Novel Studies on a “Silent” High Affinity Mutant Hemoglobin (San Diego, β 109 Val ⇒ Met)

Abstract

A patient with a “silent” mutant hemoglobin characterized by high oxygen affinity and erythrocytosis is described. A novel approach was used to identify the mutant chain. Functionally active α and β chains were prepared from hemolysates of the patient and a normal control. Hybrid tetramers of patient's α chain with normal β chain and normal α chain with patient's β chain were prepared. Functional studies revealed that the patient's β chains had a higher oxygen affinity (P₅₀. 1.1 torr) than normal β chains (P₅₀, 1.7 torr) and the hybrid containing the patient's β chains had a P₅₀ similar to the patient's “stripped” hemolysate. It was assumed Therefore that the mutation was in the β chain; structural studies using cyanogen bromide cleavage revealed that the patient had Hb San Diego, β 109 Val⇒ Met, and that the patient's cells contained approximately 50 percent mutant hemoglobin.

Functional studies were performed on a. series of mixtures of patient's hemolysate diluted with varying amounts of normal hemo-lysate. Regression analysis of the data extrapolated to an assumed 100% Hb San Diego gave values for P₅₀ and Hill's constant of 4.8 torr and 1.0. It is proposed that this method, not previously reported, may be useful in evaluating the functional properties of “silent” mutant hemoglobins that cannot be studied in the purified state.