Roles of helix H69 of 23S rRNA in translation initiation.
Initiation of translation involves the assembly of a ribosome complex with initiator tRNA bound to the peptidyl site and paired to the start codon of the mRNA. In bacteria, this process is kinetically controlled by three initiation factors--IF1, IF2, and IF3. Here, we show that deletion of helix H69 (∆H69) of 23S rRNA allows rapid 50S docking without concomitant IF3 release and virtually eliminates the dependence of subunit joining on start codon identity. Despite this, overall accuracy of start codon selection, based on rates of formation of elongation-competent 70S ribosomes, is largely uncompromised in the absence of H69. Thus, the fidelity function of IF3 stems primarily from its interplay with initiator tRNA rather than its anti-subunit association activity. While retaining fidelity, ∆H69 ribosomes exhibit much slower rates of overall initiation, due to the delay in IF3 release and impedance of an IF3-independent step, presumably initiator tRNA positioning. These findings clarify the roles of H69 and IF3 in the mechanism of translation initiation and explain the dominant lethal phenotype of the ∆H69 mutation.
Ohio State Biochemistry Program, Department of Microbiology, and Center for RNA Biology, The Ohio State University, Columbus, OH 43210.
Qi
Q
Liu
Ohio State Biochemistry Program, Department of Microbiology, and Center for RNA Biology, The Ohio State University, Columbus, OH 43210 fredrick.5@osu.edu.
Kurt
K
Fredrick
D018387:Codon, Initiator; D004926:Escherichia coli / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D020128:Inhibitory Concentration 50; D009154:Mutation; D009690:Nucleic Acid Conformation; D009711:Nucleotides / Q000737:chemistry; D010448:Peptide Initiation Factors / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010641:Phenotype; D014176:Protein Biosynthesis; D017433:Protein Structure, Secondary; D012338:RNA, Ribosomal, 23S / Q000737:chemistry*; D012343:RNA, Transfer / Q000737:chemistry; D012358:RNA, Transfer, Met / Q000235:genetics*; D012270:Ribosomes / Q000737:chemistry
D018387:Codon, Initiator; D009711:Nucleotides; D010448:Peptide Initiation Factors; D010455:Peptides; D012338:RNA, Ribosomal, 23S; D012358:RNA, Transfer, Met; D012343:RNA, Transfer
IF2; IF3; fMet-tRNA; ribosome; start codon selection