Cargando…
Enzyme Architecture: Modeling the Operation of a Hydrophobic Clamp in Catalysis by Triosephosphate Isomerase
[Image: see text] 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 understa...
Autores principales: | , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Chemical
Society
2017
|
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5543394/ https://www.ncbi.nlm.nih.gov/pubmed/28683550 http://dx.doi.org/10.1021/jacs.7b05576 |
_version_ | 1783255142949191680 |
---|---|
author | Kulkarni, Yashraj S. Liao, Qinghua Petrović, Dušan Krüger, Dennis M. Strodel, Birgit Amyes, Tina L. Richard, John P. Kamerlin, Shina C. L. |
author_facet | Kulkarni, Yashraj S. Liao, Qinghua Petrović, Dušan Krüger, Dennis M. Strodel, Birgit Amyes, Tina L. Richard, John P. Kamerlin, Shina C. L. |
author_sort | Kulkarni, Yashraj S. |
collection | PubMed |
description | [Image: see text] 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. |
format | Online Article Text |
id | pubmed-5543394 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | American Chemical
Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-55433942017-08-07 Enzyme Architecture: Modeling the Operation of a Hydrophobic Clamp in Catalysis by Triosephosphate Isomerase Kulkarni, Yashraj S. Liao, Qinghua Petrović, Dušan Krüger, Dennis M. Strodel, Birgit Amyes, Tina L. Richard, John P. Kamerlin, Shina C. L. J Am Chem Soc [Image: see text] 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. American Chemical Society 2017-07-07 2017-08-02 /pmc/articles/PMC5543394/ /pubmed/28683550 http://dx.doi.org/10.1021/jacs.7b05576 Text en Copyright © 2017 American Chemical Society This is an open access article published under a Creative Commons Non-Commercial No Derivative Works (CC-BY-NC-ND) Attribution License (http://pubs.acs.org/page/policy/authorchoice_ccbyncnd_termsofuse.html) , which permits copying and redistribution of the article, and creation of adaptations, all for non-commercial purposes. |
spellingShingle | Kulkarni, Yashraj S. Liao, Qinghua Petrović, Dušan Krüger, Dennis M. Strodel, Birgit Amyes, Tina L. Richard, John P. Kamerlin, Shina C. L. Enzyme Architecture: Modeling the Operation of a Hydrophobic Clamp in Catalysis by Triosephosphate Isomerase |
title | Enzyme
Architecture: Modeling the Operation of a Hydrophobic
Clamp in Catalysis by Triosephosphate Isomerase |
title_full | Enzyme
Architecture: Modeling the Operation of a Hydrophobic
Clamp in Catalysis by Triosephosphate Isomerase |
title_fullStr | Enzyme
Architecture: Modeling the Operation of a Hydrophobic
Clamp in Catalysis by Triosephosphate Isomerase |
title_full_unstemmed | Enzyme
Architecture: Modeling the Operation of a Hydrophobic
Clamp in Catalysis by Triosephosphate Isomerase |
title_short | Enzyme
Architecture: Modeling the Operation of a Hydrophobic
Clamp in Catalysis by Triosephosphate Isomerase |
title_sort | enzyme
architecture: modeling the operation of a hydrophobic
clamp in catalysis by triosephosphate isomerase |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5543394/ https://www.ncbi.nlm.nih.gov/pubmed/28683550 http://dx.doi.org/10.1021/jacs.7b05576 |
work_keys_str_mv | AT kulkarniyashrajs enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT liaoqinghua enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT petrovicdusan enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT krugerdennism enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT strodelbirgit enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT amyestinal enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT richardjohnp enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase AT kamerlinshinacl enzymearchitecturemodelingtheoperationofahydrophobicclampincatalysisbytriosephosphateisomerase |