UCL Discovery
UCL home » Library Services » Electronic resources » UCL Discovery

Exploiting correlated molecular-dynamics networks to counteract enzyme activity–stability trade-off

Yu, H; Dalby, P; (2018) Exploiting correlated molecular-dynamics networks to counteract enzyme activity–stability trade-off. Proceedings of the National Academy of Sciences of the United States of America , 115 (52) E12192-E12200. 10.1073/pnas.1812204115. Green open access

[thumbnail of Dalby VoR E12192.full.pdf]
Preview
Text
Dalby VoR E12192.full.pdf - Published Version

Download (1MB) | Preview

Abstract

The directed evolution of enzymes for improved activity or substrate specificity commonly leads to a trade-off in stability. We have identified an activity–stability trade-off and a loss in unfolding cooperativity for a variant (3M) of Escherichia coli transketolase (TK) engineered to accept aromatic substrates. Molecular dynamics simulations of 3M revealed increased flexibility in several interconnected active-site regions that also form part of the dimer interface. Mutating the newly flexible active-site residues to regain stability risked losing the new activity. We hypothesized that stabilizing mutations could be targeted to residues outside of the active site, whose dynamics were correlated with the newly flexible active-site residues. We previously stabilized WT TK by targeting mutations to highly flexible regions. These regions were much less flexible in 3M and would not have been selected a priori as targets using the same strategy based on flexibility alone. However, their dynamics were highly correlated with the newly flexible active-site regions of 3M. Introducing the previous mutations into 3M reestablished the WT level of stability and unfolding cooperativity, giving a 10.8-fold improved half-life at 55 °C, and increased midpoint and aggregation onset temperatures by 3 °C and 4.3 °C, respectively. Even the activity toward aromatic aldehydes increased up to threefold. Molecular dynamics simulations confirmed that the mutations rigidified the active-site via the correlated network. This work provides insights into the impact of rigidifying mutations within highly correlated dynamic networks that could also be useful for developing improved computational protein engineering strategies.

Type: Article
Title: Exploiting correlated molecular-dynamics networks to counteract enzyme activity–stability trade-off
Open access status: An open access version is available from UCL Discovery
DOI: 10.1073/pnas.1812204115
Publisher version: https://doi.org/10.1073/pnas.1812204115
Language: English
Additional information: Copyright © 2018 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution License 4.0 (CC BY): http://creativecommons.org/licenses/by/4.0/
Keywords: dynamics, trade-off, protein engineering, transketolase, stability
UCL classification: UCL
UCL > Provost and Vice Provost Offices > UCL BEAMS
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science > Dept of Biochemical Engineering
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences > Dept of Chemistry
URI: https://discovery.ucl.ac.uk/id/eprint/10064089
Downloads since deposit
86Downloads
Download activity - last month
Download activity - last 12 months
Downloads by country - last 12 months

Archive Staff Only

View Item View Item