{"doi":"10.1101/2024.06.10.598011","title":"Scaffold-enabled high-resolution cryo-EM structure determination of RNA","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, which we demonstrate with the 86-nucleotide thiamine pyrophosphate (TPP) riboswitch, and visualizing 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 undergoes a large-scale conformational change upon ligand binding, illustrating how small molecule binding to an RNA can induce large effects on gene expression. This study both sets a new standard for cryo-EM riboswitch visualization and offers a versatile strategy applicable to a broad range of small to moderate-sized RNAs, which were previously intractable for high-resolution cryo-EM studies.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2024,"id":485588,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":5,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9546,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1074938,"name":"Boris Rudolfs","orcid":"0000-0002-8791-9487","position":1,"is_corresponding":false},{"id":1329102,"name":"Shouhong Jin","orcid":null,"position":2,"is_corresponding":false},{"id":5954,"name":"Kevin M. Weeks","orcid":"0000-0002-6748-9985","position":3,"is_corresponding":false},{"id":710523,"name":"Navtej Toor","orcid":"0000-0002-6134-163X","position":4,"is_corresponding":false},{"id":1074937,"name":"Daniel B. Haack","orcid":"0000-0002-4926-7772","position":0,"is_corresponding":true}],"reference_count":49,"raw_metadata":null,"created_at":"2026-07-19T02:07:52.536246Z","pmid":"38915706","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}