The Dilemma of Conformational Dynamics in Enzyme Catalysis: Perspectives from Theory and Experiment

2013 ◽  
pp. 221-243 ◽  
Author(s):  
Urmi Doshi ◽  
Donald Hamelberg
Keyword(s):  
Enzyme Catalysis ◽  
Conformational Dynamics ◽  
Theory And Experiment

scholarly journals Rescue of conformational dynamics in enzyme catalysis by directed evolution

2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Renee Otten ◽  
Lin Liu ◽  
Lillian R. Kenner ◽  
Michael W. Clarkson ◽  
David Mavor ◽  
...  
Keyword(s):  
Directed Evolution ◽  
Enzyme Catalysis ◽  
Conformational Dynamics

scholarly journals Mix-and-inject XFEL crystallography reveals gated conformational dynamics during enzyme catalysis

2019 ◽  
Author(s):  
Medhanjali Dasgupta ◽  
Dominik Budday ◽  
Saulo H.P. de Oliveira ◽  
Peter Madzelan ◽  
Darya Marchany-Rivera ◽  
...  
Keyword(s):  
Ambient Temperature ◽  
Computer Simulations ◽  
Active Site ◽  
Enzyme Catalysis ◽  
Conformational Dynamics ◽  
Covalent Modification ◽  
Enzyme Structure ◽  
Conformational Ensemble ◽  
Cysteine Modification ◽  
X Ray

Summary ParagraphProtein dynamics play an important role in enzyme catalysis1-4. Many enzymes form covalent catalytic intermediates that can alter enzyme structure and conformational dynamics5,6. How these changes in enzyme structure and dynamics facilitate passage along the reaction coordinate is a fundamental unanswered question in structural enzymology. Here, we use Mix-and-Inject Serial Femtosecond X-ray Crystallography (MISC) at an X-ray Free Electron Laser (XFEL)7-10, ambient temperature X-ray crystallography, computer simulations, and enzyme kinetics to characterize how covalent modification of the active site cysteine residue in isocyanide hydratase (ICH) alters the enzyme’s conformational ensemble throughout the catalytic cycle. With MISC, we directly observe formation of a thioimidate covalent intermediate during ICH catalysis. The intermediate exhibits changes in the active site electrostatic environment, disrupting a hydrogen bond and triggering a cascade of conformational changes in ICH. X-ray-induced formation of a cysteine-sulfenic acid at the catalytic nucleophile (Cys101-SOH) with conventional crystallography at ambient temperature induces similar conformational shifts, demonstrating that these enzyme motions result from cysteine modification. Computer simulations show how cysteine modification-gated structural changes allosterically propagate through the ICH dimer. Mutations at Gly150 that modulate helical mobility reduce ICH catalytic turnover and alter its pre-steady state kinetic behavior, establishing that helical mobility is important for ICH catalytic efficiency. Taken together, our results demonstrate the potential of mix-and-inject XFEL crystallography to capture otherwise elusive mechanistic details of enzyme catalysis and dynamics from microcrystalline samples7,11. This approach can connect conformational dynamics to function for the large class of systems that rely on covalently modified cysteine residues for catalysis or regulation, resolving long-standing questions about enzyme mechanism and functionally relevant non-equilibrium enzyme motions.

scholarly journals Are Conformational Dynamics of Enzymes Important for Enzyme Catalysis?

2019 ◽  
Vol 59 (5) ◽  
pp. 271-272
Author(s):  
Toshifumi MORI
Keyword(s):  
Enzyme Catalysis ◽  
Conformational Dynamics

scholarly journals The role of ligand-gated conformational changes in enzyme catalysis

2019 ◽  
Vol 47 (5) ◽  
pp. 1449-1460 ◽  
Author(s):  
Cátia Moreira ◽  
Ana Rita Calixto ◽  
John P. Richard ◽  
Shina Caroline Lynn Kamerlin
Keyword(s):  
Binding Energy ◽  
Conformational Changes ◽  
Enzyme Catalysis ◽  
Phosphoryl Group ◽  
Computational Studies ◽  
Loop Dynamics ◽  
Biochemical Studies ◽  
The Common ◽  
Theory And Experiment

Abstract Structural and biochemical studies on diverse enzymes have highlighted the importance of ligand-gated conformational changes in enzyme catalysis, where the intrinsic binding energy of the common phosphoryl group of their substrates is used to drive energetically unfavorable conformational changes in catalytic loops, from inactive open to catalytically competent closed conformations. However, computational studies have historically been unable to capture the activating role of these conformational changes. Here, we discuss recent experimental and computational studies, which can remarkably pinpoint the role of ligand-gated conformational changes in enzyme catalysis, even when not modeling the loop dynamics explicitly. Finally, through our joint analyses of these data, we demonstrate how the synergy between theory and experiment is crucial for furthering our understanding of enzyme catalysis.

scholarly journals Minimization of dynamic effects in the evolution of dihydrofolate reductase

2016 ◽  
Vol 7 (5) ◽  
pp. 3248-3255 ◽  
Author(s):  
J. Javier Ruiz-Pernía ◽  
Enas Behiry ◽  
Louis Y. P. Luk ◽  
E. Joel Loveridge ◽  
Iñaki Tuñón ◽  
...  
Keyword(s):  
Dihydrofolate Reductase ◽  
Enzyme Catalysis ◽  
Isotope Labeling ◽  
Conformational Dynamics ◽  
Dynamic Effects ◽  
Powerful Technique ◽  
P Protein

Protein isotope labeling is a powerful technique to probe functionally important motions in enzyme catalysis and can be applied to investigate the conformational dynamics of proteins.

scholarly journals Mix-and-inject XFEL crystallography reveals gated conformational dynamics during enzyme catalysis

2019 ◽  
Vol 116 (51) ◽  
pp. 25634-25640 ◽  
Author(s):  
Medhanjali Dasgupta ◽  
Dominik Budday ◽  
Saulo H. P. de Oliveira ◽  
Peter Madzelan ◽  
Darya Marchany-Rivera ◽  
...  
Keyword(s):  
Active Site ◽  
Enzyme Catalysis ◽  
Conformational Dynamics ◽  
Catalytic Efficiency ◽  
Enzyme Mechanism ◽  
Time Resolved ◽  
X Ray ◽  
Protein Motions ◽  
X Ray Crystallography ◽  
Active Site Cysteine

How changes in enzyme structure and dynamics facilitate passage along the reaction coordinate is a fundamental unanswered question. Here, we use time-resolved mix-and-inject serial crystallography (MISC) at an X-ray free electron laser (XFEL), ambient-temperature X-ray crystallography, computer simulations, and enzyme kinetics to characterize how covalent catalysis modulates isocyanide hydratase (ICH) conformational dynamics throughout its catalytic cycle. We visualize this previously hypothetical reaction mechanism, directly observing formation of a thioimidate covalent intermediate in ICH microcrystals during catalysis. ICH exhibits a concerted helical displacement upon active-site cysteine modification that is gated by changes in hydrogen bond strength between the cysteine thiolate and the backbone amide of the highly strained Ile152 residue. These catalysis-activated motions permit water entry into the ICH active site for intermediate hydrolysis. Mutations at a Gly residue (Gly150) that modulate helical mobility reduce ICH catalytic turnover and alter its pre-steady-state kinetic behavior, establishing that helical mobility is important for ICH catalytic efficiency. These results demonstrate that MISC can capture otherwise elusive aspects of enzyme mechanism and dynamics in microcrystalline samples, resolving long-standing questions about the connection between nonequilibrium protein motions and enzyme catalysis.

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