Immunological characterization of rabbit hemoglobin α and β chain synthesizing polysomes. Estimation of relative numbers of active α and β messenger ribonucleic acid

Anti hemoglobin chain antibodies, purified by immuno adsorption to Sepharose linked antigen and thereby made both RNase free and immunologically specific for either β or α chain, are readily bound to nascent hemoglobin chains attached to rabbit reticulocyte polysomes. Antibodies to other antigens, e.g. anti immunoglobulin G, are not bound. Use of chain specific antibodies enabled us to determine quantitatively the mean number of ribosomes associated with rabbit hemoglobin β and α messenger RNA. Such mean sizes of β and α polysomes could be calculated, for example, from density gradient profiles of polysomes previously coated with either ¹²⁵I labeled anti β chain or ¹²⁵I labeled anti α chain. Complementary estimates were obtained, pre and postimmunoprecipitation, from the A₂₆₀ density gradient profiles of polysomes and the profiles of β and α polysomes differentially labeled with [³H]isoleucine and [¹⁴C]valine. All estimates were similar; viz. the β:α ratio for mean number of ribosomes per mRNA ranged from 1.5 to 1.86. Among unaltered polysomes from the same rabbits, the β:α ratio for total relative numbers of synthetically active β and α ribosomes was calculated from y intercepts of Dintzis Naughton plots to be 1.04 to 1.17. With these two kinds of β:α ratios as a basis, the relative numbers of active α mRNA and β mRNA were found to be somewhat greater than previously described by Lodish and Jacobsen. For individual animals, the α mRNA:β mRNA ratio ranged from 1.28 to 1.78. The addition of the further 25% of α mRNA which is unattached to polysomes suggests that total α mRNA is 2 fold more abundant than β mRNA. (28 references.)