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Using sm-FRET and denaturants to reveal folding landscapes

Research output: Contribution to journalReview articlepeer-review

Author(s)

Euan Stuart Shaw, Patrick St-Pierre, Kaley McCluskey, Daniel A Lafontaine, Carlos Penedo

School/Research organisations

Abstract

RNA folding studies aim to clarify the relationship among sequence, tridimensional structure, and biological function. In the last decade, the application of single-molecule fluorescence resonance energy transfer (sm-FRET) techniques to investigate RNA structure and folding has revealed the details of conformational changes and timescale of the process leading to the formation of biologically active RNA structures with subnanometer resolution on millisecond timescales. In this review, we initially summarize the first wave of single-molecule FRET-based RNA techniques that focused on analyzing the influence of mono- and divalent metal ions on RNA function, and how these studies have provided very valuable information about folding pathways and the presence of intermediate and low-populated states. Next, we describe a second generation of single-molecule techniques that combine sm-FRET with the use of chemical denaturants as an emerging powerful approach to reveal information about the dynamics and energetics of RNA folding that remains hidden using conventional sm-FRET approaches. The main advantages of using the competing interplay between folding agents such as metal ions and denaturants to observe and manipulate the dynamics of RNA folding and RNA–ligand interactions is discussed in the context of the adenine riboswitch aptamer.
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Details

Original languageEnglish
Pages (from-to)313-341
JournalMethods In Enzymology
Volume549
Early online date25 Nov 2014
DOIs
Publication statusPublished - 2014

    Research areas

  • Adenine aptamer, Chemical denaturation, Single-molecule FRET, RNA tertiary folding, Rate-limiting step

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