Evolution Enhances Kemp Eliminase Activity by Optimizing Oxyanion Stabilization and Conformational Flexibility
dc.contributor.author
dc.date.accessioned
2024-12-10T15:10:15Z
dc.date.available
2024-12-10T15:10:15Z
dc.date.issued
2025-01-27
dc.identifier.issn
0947-6539
dc.identifier.uri
dc.description.abstract
The base-promoted Kemp elimination reaction has been used as a model system for enzyme design. Among the multiple computationally designed and evolved Kemp eliminases generated along the years, the HG3-to-HG3.17 evolutionary trajectory is particularly interesting due to the high catalytic efficiency of HG3.17 and the debated role of glutamine 50 (Gln50) as potential oxyanion stabilizer. This study aims to elucidate the structural and dynamic changes along the evolutionary pathway from HG3 to HG3.17 that contribute to improved catalytic efficiency. In particular, we evaluate key variants along the HG3 evolutionary trajectory via molecular dynamics simulations coupled to non-covalent interactions and water analysis. Our computational study indicates that HG3.17 can adopt a catalytically competent conformation promoted by a water-mediated network of non-covalent interactions, in which aspartate 127 (Asp127) is properly positioned for proton abstraction and Gln50 and to some extent mutation cysteine 84 (Cys84) contribute to oxyanion stabilization. We find that HG3.17 exhibits a rather high flexibility of Gln50, which is regulated by the conformation adopted by the active site residue tryptophan 44 (Trp44). This interplay between Gln50 and Trp44 positioning induced by distal active site mutations affects the water-mediated network of non-covalent interactions, Gln50 preorganization, and water content of the active site pocket.
dc.description.sponsorship
Research funding: H2020 European Research Council. Grant Numbers: ERC-2015-StG-679001, ERC-2022-CoG-101088032, ERC-2022-POC-101112805, ERC-2023-POC-101158166; Ministerio de Ciencia e Innovación. Grant Numbers: PID2021-129034NB-I00, PDC2022-133950-I00, PRE2022-105114; Human Frontier Science Program Grants. Grant Number: RGP0054/2020 & Generalitat de Catalunya. Grant Number: SGR 2021 00487
Open Access funding provided thanks to the CRUE-CSIC agreement with Wiley
dc.format.mimetype
application/pdf
dc.language.iso
eng
dc.publisher
Wiley
dc.relation
PID2021-129034NB-I00
PDC2022-133950-I00
dc.relation.isformatof
Reproducció digital del document publicat a: https://doi.org/10.1002/chem.202403747
dc.relation.ispartof
Chemistry - A European Journal, 2025, vol. 31, núm. 6, p. e202403747
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Articles publicats (D-Q)
dc.rights
Attribution-NonCommercial-NoDerivatives 4.0 International
dc.rights.uri
dc.subject
dc.title
Evolution Enhances Kemp Eliminase Activity by Optimizing Oxyanion Stabilization and Conformational Flexibility
dc.type
info:eu-repo/semantics/article
dc.rights.accessRights
info:eu-repo/semantics/openAccess
dc.relation.projectID
info:eu-repo/grantAgreement/EC/H2020/679001/EU/Network models for the computational design of proficient enzymes/NetMoDEzyme
info:eu-repo/grantAgreement/EC/HE/101088032/EU/Fast yet accurate routine rational design of novel enzymes/FASTEN
info:eu-repo/grantAgreement/EC/HE/101112805/EU/Computational design of industrial enzymes for green chemistry/GREENZYME
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PID2021-129034NB-I00/ES/DISEÑO COMPUTACIONAL DE ENZIMAS CONFORMACIONALMENTE DIRIGIDO PARA MEJORAR LA ACTIVIDAD AISLADA O EN COMPLEJO/
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PDC2022-133950-I00/ES/EVOLUCION COMPUTACIONAL DE NUEVOS (BIO)CATALIZADORES/
dc.type.version
info:eu-repo/semantics/publishedVersion
dc.identifier.doi
dc.contributor.funder
dc.type.peerreviewed
peer-reviewed
dc.relation.FundingProgramme
dc.relation.ProjectAcronym
dc.identifier.eissn
1521-3765
dc.identifier.PMID
39541157