Destabilization of secondary structure in 16S ribosomal RNA by dimethylation of two adjacent adenosines

Abstract

Fragments comprising the 49 nucleotides from the 3′-end have been purified from 16s ribosomal RNA of wild-type Esoheriohia coli and from a kasugamycin-resistant mutant that specifically lacks dimethylation of two adjacent adenines near the 3′-terminus. These fragments, obtained after treatment of ribosomes in vitro with the bacteriocin cloacin DF ₁₃ , were used to study the effect of the methylgroups on the temperature dependent unfolding of double-stranded regions. Both fragments contain at least 3 independent melting transitions, of which the one with the highest T _m corresponds with the unfolding of a nine-basepair long central hairpin. Dimethylation of the adenines in the loop of this hairpin lowers the melting temperature (T _m ) by approximately 2°C at 0.2 M NaCl and by about 5°C at 0.015 M NaCl. It is suggested that m ₂⁶ Am ₂⁶ A is more antagonistic to loop formation than ApA and that the function of the methylgroups is to help to destabilize the 3′-terminal hairpin in 16S rRNA in order to facilitate intermolecular interactions.