Crystallization and preliminary X-ray structural studies of hemoglobin A 2 and hemoglobin E, isolated from the blood samples of β-thalassemic patients

Abstract

Hemoglobin A 2 (α 2δ 2), a minor (2–3%) component of circulating red blood cells, acts as an anti-sickling agent and its elevated concentration in β-thalassemia is a useful clinical diagnostic. In β-thalassemia major, where there is a failure of β-chain production, HbA 2 acts as the predominant oxygen deliverer. Hemoglobin E, is another common abnormal hemoglobin, caused by splice site mutation in exon 1 of β globin gene, when combines with β-thalassemia, causes severe microcytic anemia. The purification, crystallization, and preliminary structural studies of HbA 2 and HbE are reported here. HbA 2 and HbE are purified by cation exchange column chromatography in presence of KCN from the blood samples of individuals suffering from β-thalassemia minor and Eβ-thalassemia. X-ray diffraction data of HbA 2 and HbE were collected upto 2.1 and 1.73 Å, respectively. HbA 2 crystallized in space group P2 1 with unit cell parameters a=54.33 Å, b=83.73 Å, c=62.87 Å, and β=99.80° whereas HbE crystallized in space group P2 12 12 1 with unit cell parameters a=60.89 Å, b=95.81 Å, and c=99.08 Å. Asymmetric unit in each case contains one Hb tetramer in R 2 state.