CROSSLINKING STUDIES ON THE ORGANIZATION OF THE 16-S RIBOSOMAL-RNA WITHIN THE 30-S ESCHERICHIA-COLI RIBOSOMAL-SUBUNIT

The location and frequency of RNA crosslinks induced by photoreaction of hydroxymethyltrimethylpsoralen with 30 S Escherichia coli ribosomal subunits have been determined by electron microscopy. At least seven distinct crosslinks between regions distant in the 16 S rRNA primary structure are seen in the inactive conformation of the 30 S particle. All correspond to crosslinked features seen when the free 16 S rRNA is treated with hydroxymethyltrimethylpsoralen. The most frequently observed crosslink occurs between residues near one end of the molecule and residues about 600 nucleotides away to generate a loop of 570 bases. The size and orientation of this feature indicate it corresponds to the crosslinked feature located at the 3′ end of free 16 S rRNA. When active 30 S particles are crosslinked in 5 mm-Mg2+, six of the seven features seen in the inactive 30 S particle can still be detected. However, the frequency of several of the features, and particularly the 570-base loop feature, is dramatically decreased. This suggests that the long-range contacts that lead to these crosslinks are either absent or inaccessible in the active conformation. Crosslinking results in some loss of functional activities of the 30 S particle. This is consistent with the notion that the presence of the crosslink that generates the 570-base loop traps the subunit in an inactive form, which cannot associate with 50 S particles. The arrangement of the interacting regions crosslinked by hydroxymethyltrimethylpsoralen suggests that the RNA may be organized into three general domains. A striking feature of the Crosslinking pattern is that three of the seven products involve regions near the 3′ end of the 16 S rRNA. These serve to tie together large sections of rRNA. Thus structural changes at the 3′ end could, in principle, be felt through the entire 30 S particle. © 1979.