THE (M)GAMMA-CHAIN OF HUMAN FETAL HEMOGLOBIN IS AN (A)GAMMA-CHAIN WITH AN IN-VITRO MODIFICATION OF GAMMA-141 LEUCINE TO HYDROXYLEUCINE

We have reanalyzed the structure of the gammaT-15 peptide from the min or (m)gamma chain of human hemoglobin (Hb) F. Amino acid analysis conf irmed that the Leu 141 residue was missing from position 9 of this pep tide, and liquid secondary ion mass spectrometry indicated that it was replaced, not by methionine (residue mass 131) as previously believed , but by an amino acid of mass 129. By analogy with the recently repor ted oxidation of the corresponding leucine at position gamma141 of the unstable Hb Atlanta, it appears that the (M)gamma chain also results from the oxidation of gamma141 to hydroxyleucine (residue mass 129). T he finding that the proportion of the (M)gamma chain increased when re d cell lysates were prepared with carbon tetrachloride prompted us to reinvestigate the oxidation mechanism involved in the formation of bet a141 hydroxyleucine in Hb Atlanta. Oxidation of the beta141 residue co uld be detected when carbon tetrachloride was used in the lysis protor ol, while conversion of oxyhemoglobin to carbon monoxyhemoglobin prior to carbon tetrachloride treatment prevented oxidation. It therefore a ppears that the hydroxylation of Leu 141 is not an in vivo process in the circulating red cell. Perhaps leucine at position 141 of the beta, gamma, and delta chains (and at position 136 of the alpha chain), whi ch forms a contact with heme and is located directly across the heme p late from the E helix, is oxidized to hydroxyleucine at a very low rat e forming minute amounts of modified chains; this process is accelerat ed by treatment with agents such as carbon tetrachloride and prolonged exposure to air.