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Related Experiment Video

Updated: Apr 15, 2026

Genetic and Biochemical Approaches for In Vivo and In Vitro Assessment of Protein Oligomerization: The Ryanodine Receptor Case Study
12:43

Genetic and Biochemical Approaches for In Vivo and In Vitro Assessment of Protein Oligomerization: The Ryanodine Receptor Case Study

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In vivo protein cross-linking.

Fabrice Agou1, Michel Véron

  • 1Département de Biologie Cellulaire et Infection, Institut Pasteur, Unité de Signalisation et Pathogenèse, 25 rue du Docteur Roux, F-75015, Paris, France, fagou@pasteur.fr.

Methods in Molecular Biology (Clifton, N.J.)
|April 11, 2015
PubMed
Summary

This study guides selecting cell-permeable cross-linkers for in vivo protein interaction studies. Researchers can selectively identify protein dimers or hexamers using specific cross-linking strategies.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Cell Biology

Background:

  • Cellular compartments are crowded with macromolecules, influencing protein associations.
  • In vivo chemical cross-linking is vital for studying protein oligomerization and interactions during cellular events like signal transduction.

Purpose of the Study:

  • To provide a guide for selecting cell membrane-permeable cross-linkers.
  • To optimize in vivo cross-linking conditions for identifying specific protein cross-links.
  • To demonstrate selective cross-linking of specific protein oligomeric states.

Main Methods:

  • Selection of homo-bifunctional cross-linkers with specific spacer arm lengths.
  • Optimization of in vivo cross-linking conditions in a crowded cellular environment.

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  • Utilizing chemoselectivity and protein structure knowledge for targeted cross-linking.

    • Main Results:

    • Demonstrated selective cross-linking of specific oligomeric forms (dimer or hexamer) of a polypeptide.
    • Successfully applied in vitro and in vivo cross-linking techniques using human type B nucleoside diphosphate kinase as a model.
    • Showcased the ability to introduce cross-links specifically on either the dimer or hexamer form of the same polypeptide.

    Conclusions:

    • Selective in vivo protein cross-linking is achievable by carefully choosing cross-linkers and conditions.
    • This approach allows for the specific identification of protein oligomeric states within a complex cellular milieu.
    • The study provides a valuable methodology for investigating protein-protein interactions and their dynamics.