TY - JOUR
T1 - Expression and properties of recombinant HbA2 (α2δ2) and hybrids containing δ-β sequences
AU - Inagaki, K.
AU - Inagaki, J.
AU - Dumoulin, A.
AU - Padovan, J. C.
AU - Chait, B. T.
AU - Popowicz, A.
AU - Manning, L. R.
AU - Manning, J. M.
PY - 2000/12/1
Y1 - 2000/12/1
N2 - 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.
AB - 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.
KW - Allosteric regulator
KW - Delta gene
KW - Hemoglobin
KW - Tetramer strength
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U2 - 10.1023/A:1007196118200
DO - 10.1023/A:1007196118200
M3 - Article
C2 - 11307949
AN - SCOPUS:0034466912
VL - 19
SP - 649
EP - 662
JO - Protein Journal
JF - Protein Journal
SN - 1572-3887
IS - 8
ER -