Binding sites of ribosomal proteins on prokaryotic 5S RNAs: a study with ribonucleases

Abstract

The binding sites of ribosomal proteins L18 and L25 on 5S RNA from Escherichia coli were probed with RNases A, T1 and T2 and a double helix specific cobra venom endonuclease. The results for the protein-RNA complexes, which were compared with those for the free RNA, reveal an extensive interaction site for protein L18 and a more localized one for L25. Generally comparable results, with a few important differences, were obtained in a study of the binding sites of the 2 E. coli proteins on Bacillus stearothermophilus 5S RNA. Several protein-induced changes in the RNA structures were identified; some are possibly allosteric in nature. The 2 prokaryotic 5S RNA were also incubated with total 50S subunit proteins from E. coli and B. stearothermophilus ribosomes. Homologous and heterologous reconstitution experiments were performed for both RNA. The effects of the bound proteins on RNase digestion of the RNA could generally be correlated with the results obtained with the E. coli proteins L18 and L25, although there was evidence for an additional protein-induced conformational change in the B. stearothermophilus 5S RNA, which may have been due to a 3rd ribosomal protein L5.