Techniques to Analyze sRNA Protein Cofactor Self-Assembly In Vitro
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.
KeywordsProtein self-assembly Noncoding RNA cofactor Functional amyloid
Atomic force microscopy
Fourier transform infrared spectroscopy
Small angle X-ray scattering
Synchrotron radiation circular dichroism
Transmission electron microscopy
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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