9C6K

Cryo-EM structure of the TPP riboswitch embedded in an RNA scaffold bound to thiamine pyrophosphate

Summary for 9C6K

• Entry DOI: 10.2210/pdb9c6k/pdb
• Related: 9C6I 9C6J 9C6K
• EMDB information: 45247 45248 45249 45250 45251
• Descriptor: TPP riboswitch embedded in an RNA scaffold bound to thiamine pyrophosphate, MAGNESIUM ION, THIAMINE DIPHOSPHATE, ... (4 entities in total)
• Functional Keywords: rna, riboswitch, ligand
• Biological source: Escherichia coli
• Total number of polymer chains: 1
• Total formula weight: 161719.61

Authors

Toor, N. (deposition date: 2024-06-07, release date: 2024-12-04, Last modification date: 2025-02-05)

Primary citation

Haack, D.B.,Rudolfs, B.,Jin, S.,Khitun, A.,Weeks, K.M.,Toor, N.
Scaffold-enabled high-resolution cryo-EM structure determination of RNA.
Nat Commun, 16:880-880, 2025

PubMed Abstract: Cryo-EM structure determination of protein-free RNAs has remained difficult with most attempts yielding low to moderate resolution and lacking nucleotide-level detail. These difficulties are compounded for small RNAs as cryo-EM is inherently more difficult for lower molecular weight macromolecules. Here we present a strategy for fusing small RNAs to a group II intron that yields high resolution structures of the appended RNA. We demonstrate this technology by determining the structures of the 86-nucleotide (nt) thiamine pyrophosphate (TPP) riboswitch aptamer domain and the recently described 210-nt raiA bacterial non-coding RNA involved in sporulation and biofilm formation. In the case of the TPP riboswitch aptamer domain, the scaffolding approach allowed visualization of the riboswitch ligand binding pocket at 2.5 Å resolution. We also determined the structure of the ligand-free apo state and observe that the aptamer domain of the riboswitch adopts an open Y-shaped conformation in the absence of ligand. Using this scaffold approach, we determined the structure of raiA at 2.5 Å in the core. Our versatile scaffolding strategy enables efficient RNA structure determination for a broad range of small to moderate-sized RNAs, which were previously intractable for high-resolution cryo-EM studies.

PubMed: 39837824
DOI: 10.1038/s41467-024-55699-5

Experimental method

ELECTRON MICROSCOPY (2.94 Å)