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Photo Lynn Kamerlin

Lynn Kamerlin

Professor

Photo Lynn Kamerlin

Enzyme Architecture : Modeling the Operation of a Hydrophobic Clamp in Catalysis by Triosephosphate Isomerase

Author

  • Yashraj S Kulkarni
  • Qinghua Liao
  • Dušan Petrović
  • Dennis M Krüger
  • Birgit Strodel
  • Tina L Amyes
  • John P Richard
  • Shina C L Kamerlin

Summary, in English

Triosephosphate isomerase (TIM) is a proficient catalyst of the reversible isomerization of dihydroxyacetone phosphate (DHAP) to d-glyceraldehyde phosphate (GAP), via general base catalysis by E165. Historically, this enzyme has been an extremely important model system for understanding the fundamentals of biological catalysis. TIM is activated through an energetically demanding conformational change, which helps position the side chains of two key hydrophobic residues (I170 and L230), over the carboxylate side chain of E165. This is critical both for creating a hydrophobic pocket for the catalytic base and for maintaining correct active site architecture. Truncation of these residues to alanine causes significant falloffs in TIM's catalytic activity, but experiments have failed to provide a full description of the action of this clamp in promoting substrate deprotonation. We perform here detailed empirical valence bond calculations of the TIM-catalyzed deprotonation of DHAP and GAP by both wild-type TIM and its I170A, L230A, and I170A/L230A mutants, obtaining exceptional quantitative agreement with experiment. Our calculations provide a linear free energy relationship, with slope 0.8, between the activation barriers and Gibbs free energies for these TIM-catalyzed reactions. We conclude that these clamping side chains minimize the Gibbs free energy for substrate deprotonation, and that the effects on reaction driving force are largely expressed at the transition state for proton transfer. Our combined analysis of previous experimental and current computational results allows us to provide an overview of the breakdown of ground-state and transition state effects in enzyme catalysis in unprecedented detail, providing a molecular description of the operation of a hydrophobic clamp in triosephosphate isomerase.

Publishing year

2017-08-02

Language

English

Pages

10514-10525

Publication/Series

Journal of the American Chemical Society

Volume

139

Issue

30

Document type

Journal article

Publisher

The American Chemical Society (ACS)

Keywords

  • Biocatalysis
  • Dihydroxyacetone Phosphate/chemistry
  • Glyceraldehyde 3-Phosphate/chemistry
  • Hydrophobic and Hydrophilic Interactions
  • Molecular Conformation
  • Molecular Dynamics Simulation
  • Saccharomyces cerevisiae/enzymology
  • Thermodynamics
  • Triose-Phosphate Isomerase/chemistry

Status

Published

ISBN/ISSN/Other

  • ISSN: 1520-5126