Abstract
To clarify the role of γN-terminal Gly, γ5 Glu, and γ143 Ser in 2, 3-biphosphosphoglycerate (BPG) binding to fetal hemoglobin (Hb F ), we engineered and produced normal human Hb F and two Hb F variants (Hb F γG1V, γS143H, and Hb F γG1V, γE5P, γS143H) using a yeast expression system and then compared their oxygen-binding properties with those of native human Hb F and adult Hb (Hb A). Oxygen affinity of Hb F γG1V, γS143H in the absence of 2, 3-BPG was slightly higher than that of normal Hb F. The decrease in oxygen affinities for Hb F γG1V, γS143H with increasing 2, 3-BPG concentrations was larger than that of normal Hb F, but significantly less than that of Hb A. In contrast, oxygen affinities of Hb F γG1V, γE5P, γS143H in the absence and presence of 2, 3-BPG were much lower than those of Hb F γG1V, γS143H and were similar to those of Hb A. These results indicate that differences between Pro and Glu at the A2 position in the A helix in Hb A and Hb F, respectively, are critical for reduced binding of 2, 3-BPG to Hb F, even though β5 Pro does not interact directly with 2, 3-BPG in Hb A. Hb F variants such as Hb F γG1V, γE5P, γS143H, which exhibit reduced oxygen affinity, should facilitate design of efficient antisickling fetal Hb variants for potential use in gene therapy for sickle cell disease.
The Role of Cysteine Residues in the Erythrocyte Plasma Membrane Anion Exchange Protein, AE1
