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Publication Abstract from PubMed

Enzymes populate ensembles of structures necessary for catalysis that are difficult to experimentally characterize. We use time-resolved mix-and-inject serial crystallography at an x-ray free electron laser to observe catalysis in a designed mutant isocyanide hydratase (ICH) enzyme that enhances sampling of important minor conformations. The active site exists in a mixture of conformations, and formation of the thioimidate intermediate selects for catalytically competent substates. The influence of cysteine ionization on the ICH ensemble is validated by determining structures of the enzyme at multiple pH values. Large molecular dynamics simulations in crystallo and time-resolved electron density maps show that Asp(17) ionizes during catalysis and causes conformational changes that propagate across the dimer, permitting water to enter the active site for intermediate hydrolysis. ICH exhibits a tight coupling between ionization of active site residues and catalysis-activated protein motions, exemplifying a mechanism of electrostatic control of enzyme dynamics.

Changes in an enzyme ensemble during catalysis observed by high-resolution XFEL crystallography.,Smith N, Dasgupta M, Wych DC, Dolamore C, Sierra RG, Lisova S, Marchany-Rivera D, Cohen AE, Boutet S, Hunter MS, Kupitz C, Poitevin F, Moss FR 3rd, Mittan-Moreau DW, Brewster AS, Sauter NK, Young ID, Wolff AM, Tiwari VK, Kumar N, Berkowitz DB, Hadt RG, Thompson MC, Follmer AH, Wall ME, Wilson MA Sci Adv. 2024 Mar 29;10(13):eadk7201. doi: 10.1126/sciadv.adk7201. Epub 2024 Mar , 27. PMID:38536910^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.