Structural and functional studies on hemoglobin Bethesda (alpha2beta2 145His), a varient associated with compensatory erythrocytosis.

Studies have been performed on a 12-yr-old Chinese girl with compensatory erythrocytosis due to the presence of hemoglobin Bethesda comprising about 45% of the red cell hemoglobin. Her parents and three siblings were normal. The oxygen affinity of her blood was markedly increased: under physiological conditions (pH 7.40, 37 degrees C). P(50) was 12.8 mm Hg (normal = 26.5 mm Hg). The red cell 2,3-diphosphoglycerate (2.3-DPG) level was normal. The abnormal hemoglobin could not be separated from hemoglobin A by zone electrophoresis at pH 8.6 or isoelectric focusing on polyacrylamide gel. However, after the hemoglobin was split into free alpha and beta chains by treatment with p-hydroxymercuribenzoate (PMB) or 6 M urea, an abnormal beta chain was readily demonstrated having a higher isoelectric point (more positive net charge) than normal beta(A). Structural analysis of the variant beta chain demonstrated the substitution of histidine for tyrosine at position 145: hemoglobin Bethesda (alpha(2)beta(2) (145His)). From earlier chemical and crystallographic studies, it has been postulated that this residue is a critical determinant of hemoglobin function. Hemoglobin Bethesda was separated from hemoglobin A by column chromatography. Oxygen equilibria of purified hemoglobin Bethesda revealed an extremely high oxygen affinity (exceeding that of isolated alpha and beta chains), and markedly reduced cooperativity. The Bohr effect of hemoglobin Bethesda was 1/3 that of hemoglobin A. However, hemoglobin Bethesda showed a significant interaction with 2.3-DPG and inositol hexaphosphate.