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Updated: Sep 15, 2025

Single Molecule Fluorescence Energy Transfer Study of Ribosome Protein Synthesis
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Observing intersubunit dynamics in single yeast ribosomes.

Ananya Das1, Aleksandr V Ivanov1, Hironao Wakabayashi1

  • 1Department of Biochemistry & Biophysics, School of Medicine and Dentistry, and Center for RNA Biology, University of Rochester, Rochester, NY 14642.

Biorxiv : the Preprint Server for Biology
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PubMed
Summary
This summary is machine-generated.

Yeast ribosomes rotate between nonrotated and rotated states during translation. This intersubunit rotation is coupled to tRNA movement and elongation factors, offering insights into translation regulation.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Structural Biology

Background:

  • Ribosomes translate genetic information into proteins.
  • Ribosomal subunit rotation is crucial for translation.
  • Eukaryotic translation involves complex dynamics and regulatory factors.

Purpose of the Study:

  • To investigate the intersubunit dynamics of Saccharomyces cerevisiae ribosomes during translation.
  • To elucidate the role of elongation factors in ribosome conformational changes.
  • To establish a method for studying eukaryotic translation regulation.

Main Methods:

  • Single-molecule Förster resonance energy transfer (smFRET) assay.
  • Labeling of ribosomal proteins uS15 and eL30 with fluorophores.
  • Monitoring FRET states corresponding to nonrotated (NR) and rotated (R) conformations.

Main Results:

  • Yeast ribosomes exhibit two predominant FRET states (NR and R), similar to bacterial ribosomes.
  • Intersubunit rotation is coupled to tRNA transitions between classical and hybrid binding states.
  • The elongation cycle involves sequential transitions between NR and R states.
  • Elongation factor 3 (eEF3) facilitates the late step of translocation, following eEF2-induced reverse rotation.

Conclusions:

  • The study provides evidence for coupled intersubunit rotation and tRNA dynamics in yeast translation.
  • The developed smFRET assay is a valuable tool for studying eukaryotic translation regulation mechanisms.
  • Understanding these dynamics is key to investigating ribosome pausing, stalling, and frameshifting.