{"doi":"10.1101/2020.07.24.215095","title":"Effects of individual base-pairs on <i>in vivo</i> target search and destruction kinetics of small RNA","abstract":"Abstract Base-pairing interactions mediate intermolecular target recognition in many biological systems and applications, including DNA repair, CRISPR, microRNA, small RNA (sRNA) and antisense oligo therapies. Even a single base-pair mismatch can cause a substantial difference in biological activity but presently we do not yet know how the target search kinetics in vivo are influenced by single nucleotide level changes. Here, we used high-throughput sequencing to identify functionally relevant single point mutants of the bacterial sRNA, SgrS, and quantitative super-resolution microscopy to probe the mutational impact on the regulation of its primary target, ptsG mRNA. Our super-resolution imaging and analysis platform allowed us to further dissect mutational effects on SgrS lifetimes, and even subtle changes in the in vivo rates of target association, k on , and dissociation, k off . Mutations that disrupt binding of a chaperone protein, Hfq, and are distal to the mRNA annealing region still decreased k on and increased k off , providing an in vivo demonstration that Hfq directly facilitates sRNA-mRNA annealing. Single base-pair mismatches in the annealing region reduced k on by 24-31% and increased k off by 14-25%, extending the time it takes to find and destroy the target mRNA by about a third, depending on whether an AU or GC base-pair is disrupted. The effects of disrupting contiguous base-pairing are much more modest than that expected from thermodynamics, suggesting that Hfq also buffers base-pair disruptions.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2020,"id":130474,"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":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.944,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2020-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":530828,"name":"Muhammad S. Azam","orcid":"0000-0003-2312-6573","position":1,"is_corresponding":false},{"id":48090,"name":"Tunc Kayikcioglu","orcid":"0000-0003-2205-8062","position":2,"is_corresponding":false},{"id":583162,"name":"Maksym Bobrovskyy","orcid":"0000-0002-2541-5451","position":3,"is_corresponding":false},{"id":583163,"name":"Jichuan Zhang","orcid":"0000-0003-1966-9436","position":4,"is_corresponding":false},{"id":583774,"name":"Xiangqian Ma","orcid":null,"position":5,"is_corresponding":false},{"id":583164,"name":"Piyush Labhsetwar","orcid":"0000-0001-5933-3609","position":6,"is_corresponding":false},{"id":535457,"name":"Jingyi Fei","orcid":"0000-0002-9775-3820","position":7,"is_corresponding":false},{"id":467230,"name":"Digvijay Singh","orcid":"0000-0002-7411-4436","position":8,"is_corresponding":false},{"id":254091,"name":"Zaida Luthey‐Schulten","orcid":"0000-0001-9749-8367","position":9,"is_corresponding":false},{"id":530829,"name":"Carin K. Vanderpool","orcid":"0000-0002-2850-5532","position":10,"is_corresponding":false},{"id":22168,"name":"Taekjip Ha","orcid":"0000-0003-2195-6258","position":11,"is_corresponding":false},{"id":530827,"name":"Anustup Poddar","orcid":"0000-0001-6792-2956","position":0,"is_corresponding":true}],"reference_count":78,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-18T23:15:56.698770Z","pmid":null,"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":[]}