Nathan Smith, Medhanjali Dasgupta, David C Wych, Cole Dolamore, Raymond G Sierra, Stella Lisova, Darya Marchany-Rivera, Aina E Cohen, Sébastien Boutet, Mark S Hunter, Christopher Kupitz, Frédéric Poitevin, Frank R Moss, David W Mittan-Moreau, Aaron S Brewster, Nicholas K Sauter, Iris D Young, Alexander M Wolff, Virendra K Tiwari, Nivesh Kumar, David B Berkowitz, Ryan G Hadt, Michael C Thompson, Alec H Follmer, Michael E Wall, Mark A Wilson
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...
March 29, 2024: Science Advances