Effect of methylation of the N6 position of the penultimate adenosine of capped mRNA on ribosome binding.

An RNA(2'-O-methyladenosine-N6)-methyltransferase isolated from HeLa cells was used to convert the ends of vaccinia virus mRNAs containing m7G(5')pppAm-to m7G(5')pppm6Am-. Under competitive conditions, there was no preferential binding of mRNAs containing m6Am residues within the capped ends relative to those containing Am either to wheat germ ribosomes at 70 mM potassium acetate or to reticulocyte ribosomes at 120 mM potassium acetate. Only at high concentrations of potassium acetate (200 mM) and at near-saturating mRNA concentrations, was a slight enrichment of mRNAs containing m6Am over those containing Am detected on reticulocyte ribosomes. It appears that unlike methylation of the N7 position of the terminal guanosine, methylation at the N6 position of the penultimate adenosine of mRNA has, at most, a small effect on binding to ribosomes under present in vitro conditions.