Monitoring Real-Time Temperature Dynamics of a Short RNA Hairpin Using Förster Resonance Energy Transfer and Circular Dichroism.
Circular dichroism
FRET
Hairpins
RNA structures
RNA thermometer
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2024
2024
Historique:
medline:
10
4
2024
pubmed:
10
4
2024
entrez:
9
4
2024
Statut:
ppublish
Résumé
RNA molecules play crucial roles in gene expression regulation and cellular signaling, and these functions are governed by the formation of RNA secondary and tertiary structures. These structures are highly dynamic and subject to rapid changes in response to environmental cues, temperature in particular. Thermosensitive RNA secondary structures have been harnessed by multiple organisms to survey their temperature environment and to adjust gene expression accordingly. It is thus highly desirable to observe RNA structural changes in real time over a range of temperatures. Multiple approaches have been developed to study structural dynamics, but many of these require extensive processing of the RNA, large amounts of RNA input, and/or cannot be applied under physiological conditions. Here, we describe the use of a dually fluorescently labeled RNA oligonucleotide (containing a predicted hairpin structure) to monitor subtle RNA structural dynamics in vitro by Förster resonance energy transfer (FRET) and circular dichroism (CD) spectroscopy. These approaches can be employed under physiologically relevant conditions over a range of temperatures and with RNA concentrations as low as 200 nM; they enable us to observe RNA structural dynamics in real time and to correlate these dynamics with changes in biological processes such as translation.
Identifiants
pubmed: 38594536
doi: 10.1007/978-1-0716-3814-9_15
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
149-158Informations de copyright
© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
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