Observation of SAM-VI Riboswitch Dynamics Using Single-Molecule FRET

Observation of SAM-VI Riboswitch Dynamics Using Single-Molecule FRET

Riboswitches regulate gene expression through intricate dynamic conformational transitions, with divalent cation Mg<sup>2+</sup> and their ligands playing pivotal roles in this process. The dynamic structural mechanism by which the S-adenosyl-L-methionine (SAM) responsive SAM-VI riboswit...

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Bibliographic Details
Main Authors: Yanyan Xue, Yi Sun, Yichun Xia, Xiuming Liu, Hua Dai
Format: Article
Language:English
Published: MDPI AG 2025-06-01
Series:Biomolecules
Subjects:
SAM-VI riboswitch
dynamic structure
PLOR
fluorescent labeling
smFRET
Online Access:https://www.mdpi.com/2218-273X/15/6/841
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Summary:Riboswitches regulate gene expression through intricate dynamic conformational transitions, with divalent cation Mg<sup>2+</sup> and their ligands playing pivotal roles in this process. The dynamic structural mechanism by which the S-adenosyl-L-methionine (SAM) responsive SAM-VI riboswitch (riboSAM) regulates the downstream SAM synthase gene translation remains unclear. In this study, we employed position-selective labeling of RNA (PLOR) to incorporate Cy3-Cy5 into designated positions of riboSAM, applying single-molecule Förster resonance energy transfer (smFRET) method to track its conformational switches in response to Mg<sup>2+</sup> and SAM. smFRET analysis revealed that in the absence of Mg<sup>2+</sup> and ligand, riboSAM predominantly adopted a translation-activating <i>apo</i> conformation. Physiological concentrations of Mg<sup>2+</sup> induced riboSAM to fold into dynamic <i>transit-p</i> and <i>holo-p</i> states, creating a transient and structurally pliable binding pocket for ligand binding. SAM binding locks the dynamic <i>transit-p</i> and <i>holo-p</i> states into their final stable <i>transit</i> and <i>holo</i> conformations through conformational selection, turning off downstream cis-gene expression and completing feedback regulation of cellular SAM concentration. The observed synergistic regulatory effect of Mg<sup>2+</sup> ions and ligand on riboSAM’s conformational dynamics at single-molecule resolution provides new mechanistic insights into gene regulation by diverse riboswitch classes.
ISSN:2218-273X

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