CONTRIBUTIONS OF MULTIPLE BASIC-AMINO-ACIDS IN THE C-TERMINAL REGION OF YEAST RIBOSOMAL-PROTEIN L1 TO 5-S RIBOSOMAL-RNA BINDING AND 60-S RIBOSOME STABILITY

Previous studies suggest that the C-terminal region of ribosomal prote in L1 from Saccharomyces cerevisiae is important for its interaction w ith the 5 S rRNA molecule. Within this region are several highly conse rved basic amino acids including Lys276, Lys279, Lys289, Arg282, Arg28 5. To examine potential contributions of these amino acids to RNA-prot ein interaction and ribosomal assembly, effects of substitutions of th ese residues by methionine either individually or in combinations were examined. A methionine substitution of any one of the lysine residues did not significantly affect RNA binding in vitro. The mutant RNPs we re as stable as the wild type RNP. Yeast transformants expressing thes e mutant proteins grew at the same rate as the wild-type. However, mut ant proteins containing substitutions of any two of these basic amino acids bound RNA weakly The resultant RNPs were significantly less stab le than the wild-type. Whereas cells expressing mutant L1 with a singl e substitution at 289 was not lethal, cells expressing mutant LI with any double substitutions involving Lys289 as one of the substituted am ino acids were lethal. These data suggest that Lys289 plays a key role in the binding of ribosomal protein L1 to 5 S rRNA. The other basic r esidues, particularly Arg282, and Arg285, in this region also contribu te to RNA binding. These residues are predicted to locate on the same side of an a helix. We would like to propose a structural model for th e yeast RNP that involves multiple contact sites located on one side o f the helix in the C terminus of the protein and the 5 S rRNA. These b asic amino acids also participate, directly or indirectly, in the inte raction of the RNP complex with other components of the 60 S ribosomal subunit.