Bases defining an ammonium and magnesium ion-dependent tertiary structure within the large subunit ribosomal RNA

A 58 nucleotide RNA derived from a highly conserved domain of the large subunit ribosomal RNA (Escherichia coli 1051 to 1108) has a set of tertiary interactions that is stabilized by NH4+ and Mg2+ in preference to other ions. We have mapped the nucleotides contributing to this structure by examining the thermal denaturation of 25 sequence variants. Where necessary compensatory mutations were made to preserve the phylogenetically conserved secondary structure. Substitutions of bases or base-pairs at eight positions specifically eliminate the ion-dependent tertiary structure without affecting the secondary structure stability; most of these positions are conserved among all large subunit RNA sequences. At two positions, substitutions of bases found in other organisms stabilize the E. coli tertiary structure by substantial amounts (delta G 37 degrees becomes more favorable by -1.8 to -4.5 kcal/mol). One of these variants disrupts a potential A.U base-pair within a helix, suggesting that the tertiary structure competes with alternative structures. The results show that this rRNA domain contains an extensive, highly conserved, and very stable set of tertiary interactions. The sequence in E. coli, and probably most other organisms, has not evolved to maximum stability. It is possible that natural selection has "tuned" the tertiary structure to an optimum stability, perhaps because the structure must open and close during the ribosome cycle.