Expression and properties of recombinant HbA₂ (α₂δ₂) and hybrids containing δ-β sequences

Hemoglobin A₂ (α₂δ₂), which is present at low concentration (1-2%) in the circulating red cells of normal individuals, has two important features that merit its study, i.e., it inhibits polymerization of sickle HbS and its elevated concentration in some thalassemias is a useful clinical diagnostic. However, reports on its functional properties regarding O₂ binding are conflicting. We have attempted to resolve these discrepancies by expressing, for the first time, recombinant hemoglobin A₂ and systematically studying its functional properties. The construct expressing HbA₂ contains only α and δ genes so that the extensive purification required to isolate natural HbA₂ is circumvented. Although natural hemoglobin A₂ is expressed at low levels in vivo, the amount of recombinant α₂δ₂ expressed in yeast is similar to that found for adult hemoglobin A and for fetal hemoglobin F when the α + β or the α + γ genes, respectively, are present on the construct. Recombinant HbA₂ is stable, i.e., not easily oxidized, and it is a cooperative functional hemoglobin with tetramer-dimer dissociation properties like those of adult HbA. However, its intrinsic oxygen affinity and response to the allosteric regulators chloride and 2,3-diphosphoglycerate are lower than the corresponding properties for adult hemoglobin. Molecular modeling studies which attempt to understand these properties of HbA₂ are described.