Synthesis of hemoglobin Gun Hill: increased synthesis of the heme-free beta-GH globin chain and subunit exchange with a free alpha-chain pool.

Hemoglobin Gun Hill is an unstable mutant hemoglobin associated with mild compensated hemolysis. This abnormal protein has a deletion of five amino acids in the beta-chains. The deletion includes the heme-binding proximal histidine at position 92. The beta-chains of hemoglobin Gun Hill lack heme groups. Approximately 32% of the circulating hemoglobin in heterozygous subjects consists of the mutant hemoglobin. When reticulocytes were incubated with radioactive amino acid the specific activity of hemoglobin Gun Hill was three to six times that of hemoglobin A. Total incorporation of radioactivity into hemoglobin Gun Hill was two to three times that into hemoglobin A. There were 20-50% more total counts in beta-Gun Hill (beta(GH)) than in beta(A). These results indicate that in reticulocytes there was greater synthesis of the abnormal beta-chains than beta(A)-chains. The ratio of the specific activities of the alpha-chains of hemoglobin Gun Hill to the alpha-chains of hemoglobin A was 20: 1. There was evidence of a free pool of alpha-chains in the reticulocytes containing hemoglobin Gun Hill. After 10 min of incubation approximately 40% of the total alpha-chain radioactivity was in the free pool. When protein synthesis was blocked by incubation of reticulocytes with puromycin, the specific activity of the alpha-chains of hemoglobin Gun Hill continued to increase due to direct exchange of alpha-subunits between the free pool and preformed hemoglobin Gun Hill. Studies of the assembly of beta(A) and beta(GH) revealed that the rates of translation of the two polypeptide chains were equal and uniform. No evidence was obtained for the existence of "slow points" in the process of globin chain assembly. The studies also suggest that lack of strong heme-globin binding does not hinder the synthesis of globin chains.