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Related Concept Videos

Ribosome Profiling02:24

Ribosome Profiling

Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...

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Related Experiment Video

Updated: May 9, 2026

Quantitative Analysis of the Cellular Lipidome of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry
08:56

Quantitative Analysis of the Cellular Lipidome of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry

Published on: March 8, 2020

Offline Two-Stage SEC and LC-MS/MS for Comprehensive Characterization of Yeast Ribosomal Populations.

Andrew J Spiteri1,2, Joshua J Hamey1,2, Marc R Wilkins1,2

  • 1Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW 2052, Australia.

Journal of Proteome Research
|May 8, 2026
PubMed
Summary

This study introduces two-dimensional size exclusion chromatography tandem mass spectrometry (2D-SEC MS/MS) for rapid yeast ribosome analysis. The method quickly characterizes ribosomal particles and their protein composition, revealing insights into ribosome function and regulation.

Keywords:
Liquid chromatography−tandem mass spectrometry (LC−MS/MS)Ribosome heterogeneityRibosome protein paralogsSaccharomyces cerevisiaeSize exclusion chromatography (SEC)post-translational modifications

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Related Experiment Videos

Last Updated: May 9, 2026

Quantitative Analysis of the Cellular Lipidome of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry
08:56

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Quantitative Metabolomics of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry
07:25

Quantitative Metabolomics of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry

Published on: January 5, 2021

Area of Science:

  • Proteomics
  • Molecular Biology
  • Biochemistry

Background:

  • Ribosomes are essential cellular machinery responsible for protein synthesis.
  • Understanding ribosome composition and dynamics is crucial for deciphering cellular function and disease.
  • Current methods for ribosome characterization can be time-consuming and lack resolution.

Purpose of the Study:

  • To develop a rapid, high-resolution workflow for characterizing Saccharomyces cerevisiae ribosomes.
  • To enable detailed analysis of ribosomal subpopulations, protein composition, and post-translational modifications.
  • To link ribosome composition changes to functional perturbations under different cellular conditions.

Main Methods:

  • Coupling two size exclusion chromatography (SEC) steps with mass spectrometry (MS/MS) for a two-dimensional separation (2D-SEC MS/MS).
  • Utilizing MS-compatible SEC columns (1000 Å and 500 Å) to resolve intact polysomes, monosomes, and ribosomal subunits (80S, 60S, 40S).
  • Applying bottom-up liquid chromatography-tandem mass spectrometry (LC-MS/MS) to analyze protein content and modifications in separated fractions.

Main Results:

  • Successfully separated intact polysomes, 80S monosomes, and 60S/40S subunits into four distinct fractions in under 90 minutes.
  • Identified 78 out of 79 core yeast ribosomal proteins with high sequence coverage (93%) in the polysome fraction.
  • Resolved and quantified 36 out of 38 paralogous protein pairs, detecting isoform-specific incorporation biases.
  • Mapped all known yeast ribosomal methylation and canonical phosphorylation sites, enabling site-specific PTM measurements.
  • Detected rapid polysome collapse and altered profiling ratios under nutrient deprivation and RPL7A knockout conditions.

Conclusions:

  • 2D-SEC MS/MS provides a rapid and high-resolution method for comprehensive yeast ribosome characterization.
  • The workflow effectively links ribosome composition and post-translational modifications to functional states.
  • This technique offers a powerful tool for studying ribosome heterogeneity and its role in cellular processes and disease.