Synthesis of alpha and beta chains of hemoglobin was studied in vitro in intact reticulocytes and bone marrow cells. The cells were from rabbits having a variant form of hemoglobin in which L-isoleucine is in the alpha but not in the beta chains. This characteristic permitted a selective inhibition of alpha-chain synthesis to be produced by addition to the incubation medium of L-O-methylthreonine, an inhibitor of protein synthesis that is a specific antagonist of L-isoleucine. In studies with reticulocytes, 25 mM L-O-methylthreonine produced a 60-70% inhibition of alpha-chain synthesis, but beta-chain synthesis was unaffected even after incubation times for 4 hr. Because reticulocytes contain a pool of uncombined alpha chains which might have obscured the demonstration of an alpha chain-dependent mechanism for beta-chain synthesis, subsequent studies were done with bone marrow cells. The latter had little or no detectable alpha-chain pool. A substantial inhibition of alpha-chain synthesis by the bone marrow cells was produced by the isoleucine antagonist but was also accompanied by a significantly decreased rate of beta-chain synthesis. These findings suggest that the coordinated synthesis of the complementary alpha- and beta-globin chains of hemoglobin may reflect in part a modifying effect of alpha-chain synthesis on the synthesis of beta chains.

Arginine↗

PubMed1973

Hemoglobin Abraham Lincoln, beta32 (B14) leucine leads to proline. An unstable variant producing severe hemolytic disease.

An unstable hemoglobin variant was identified in a Negro woman with hemolytic anemia since infancy. A splenectomy had been performed when the patient was a child. The anemia was accompanied by erythrocyte inclusion bodies and excretion of darkly pigmented urine. Neither parent of the proposita demonstrated any hematologic abnormality, and it appeared that this hemoglobin variant arose as a new mutation. Erythrocyte survival in the patient was greatly reduced: the erythrocyte t(1/2) using radiochromium as a tag was 2.4 days, and a reticulocyte survival study performed after labeling the cells with L-[(14)C]leucine indicated a t(1/2) of 7.2 days. When stroma-free hemolysates were heated at 50 degrees C, 16-20% of the hemoglobin precipitated. The thermolability was prevented by the addition of hemin, carbon monoxide, or dithionite, suggesting an abnormality of heme binding. An increased rate of methemoglobin formation was also observed after incubation of erythrocytes at 37 degrees C. The abnormal hemoglobin could not be separated from hemoglobin A by electrophoresis or chromatography, but it was possible to isolate the variant beta-chain by precipitation with p-hydroxymercuribenzoate. Purification of the beta-chain by column chromatography followed by peptide mapping and amino acid analysis demonstrated a substitution of proline for beta32 leucine. It appears likely that a major effect of this substitution is a disruption of the normal orientation of the adjacent leucine residue at beta31 to impair heme stabilization.

Adult↗

PubMed1973