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Techniques to Analyze sRNA Protein Cofactor Self-Assembly In Vitro

  • David Partouche
  • Antoine Malabirade
  • Thomas Bizien
  • Marisela Velez
  • Sylvain Trépout
  • Sergio Marco
  • Valeria Militello
  • Christophe Sandt
  • Frank Wien
  • Véronique Arluison
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1737)

Abstract

Post-transcriptional control of gene expression by small regulatory noncoding RNA (sRNA) needs protein accomplices to occur. Past research mainly focused on the RNA chaperone Hfq as cofactor. Nevertheless, recent studies indicated that other proteins might be involved in sRNA-based regulations. As some of these proteins have been shown to self-assemble, we describe in this chapter protocols to analyze the nano-assemblies formed. Precisely, we focus our analysis on Escherichia coli Hfq as a model, but the protocols presented here can be applied to analyze any polymer of proteins. This chapter thus provides a guideline to develop commonly used approaches to detect prokaryotic protein self-assembly, with a special focus on the detection of amyloidogenic polymers.

Keywords

Protein self-assembly Noncoding RNA cofactor Functional amyloid 

Abbreviations

AFM

Atomic force microscopy

CF

Curve fitting

FTIR

Fourier transform infrared spectroscopy

SAXS

Small angle X-ray scattering

SRCD

Synchrotron radiation circular dichroism

TEM

Transmission electron microscopy

ThT

Thioflavin T

Notes

Acknowledgments

This work was supported by Université Paris Diderot, CNRS, CEA, and Synchrotron SOLEIL. We gratefully acknowledge help to MV from the French Embassy for their program for scientific and university cooperation. We are indebted to F. Gobeaux (CEA Saclay, Gif-sur-Yvette, France) and A. Deniset-Besseau (LCP, Université Paris-Sud, France) for many fruitful discussions. We thank Kimberly Stanek (University of Virginia) for her careful and critical reading of our manuscript.

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Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  • David Partouche
    • 1
    • 2
  • Antoine Malabirade
    • 2
  • Thomas Bizien
    • 1
  • Marisela Velez
    • 3
  • Sylvain Trépout
    • 4
    • 5
  • Sergio Marco
    • 4
    • 5
  • Valeria Militello
    • 6
  • Christophe Sandt
    • 1
  • Frank Wien
    • 1
  • Véronique Arluison
    • 7
  1. 1.Synchrotron SOLEIL, L’Orme des Merisiers Saint AubinGif-sur-YvetteFrance
  2. 2.Université Paris Diderot-Paris 7, Sorbonne Paris CitéParisFrance
  3. 3.Instituto de Catálisis y Petroleoquímica, CSICMadridSpain
  4. 4.INSERM, U1196, Université Paris Sud, Université Paris-SaclayOrsayFrance
  5. 5.Institut Curie, PSL Research University, CNRS, UMR 91873348OrsayFrance
  6. 6.Department of Physics and ChemistryUniversity of PalermoPalermoItaly
  7. 7.Université Paris Diderot & LLB/CEA/CNRSGif-sur-YvetteFrance

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