Expression and properties of recombinant HbA2 (alpha2delta2) and hybrids containing delta-beta sequences

Hemoglobin A2 (alpha2delta2), 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 alpha and delta 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 alpha2delta2 expressed in yeast is similar to that found for adult hemoglobin A and for fetal hemoglobin F when the alpha + beta or the alpha + gamma 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.