{"doi":"10.1101/2020.08.12.247874","title":"Quantitative yeast-yeast two hybrid for discovery and binding affinity estimation of protein-protein interactions","abstract":"Abstract Quantifying the binding affinity of protein-protein interactions is important for elucidating connections within biochemical signaling pathways, as well as characterization of binding proteins isolated from combinatorial libraries. We describe a quantitative yeast-yeast two hybrid (qYY2H) system that not only enables discovery of specific protein-protein interactions, but also efficient, quantitative estimation of their binding affinities ( K D ). In qYY2H, the bait and prey proteins are expressed as yeast cell surface fusions using yeast surface display. We developed a semi-empirical framework for estimating the K D of monovalent bait-prey interactions, using measurements of the apparent K D of yeast-yeast binding, which is mediated by multivalent interactions between yeast-displayed bait and prey. Using qYY2H, we identified interaction partners of SMAD3 and the tandem WW domains of YAP from a cDNA library and characterized their binding affinities. Finally, we showed that qYY2H could also quantitatively evaluate binding interactions mediated by post-translational modifications on the bait protein.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2020,"id":121699,"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":6,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9523,"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":562434,"name":"Abigail Blain","orcid":null,"position":1,"is_corresponding":false},{"id":444417,"name":"John Bowen","orcid":null,"position":2,"is_corresponding":false},{"id":562435,"name":"Matthew Burroughs","orcid":null,"position":3,"is_corresponding":false},{"id":561737,"name":"Nikki McArthur","orcid":"0000-0001-9872-8961","position":4,"is_corresponding":false},{"id":304474,"name":"Stefano Menegatti","orcid":"0000-0001-5633-434X","position":5,"is_corresponding":false},{"id":304473,"name":"Balaji M. Rao","orcid":"0000-0001-5695-8953","position":6,"is_corresponding":false},{"id":306383,"name":"Kaitlyn Bacon","orcid":null,"position":0,"is_corresponding":true}],"reference_count":63,"raw_metadata":null,"created_at":"2026-07-18T23:14:46.979435Z","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":[]}