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Isolation of Translating Ribosomes Containing Peptidyl-tRNAs for Functional and Structural Analyses
Published on: February 25, 2011
A cryo-electron microscopic study of ribosome-bound termination factor RF2
Urmila B S Rawat1, Andrey V Zavialov, Jayati Sengupta
1Howard Hughes Medical Institute, Health Research, Inc., Empire State Plaza, Albany NY 12201-0509, USA.
Nature
|January 4, 2003
Summary
Release factor 2 (RF2) adopts an open conformation on the ribosome, enabling simultaneous interaction with stop codons and the peptidyl-transferase center for accurate protein synthesis termination.
Area of Science:
- Molecular Biology
- Structural Biology
- Genetics
Background:
- Protein synthesis terminates via class-1 release factors (RFs) recognizing stop codons.
- RFs possess conserved GGQ and SPF motifs critical for termination.
- The spatial arrangement of RFs on the ribosome has been a key question in understanding termination accuracy.
Purpose of the Study:
- To investigate the conformational state of RF2 when bound to the ribosome during protein synthesis termination.
- To elucidate how RF2 simultaneously interacts with the stop codon and the peptidyl-transferase center.
- To provide new insights into the accuracy of ribosomal protein synthesis termination.
Main Methods:
- X-ray crystallography of RF2 bound to the ribosome.
- Structural analysis of RF2 conformations.
- Biochemical assays to assess RF2 function.
Main Results:
- RF2 adopts an open conformation when bound to the ribosome.
- This open conformation allows the GGQ motif to reach the peptidyl-transferase center (PTC) while the SPF motif interacts with the stop codon in the decoding center (DC).
- The observed distance between motifs in the crystal structure contradicts simultaneous DC and PTC interaction in a closed conformation.
Conclusions:
- The open conformation of RF2 is crucial for simultaneous interaction with both the stop codon and the PTC.
- This mechanism explains how the ribosome signals stop codon presence to the PTC for accurate termination.
- Findings offer new interpretations of accuracy in protein synthesis termination.
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