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Natl. Acad. Sci. U.S.A."],"published-print":{"date-parts":[[2026,10,20]]},"abstract":"<jats:p>\n                    Capturing enzymes under native turnover conditions remains a grand challenge in structural biology. Here, we develop a reaction-based cryo\u2013electron microscopy (cryo-EM) strategy that directly samples\n                    <jats:italic toggle=\"yes\">Drosophila melanogaster<\/jats:italic>\n                    cytidine triphosphate synthase (CTPS) from actively catalyzing mixtures containing only natural substrates and allosteric effectors. By integrating reaction-based sampling with three-dimensional variability analysis, we resolve a continuous conformational spectrum of CTPS during catalysis. This approach visualizes the chemical progression of 4-phosphoryl-uridine triphosphate (4Pi-UTP) formation and reveals a previously unobserved co-occupancy pattern of adenosine triphosphate (ATP) and CTP at the reaction end point. Cross-validation with nonhydrolyzable ATP analogs demonstrates that UTP phosphorylation by ATP, rather than mere ATP binding, shifts the conformational ensemble toward more closed states. Our findings establish reaction-based cryo-EM as a framework for resolving chemically annotated conformational ensembles without predefining a single trapped intermediate.\n                  <\/jats:p>","DOI":"10.1073\/pnas.2620271123","type":"journal-article","created":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T16:32:35Z","timestamp":1791563555000},"update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":0,"title":["Reaction-based cryo-EM resolves the continuous conformational spectrum of CTP synthase catalysis"],"prefix":"10.1073","volume":"123","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5342-4761","authenticated-orcid":false,"given":"Chen-Jun","family":"Guo","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/030bhh786","id-type":"ROR","asserted-by":"publisher"}],"name":"School of Life Science and Technology, ShanghaiTech University","place":["Shanghai, 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