Interaction of ribosomal proteins L25 from yeast and EL23 from E. coli with yeast 26S and mouse 28S rRNA.
The interaction of ribosomal protein EL23 from E. coli and L25 from yeast with yeast 26S rRNA was analysed by nitrocellulose filter binding and RNase protection experiments using both intact rRNA and various fragments prepared by in vitro transcription of cloned yeast rDNA regions in the SP6 system. The results show that EL23 efficiently and specifically interacts with the region of 26S rRNA previously identified as the binding site for the yeast ribosomal protein L25. A comparison of the oligonucleotides resulting from limited RNase T1 digestion of the heterologous EL23/26S rRNA complex with those obtained by the same treatment of the homologous L25/26S rRNA complex showed that the molecular details of the two r-protein/rRNA interactions are highly similar if not identical. Using the synthetic 26S rRNA fragments we could demonstrate that all information for the formation of a biologically active binding site is located within the region of the rRNA delimited by the sequences protected by L25 against RNase T1 digestion. Part of the sequence at the 3' end of the 5'-distal protected region, however, was found not to be essential for r-protein binding although it does enhance the efficiency of this binding. Binding experiments using synthetic mouse 28S rRNA fragments showed that neither EL23 nor L25 interact with the structural equivalent of their respective cognate binding sites present in this mammalian rRNA. We argue that the structure of the expansion sequence present in this region of mouse 28S rRNA is a major cause of this failure.
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D002872:Chromosome Deletion; D004275:DNA, Ribosomal / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D005656:Fungal Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids; D012335:RNA, Ribosomal / Q000378:metabolism*; D012339:RNA, Ribosomal, 28S / Q000235:genetics / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013045:Species Specificity
D001426:Bacterial Proteins; D004275:DNA, Ribosomal; D029968:Escherichia coli Proteins; D005656:Fungal Proteins; D012335:RNA, Ribosomal; D012339:RNA, Ribosomal, 28S; D012269:Ribosomal Proteins; C043816:ribosomal protein L23, E coli; C031127:ribosomal protein L25