetramer to dimer dissociation constant (&,.J of about 5 PM.(b) Haptoglobin bound to deoxyhemoglobin Kempsey about as rapidly as to oxyhemoglobin A. The former reaction was inhibited strongly by inositol hexaphosphate.(c) The rapid component in the reaction of deoxyhemoglobin Kempsey with carbon monoxide and inositol hexaphosphate (in the same syringe) was increased at low hemoglobin concentration.(d) The hybrid hemoglobin a2/3A/3Kempsey was relatively less stable than (~~p*fl'.Deoxyhemoglobin Kempsey resembled oxyhemoglobin A