Changes in an enzyme ensemble during catalysis observed by high-resolution XFEL crystallography-Reference-Cited by-同舟云学术

Changes in an enzyme ensemble during catalysis observed by high-resolution XFEL crystallography

Published:2024-03-29 Issue:13 Volume:10 Page:
ISSN:2375-2548
Container-title:Science Advances
language:en
Short-container-title:Sci. Adv.

Author:

Smith Nathan¹^ORCID,Dasgupta Medhanjali¹,Wych David C.²³,Dolamore Cole¹,Sierra Raymond G.⁴^ORCID,Lisova Stella⁴,Marchany-Rivera Darya⁵^ORCID,Cohen Aina E.⁵^ORCID,Boutet Sébastien⁴^ORCID,Hunter Mark S.⁴,Kupitz Christopher⁴^ORCID,Poitevin Frédéric⁴^ORCID,Moss Frank R.⁴^ORCID,Mittan-Moreau David W.⁶^ORCID,Brewster Aaron S.⁶^ORCID,Sauter Nicholas K.⁶^ORCID,Young Iris D.⁶^ORCID,Wolff Alexander M.⁷^ORCID,Tiwari Virendra K.⁸,Kumar Nivesh⁸,Berkowitz David B.⁸,Hadt Ryan G.⁹,Thompson Michael C.⁷^ORCID,Follmer Alec H.¹⁰^ORCID,Wall Michael E.²^ORCID,Wilson Mark A.¹^ORCID

Affiliation:

1. Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.

2. Computer, Computational, and Statistical Sciences Division, Los Alamos National Laboratory, Los Alamos, NM 875405, USA.

3. Center for Nonlinear Studies, Los Alamos National Laboratory, Los Alamos, NM 87545, USA.

4. Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.

5. Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.

6. Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.

7. Department of Chemistry and Biochemistry, University of California, Merced, CA 95340, USA.

8. Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.

9. Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.

10. Department of Chemistry, University of California-Irvine, Irvine, CA 92697, USA.

Abstract

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.

Publisher

American Association for the Advancement of Science (AAAS)

Link

https://www.science.org/doi/pdf/10.1126/sciadv.adk7201

Reference85 articles.

1. Taking Ockham's razor to enzyme dynamics and catalysis

2. Intrinsic dynamics of an enzyme underlies catalysis

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