Separation of reticulocyte initiation factor M 2 activity into two components.

Abstract

IF-M(2), one of three initiation factors isolated by DEAE-cellulose chromatography from the 0.5 M KCl-wash fraction of rabbit reticulocyte ribosomes, has been separated by Sephadex G-200 chromatography into two components: IF-M(2A) and IF-M(2B). IF-M(2A) elutes near the void-volume, while IF-M(2B), which is much smaller in molecular weight than IF-M(2A), elutes slightly after a hemoglobin marker. In the presence of the other appropriate factors, both IF-M(2A) and IF-M(2B) are required to stimulate poly(U)-directed polyphenylalanine synthesis at low Mg(++) concentration, ApUpG-directed Met-tRNA(F) binding to washed reticulocyte ribosomes, and initiation of globin synthesis from endogenous mRNA. IF-M(2A) stimulates ribosome-dependent GTP hydrolysis, while IF-M(2B) does not; IF-M(2B) stimulates ApUpG-directed fMet-tRNA(F) binding in the presence of IF-M(1), while IF-M(2A) does not. Although IF-M(2A) and IF-M(2B) can be distinguished from each other by size and by activity, a distinct function for IF-M(2B) has not yet been found. Therefore, its precise role in the initiation process remains unclear.