Destabilization of the P site codon-anticodon helix results from movement of tRNA into the P/E hybrid state within the ribosome.
Retention of the reading frame in ribosomal complexes after single-round translocation depends on the acylation state of the tRNA. When tRNA lacking a peptidyl group is translocated to the P site, the mRNA slips to allow re-pairing of the tRNA with a nearby out-of-frame codon. Here, we show that this ribosomal activity results from movement of tRNA into the P/E hybrid state. Slippage of mRNA is suppressed by 3' truncation of the translocated tRNA, increased MgCl2 concentration, and mutation C2394A of the 50S E site, and each of these conditions inhibits P/E-state formation. Mutation G2252U of the 50S P site stimulates mRNA slippage, suggesting that decreased affinity of tRNA for the P/P state also destabilizes mRNA in the complex. The effects of G2252U are suppressed by C2394A, further implicating the P/E state in mRNA destabilization. This work uncovers a functional attribute of the P/E state crucial for understanding translation.
McGarry
Kevin G
KG
Department of Microbiology, The Ohio State University, Columbus, Ohio 43210, USA.
Walker
Sarah E
SE
Wang
Huanyu
H
Fredrick
Kurt
K
D000926:Anticodon / Q000378:metabolism*; D001665:Binding Sites; D003062:Codon / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D015636:Magnesium Chloride; D009154:Mutation; D009690:Nucleic Acid Conformation; D010441:Peptide Chain Elongation, Translational; D010455:Peptides / Q000235:genetics / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000502:physiology; D021381:Protein Transport / Q000235:genetics; D020871:RNA Stability / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D012335:RNA, Ribosomal / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012269:Ribosomal Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism*; D046249:Transfer RNA Aminoacylation / Q000502:physiology
D000926:Anticodon; D003062:Codon; D029968:Escherichia coli Proteins; D010455:Peptides; D012333:RNA, Messenger; D012335:RNA, Ribosomal; D012346:RNA, Transfer, Amino Acyl; D012269:Ribosomal Proteins; D015636:Magnesium Chloride; D012343:RNA, Transfer