Structure and Synthesis of the Unstable Hemoglobin Sabine (α2β291 Leu → Pro)

Hemoglobin Sabine (β91 Leu→Pro) is an unstable molecule present at low concentrations (12% of total hemoglobin) in the peripheral blood of one individual. The following results of structural studies demonstrated that hemoglobin Sabine existed predominantly as an α2β2 tetramer with two heme groups associated with the α chains only. (a) Quantitative analyses of the heme and globin parts showed that hemoglobin Sabine had a molar heme-globin ratio of one-half that of hemoglobin A. (b) Hemoglobin Sabine had an s20, w of 4.04 compared to 4.34 for hemoglobin A. (c) Dissociation of the α and β chains by treatment with p-chloromercuribenzoate showed that heme was associated with α chains only. Moreover, 14C-labeled hemoglobin Sabine co-migrated with 3H-labeled α2hβ2o semihemoglobin, an artificially constructed tetramer of globin A chains having heme groups associated with only the α chains. When hemin was added to this preparation, the electrophoretic mobility of the semihemoglobin, but not that of hemoglobin Sabine, was changed to that of normal hemoglobin A. This result suggests that hemoglobin Sabine lacks a full complement of heme groups because of a decreased affinity for heme. There was a balanced production of total cellular heme and globin chains, as determined by [14C]glycine incorporation into these components. When blood cells were incubated with [3H]leucine, the rates of incorporation of radioactivity into hemoglobins Sabine and A were similar. Analyses of the rates of labeling of the individual α and β chains in both hemoglobins showed that initially unlabeled α chains combined with labeled, newly synthesized β chains. However, there was a rapid turnover of newly synthesized hemoglobin Sabine β chains, and, concomitantly, as time progressed, a significant increase in the α chain fraction of radioactivity in hemoglobin Sabine compared to that in hemoglobin A. When cells were pulse labeled for short time periods the rate of synthesis of the β chain in hemoglobin Sabine was found equal to that of the β chain in hemoglobin A. A soluble pool of rapidly labeled α chains was detected which appeared to be precursors to both of the completed hemoglobins. A model of hemoglobin Sabine synthesis and assembly is suggested in which (a) the lower blood concentration occurs entirely because of rapid removal of some of the newly synthesized βSabine-globin chains from the soluble phase of the cells and not because of decreased chain synthesis, and (b) newly synthesized α chains are brought into the hemoglobin Sabine molecule primarily by an exchange between the pool of soluble α chains and the pre-existing hemoglobin Sabine α chains. This model is in agreement with those proposed by other investigators in their studies of unstable hemoglobins of a similar nature.