Probing Transient Riboswitch Structures via Single Molecule Accessibility Analysis.
Bacterial gene regulation
Conformational dynamics
RNA folding
Riboswitch
Single molecule fluorescence microscopy
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:
2023
2023
Historique:
entrez:
13
10
2022
pubmed:
14
10
2022
medline:
18
10
2022
Statut:
ppublish
Résumé
Riboswitches are a class of RNA motifs in the untranslated regions of bacterial messenger RNAs (mRNAs) that can adopt different conformations to regulate gene expression. The binding of specific small molecule or ion ligands, or other RNAs, influences the conformation the riboswitch adopts. Single Molecule Kinetic Analysis of RNA Transient Structure (SiM-KARTS) offers an approach for probing this structural isomerization, or conformational switching, at the level of single mRNA molecules. SiM-KARTS utilizes fluorescently labeled, short, sequence-complementary DNA or RNA oligonucleotide probes that transiently access a specific RNA conformation over another. Binding and dissociation to a surface-immobilized target RNA of arbitrary length are monitored by Total Internal Reflection Fluorescence Microscopy (TIRFM) and quantitatively analyzed, via spike train and burst detection, to elucidate the rate constants of isomerization, revealing mechanistic insights into riboswitching.
Identifiants
pubmed: 36227561
doi: 10.1007/978-1-0716-2687-0_4
pmc: PMC10078578
mid: NIHMS1888024
doi:
Substances chimiques
DNA, Complementary
0
Ligands
0
Oligonucleotide Probes
0
RNA Probes
0
RNA, Bacterial
0
Riboswitch
0
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
37-51Subventions
Organisme : NIGMS NIH HHS
ID : R35 GM131922
Pays : United States
Informations de copyright
© 2023. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.
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