{"doi":"10.1101/2021.09.29.461692","title":"Ensemble–function relationships to evaluate catalysis in the ketosteroid isomerase oxyanion hole","abstract":"ABSTRACT Following decades of insights from structure–function studies, there is now a need to progress from a static to dynamic view of enzymes. Comparison of prior cryo X-ray structures suggested that deleterious effects from ketosteroid isomerase (KSI) mutants arise from misalignment of the oxyanion hole catalytic residue, Y16. However, multi-conformer models from room temperature X-ray diffraction revealed an ensemble of Y16 conformers indistinguishable from WT for Y32F/Y57F KSI and a distinct, non-native ensemble for Y16 in Y57F KSI. Functional analyses suggested rate effects arise from weakened hydrogen bonding, due to disruption of the Y16/Y57/Y32 hydrogen bond network, and repositioning of the general base. In general, catalytic changes can be deconvoluted into effects on the probability of occupying a state ( P -effects) and the reactivity of each state ( k -effects). Our results underscore the need for ensemble–function analysis to decipher enzyme function and ultimately manipulate their extraordinary capabilities.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2021,"id":227031,"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":0.9533,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2021-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":364771,"name":"Tzanko Doukov","orcid":"0000-0001-8625-2572","position":1,"is_corresponding":false},{"id":630198,"name":"Margaux M. Pinney","orcid":"0000-0002-7735-5032","position":2,"is_corresponding":false},{"id":108140,"name":"J.T. Biel","orcid":"0000-0002-0935-8362","position":3,"is_corresponding":false},{"id":63530,"name":"James S. Fraser","orcid":"0000-0002-5080-2859","position":4,"is_corresponding":false},{"id":226045,"name":"Daniel Herschlag","orcid":"0000-0002-4685-1973","position":5,"is_corresponding":false},{"id":479336,"name":"Filip Yabukarski","orcid":"0000-0002-7207-0951","position":0,"is_corresponding":true}],"reference_count":82,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-18T23:54:38.004707Z","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":[]}