Mutations in conserved regions of ribosomal RNAs decrease the productive association of peptide-chain release factors with the ribosome during translation termination.
Early studies provided evidence that peptide-chain release factors (RFs) bind to both ribosomal subunits and trigger translation termination. Although many ribosomal proteins have been implicated in termination, very few data present direct biochemical evidence for the involvement of rRNA. Particularly absent is direct evidence for a role of a large subunit rRNA in RF binding. Previously we demonstrated in vitro that mutations in Escherichia coli rRNAs, known to cause nonsense codon readthrough in vivo, reduce the efficiency of RF2-driven catalysis of peptidyl-tRNA hydrolysis. This reduction was consistent with the idea that in vivo defective termination at the mutant ribosomes contributes to the readthrough. Nevertheless, other explanations were also possible, because still missing was essential biochemical evidence for that idea, namely, decrease in productive association of RFs with the mutant ribosomes. Here we present such evidence using a new realistic in vitro termination assay. This study directly supports in vivo involvement in termination of conserved rRNA regions that also participate in other translational events. Furthermore, this study provides the first strong evidence for involvement of large subunit rRNA in RF binding, indicating that the same rRNA region interacts with factors that determine both elongation and termination of translation.
Arkov
A L
AL
Department of Molecular Genetics (Box 11), The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, TX 77030, Houston, USA.
Freistroffer
D V
DV
Pavlov
M Y
MY
Ehrenberg
M
M
Murgola
E J
EJ
D001483:Base Sequence; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics*; D007700:Kinetics; D008958:Models, Molecular; D008962:Models, Theoretical; D009154:Mutation; D009690:Nucleic Acid Conformation; D010443:Peptide Chain Termination, Translational; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics; D012335:RNA, Ribosomal / Q000737:chemistry* / Q000235:genetics*; D012270:Ribosomes / Q000378:metabolism*
D012329:RNA, Bacterial; D012335:RNA, Ribosomal