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Changes in an enzyme ensemble during catalysis observed by high-resolution XFEL crystallography.
Smith, Nathan; Dasgupta, Medhanjali; Wych, David C; Dolamore, Cole; Sierra, Raymond G; Lisova, Stella; Marchany-Rivera, Darya; Cohen, Aina E; Boutet, Sébastien; Hunter, Mark S; Kupitz, Christopher; Poitevin, Frédéric; Moss, Frank R; Mittan-Moreau, David W; Brewster, Aaron S; Sauter, Nicholas K; Young, Iris D; Wolff, Alexander M; Tiwari, Virendra K; Kumar, Nivesh; Berkowitz, David B; Hadt, Ryan G; Thompson, Michael C; Follmer, Alec H; Wall, Michael E; Wilson, Mark A.
Afiliação
  • Smith N; Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
  • Dasgupta M; Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
  • Wych DC; Computer, Computational, and Statistical Sciences Division, Los Alamos National Laboratory, Los Alamos, NM 875405, USA.
  • Dolamore C; Center for Nonlinear Studies, Los Alamos National Laboratory, Los Alamos, NM 87545, USA.
  • Sierra RG; Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
  • Lisova S; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Marchany-Rivera D; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Cohen AE; Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Boutet S; Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Hunter MS; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Kupitz C; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Poitevin F; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Moss FR; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Mittan-Moreau DW; Linac Coherent Light Source, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025, USA.
  • Brewster AS; Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
  • Sauter NK; Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
  • Young ID; Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
  • Wolff AM; Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
  • Tiwari VK; Department of Chemistry and Biochemistry, University of California, Merced, CA 95340, USA.
  • Kumar N; Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
  • Berkowitz DB; Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
  • Hadt RG; Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE 68588, USA.
  • Thompson MC; Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
  • Follmer AH; Department of Chemistry and Biochemistry, University of California, Merced, CA 95340, USA.
  • Wall ME; Department of Chemistry, University of California-Irvine, Irvine, CA 92697, USA.
  • Wilson MA; Computer, Computational, and Statistical Sciences Division, Los Alamos National Laboratory, Los Alamos, NM 875405, USA.
Sci Adv ; 10(13): eadk7201, 2024 Mar 29.
Article em En | MEDLINE | ID: mdl-38536910
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 Asp17 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.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Proteínas / Simulação de Dinâmica Molecular Idioma: En Revista: Sci Adv Ano de publicação: 2024 Tipo de documento: Article País de afiliação: Estados Unidos
Texto completo
Imprimir
XML
PubMed Links
Buscar no Google

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Proteínas / Simulação de Dinâmica Molecular Idioma: En Revista: Sci Adv Ano de publicação: 2024 Tipo de documento: Article País de afiliação: Estados Unidos