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dc.contributor.authorSantiago, Gerard
dc.contributor.authorMartínez-Martínez, Mónica
dc.contributor.authorAlonso, Sandra
dc.contributor.authorBargiela, Rafael
dc.contributor.authorCoscolín, Cristina
dc.contributor.authorGolyshing, Peter N.
dc.contributor.authorGuallar, Victor
dc.contributor.authorFerrer, Manuel
dc.contributor.otherBarcelona Supercomputing Center
dc.identifier.citationSantiago, G. [et al.]. Rational Engineering of Multiple Active Sites in an Ester Hydrolase. "Biochemistry", 30 Març 2018, vol. 57, núm. 15, p. 2245-2255.
dc.description.abstractEffects of altering the properties of an active site in an enzymatic homogeneous catalyst have been extensively reported. However, the possibility of increasing the number of such sites, as commonly done in heterogeneous catalytic materials, remains unexplored, particularly because those have to accommodate appropriate residues in specific configurations. This possibility was investigated by using a serine ester hydrolase as the target enzyme. By using the Protein Energy Landscape Exploration software, which maps ligand diffusion and binding, we found a potential binding pocket capable of holding an extra catalytic triad and oxyanion hole contacts. By introducing two mutations, this binding pocket became a catalytic site. Its substrate specificity, substrate preference, and catalytic activity were different from those of the native site of the wild type ester hydrolase and other hydrolases, due to the differences in the active site architecture. Converting the binding pocket into an extra catalytic active site was proven to be a successful approach to create a serine ester hydrolase with two functional reactive groups. Our results illustrate the accuracy and predictive nature of modern modeling techniques, opening novel catalytic opportunities coming from the presence of different catalytic environments in single enzymes.
dc.description.sponsorshipThis project received funding from the European Union’s Horizon 2020 research and innovation program [Blue Growth: Unlocking the potential of Seas and Oceans] under grant agreement no. [634486] (project acronym INMARE). This research was also supported by the grants PCIN-2014-107 (within ERA NET IB2 grant nr. ERA-IB-14-030 - MetaCat), PCIN-2017-078 (within the ERA-MarineBiotech grant ProBone), BIO2014-54494-R, CTQ2016-79138-R and BIO2017-85522-R from the Spanish Ministry of Economy, Industry and Competitiveness. P.N.G. gratefully acknowledges funding from the UK Biotechnology and Biological Sciences Research Council (grant no. BB/M029085/1). R.B. and P.N.G. acknowledge the support of the Supercomputing Wales project, which is part-funded by the European Regional De-velopment Fund (ERDF) via Welsh Government. P.N.G. acknowledges the support of the Centre of Environmental Biotechnology Project funded by the European Regional Development Fund (ERDF) through Welsh Government. The authors gratefully acknowledge financial support provided by the European Regional Development Fund (ERDF). The MALDI-TOF/TOF analysis was performed in the proteomics facility of the Spanish National Center for Biotechnology (CNB-CSIC) that belongs to ProteoRed, PRB2-ISCIII, sup-ported by grant PT13/0001.
dc.format.extent11 p.
dc.publisherAmerican Chemical Society
dc.rightsACS AuthorChoice - This is an open access article published under an ACS AuthorChoice License, which permits copying and redistribution of the article or any adaptations for non-commercial purposes.
dc.subjectÀrees temàtiques de la UPC::Enginyeria biomèdica
dc.subject.lcshEnzyme activation
dc.subject.otherProtein Energy Landscape Exploration software
dc.titleRational Engineering of Multiple Active Sites in an Ester Hydrolase
dc.description.peerreviewedPeer Reviewed
dc.rights.accessOpen Access
dc.description.versionPostprint (published version)
dc.relation.projectidinfo:eu-repo/grantAgreement/EC/H2020/634486/EU/Industrial Applications of Marine Enzymes: Innovative screening and expression platforms to discover and use the functional protein diversity from the sea/INMARE

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