Molecular dynamics explorations of active site structure in designed and evolved enzymes

This Account describes the use of molecular dynamics (MD) simulations to reveal how mutations alter the structure and organization of enzyme active sites. As proposed by Pauling about 70 years ago and elaborated by many others since then, biocatalysis is efficient when functional groups in the activ...

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Autores: Osuna Oliveras, Sílvia, Jiménez-Osés, Gonzalo, Noey, Elizabeth L., Houk, Kendall N.
Tipo de documento: artigo
Estado:Versão publicada
Data de publicação:2015
País:España
Recursos:Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya)
Repositório:Recercat. Dipósit de la Recerca de Catalunya
OAI Identifier:oai:recercat.cat:10256/11442
Acesso em linha:http://hdl.handle.net/10256/11442
Access Level:Acesso embargado
Palavra-chave:Dinàmica molecular
Molecular dynamics
Enzims
Enzymes
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dc.title.none.fl_str_mv Molecular dynamics explorations of active site structure in designed and evolved enzymes
title Molecular dynamics explorations of active site structure in designed and evolved enzymes
spellingShingle Molecular dynamics explorations of active site structure in designed and evolved enzymes
Osuna Oliveras, Sílvia
Dinàmica molecular
Molecular dynamics
Enzims
Enzymes
title_short Molecular dynamics explorations of active site structure in designed and evolved enzymes
title_full Molecular dynamics explorations of active site structure in designed and evolved enzymes
title_fullStr Molecular dynamics explorations of active site structure in designed and evolved enzymes
title_full_unstemmed Molecular dynamics explorations of active site structure in designed and evolved enzymes
title_sort Molecular dynamics explorations of active site structure in designed and evolved enzymes
dc.creator.none.fl_str_mv Osuna Oliveras, Sílvia
Jiménez-Osés, Gonzalo
Noey, Elizabeth L.
Houk, Kendall N.
author Osuna Oliveras, Sílvia
author_facet Osuna Oliveras, Sílvia
Jiménez-Osés, Gonzalo
Noey, Elizabeth L.
Houk, Kendall N.
author_role author
author2 Jiménez-Osés, Gonzalo
Noey, Elizabeth L.
Houk, Kendall N.
author2_role author
author
author
dc.contributor.none.fl_str_mv Ministerio de Ciencia e Innovación (Espanya)
dc.subject.none.fl_str_mv Dinàmica molecular
Molecular dynamics
Enzims
Enzymes
topic Dinàmica molecular
Molecular dynamics
Enzims
Enzymes
description This Account describes the use of molecular dynamics (MD) simulations to reveal how mutations alter the structure and organization of enzyme active sites. As proposed by Pauling about 70 years ago and elaborated by many others since then, biocatalysis is efficient when functional groups in the active site of an enzyme are in optimal positions for transition state stabilization. Changes in mechanism and covalent interactions are often critical parts of enzyme catalysis. We describe our explorations of the dynamical preorganization of active sites using MD, studying the fluctuations between active and inactive conformations normally concealed to static crystallography. MD shows how the various arrangements of active site residues influence the free energy of the transition state and relates the populations of the catalytic conformational ensemble to the enzyme activity. This Account is organized around three case studies from our laboratory. We first describe the importance of dynamics in evaluating a series of computationally designed and experimentally evolved enzymes for the Kemp elimination, a popular subject in the enzyme design field. We find that the dynamics of the active site is influenced not only by the original sequence design and subsequent mutations but also by the nature of the ligand present in the active site. In the second example, we show how microsecond MD has been used to uncover the role of remote mutations in the active site dynamics and catalysis of a transesterase, LovD. This enzyme was evolved by Tang at UCLA and Codexis, Inc., and is a useful commercial catalyst for the production of the drug simvastatin. X-ray analysis of inactive and active mutants did not reveal differences in the active sites, but relatively long time scale MD in solution showed that the active site of the wild-type enzyme preorganizes only upon binding of the acyl carrier protein (ACP) that delivers the natural acyl group to the active site. In the absence of bound ACP, a noncatalytic arrangement of the catalytic triad is dominant. Unnatural truncated substrates are inactive because of the lack of protein-protein interactions provided by the ACP. Directed evolution is able to gradually restore the catalytic organization of the active site by motion of the protein backbone that alters the active site geometry. In the third case, we demonstrate the key role of MD in combination with crystallography to identify the origins of substrate-dependent stereoselectivities in a number of Codexis-engineered ketoreductases, one of which is used commercially for the production of the antibiotic sulopenem. Here, mutations alter the shape of the active site as well as the accessibility of water to different regions of it. Each of these examples reveals something different about how mutations can influence enzyme activity and shows that directed evolution, like natural evolution, can increase catalytic activity in a variety of remarkable and often subtle ways
publishDate 2015
dc.date.none.fl_str_mv 2015
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dc.identifier.none.fl_str_mv http://hdl.handle.net/10256/11442
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url http://hdl.handle.net/10256/11442
dc.language.none.fl_str_mv Inglés
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dc.publisher.none.fl_str_mv American Chemical Society (ACS)
publisher.none.fl_str_mv American Chemical Society (ACS)
dc.source.none.fl_str_mv © Accounts of Chemical Research, 2015, vol. 48, núm. 4, p. 1080-1089
Articles publicats (D-Q)
reponame:Recercat. Dipósit de la Recerca de Catalunya
instname:Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya)
instname_str Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya)
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spelling Molecular dynamics explorations of active site structure in designed and evolved enzymesOsuna Oliveras, SílviaJiménez-Osés, GonzaloNoey, Elizabeth L.Houk, Kendall N.Dinàmica molecularMolecular dynamicsEnzimsEnzymesThis Account describes the use of molecular dynamics (MD) simulations to reveal how mutations alter the structure and organization of enzyme active sites. As proposed by Pauling about 70 years ago and elaborated by many others since then, biocatalysis is efficient when functional groups in the active site of an enzyme are in optimal positions for transition state stabilization. Changes in mechanism and covalent interactions are often critical parts of enzyme catalysis. We describe our explorations of the dynamical preorganization of active sites using MD, studying the fluctuations between active and inactive conformations normally concealed to static crystallography. MD shows how the various arrangements of active site residues influence the free energy of the transition state and relates the populations of the catalytic conformational ensemble to the enzyme activity. This Account is organized around three case studies from our laboratory. We first describe the importance of dynamics in evaluating a series of computationally designed and experimentally evolved enzymes for the Kemp elimination, a popular subject in the enzyme design field. We find that the dynamics of the active site is influenced not only by the original sequence design and subsequent mutations but also by the nature of the ligand present in the active site. In the second example, we show how microsecond MD has been used to uncover the role of remote mutations in the active site dynamics and catalysis of a transesterase, LovD. This enzyme was evolved by Tang at UCLA and Codexis, Inc., and is a useful commercial catalyst for the production of the drug simvastatin. X-ray analysis of inactive and active mutants did not reveal differences in the active sites, but relatively long time scale MD in solution showed that the active site of the wild-type enzyme preorganizes only upon binding of the acyl carrier protein (ACP) that delivers the natural acyl group to the active site. In the absence of bound ACP, a noncatalytic arrangement of the catalytic triad is dominant. Unnatural truncated substrates are inactive because of the lack of protein-protein interactions provided by the ACP. Directed evolution is able to gradually restore the catalytic organization of the active site by motion of the protein backbone that alters the active site geometry. In the third case, we demonstrate the key role of MD in combination with crystallography to identify the origins of substrate-dependent stereoselectivities in a number of Codexis-engineered ketoreductases, one of which is used commercially for the production of the antibiotic sulopenem. Here, mutations alter the shape of the active site as well as the accessibility of water to different regions of it. Each of these examples reveals something different about how mutations can influence enzyme activity and shows that directed evolution, like natural evolution, can increase catalytic activity in a variety of remarkable and often subtle waysThis research was supported by NIH NIGMS (GM036700, GM097200, and GM075962). ANTON simulations were performed on the National Resource for Biomedical Supercomputing at the Pittsburgh Supercomputing Center with funding from NIH NIGMS (RC2GM093307). This work used the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by NSF (OCI-1053575). The authors acknowledge Dr. Jiyong Park for computational support and discussion and Dr. Xiyun Zhang (Codexis) for helpful discussions. S.O. acknowledges the Spanish MINECO for Project CTQ2011-25086/BQU and JdC Grant JCI-2012-14438 and the European Community for CIG Project PCIG14-GA-2013-630978American Chemical Society (ACS)Ministerio de Ciencia e Innovación (Espanya)infoinfo2015info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionapplication/pdfhttp://hdl.handle.net/10256/11442http://hdl.handle.net/10256/11442© Accounts of Chemical Research, 2015, vol. 48, núm. 4, p. 1080-1089Articles publicats (D-Q)reponame:Recercat. Dipósit de la Recerca de Catalunyainstname:Varias* (Consorci de Biblioteques Universitáries de Catalunya, Centre de Serveis Científics i Acadèmics de Catalunya)Inglésinfo:eu-repo/semantics/altIdentifier/doi/10.1021/ar500452qinfo:eu-repo/semantics/altIdentifier/issn/0001-4842info:eu-repo/semantics/altIdentifier/eissn/1520-4898info:eu-repo/grantAgreement/MICINN//CTQ2011-25086info:eu-repo/grantAgreement/EC/FP7/630978Tots els drets reservatsinfo:eu-repo/semantics/embargoedAccessoai:recercat.cat:10256/114422026-05-29T05:05:01Z
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