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dc.contributor.authorŚwiderek, Katarzyna
dc.contributor.authorTuñón, Iñaki
dc.contributor.authorMartí Forés, Sergio
dc.contributor.authorMoliner, Vicent
dc.date.accessioned2016-05-12T11:40:22Z
dc.date.available2016-05-12T11:40:22Z
dc.date.issued2015
dc.identifier.citationSWIDEREK, Katarzyna; TUÑÓN, Iñaki; MARTÍ FORÉS, Sergio; MOLINER IBÁÑEZ, Vicente. Protein Conformational Landscapes and Catalysis. Influence of Active Site Conformations in the Reaction Catalyzed by L-Lactate Dehydrogenase. ACS Catalysis (2015), v. 2, n. 5, pp. 1172-1185ca_CA
dc.identifier.urihttp://hdl.handle.net/10234/159588
dc.description.abstractIn the past decade, L-Lactate Dehydrogenase (LDH) has become an extremely useful marker in both clinical diagnosis and in monitoring the course of many human diseases. It has been assumed since the 1980s that the full catalytic process of LDH starts with the binding of the cofactor and the substrate followed by the enclosure of the active site by a mobile loop of the protein before the reaction takes place. In this paper, we show that the chemical step of the LDH-catalyzed reaction can proceed within the open loop conformation, and the different reactivity of the different protein conformations would be in agreement with the broad range of rate constants measured in single-molecule spectrometry studies. Starting from a recently solved X-ray diffraction structure that presented an open loop conformation in two of the four chains of the tetramer, QM/MM free energy surfaces have been obtained at different levels of theory. Depending on the level of theory used to describe the electronic structure, the free energy barrier for the transformation of pyruvate into lactate with the open conformation of the protein varies between 12.9 and 16.3 kcal/mol, after quantizing the vibrations and adding the contributions of recrossing and tunneling effects. These values are very close to the experimentally deduced one (14.2 kcal·mol–1) and ∼2 kcal·mol–1 smaller than the one obtained with the closed loop conformer. Calculation of primary KIEs and IR spectra in both protein conformations are also consistent with our hypothesis and in agreement with experimental data. Our calculations suggest that the closure of the active site is mainly required for the inverse process—the oxidation of lactate to pyruvate. According to this hypothesis, H4-type LDH enzyme molecules should have a better ability to close the mobile loop than the M4-type LDH molecules.ca_CA
dc.description.sponsorShipWe thank the Spanish Ministerio de Economía y Competitividad for project CTQ2012-36253-C03, Universitat Jaume I (project P1•1B2011-23), Generalitat Valenciana (PROMETEOII/2014/022 and ACOMP/2014/277 projects), Polish National Center for Science (NCN) (grant 2011/02/A/ST4/00246, 2012− 2017) and the USA National Institute of Health (ref NIH R01 GM065368).ca_CA
dc.format.extent14 p.ca_CA
dc.format.mimetypeapplication/pdfca_CA
dc.language.isoengca_CA
dc.publisherACS Publicationsca_CA
dc.relation.isPartOfACS Catalysis (2015), v. 2, n. 5ca_CA
dc.subjectLDHca_CA
dc.subjectReaction mechanismca_CA
dc.subjectQM/MMca_CA
dc.subjectKIEsca_CA
dc.subjectFree energy surfacesca_CA
dc.subjectSingle-molecule experimentsca_CA
dc.titleProtein Conformational Landscapes and Catalysis. Influence of Active Site Conformations in the Reaction Catalyzed by L-Lactate Dehydrogenaseca_CA
dc.typeinfo:eu-repo/semantics/articleca_CA
dc.identifier.doihttp://dx.doi.org/10.1021/cs501704f
dc.rights.accessRightsinfo:eu-repo/semantics/restrictedAccessca_CA
dc.relation.publisherVersionhttp://pubs.acs.org/doi/abs/10.1021/cs501704fca_CA


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