71218 20201130083155.0 doi 10.1038/s41598-018-26445-x sideral 106782 ART-2018-106782 eng Carro, Juan Multiple implications of an active site phenylalanine in the catalysis of aryl-alcohol oxidase 2018 Access copy available to the general public Unrestricted Aryl-alcohol oxidase (AAO) has demonstrated to be an enzyme with a bright future ahead due to its biotechnological potential in deracemisation of chiral compounds, production of bioplastic precursors and other reactions of interest. Expanding our understanding on the AAO reaction mechanisms, through the investigation of its structure-function relationships, is crucial for its exploitation as an industrial biocatalyst. In this regard, previous computational studies suggested an active role for AAO Phe397 at the active-site entrance. This residue is located in a loop that partially covers the access to the cofactor forming a bottleneck together with two other aromatic residues. Kinetic and a nity spectroscopic studies, complemented with computational simulations using the recently developed adaptive-PELE technology, reveal that the Phe397 residue is important for product release and to help the substrates attain a catalytically relevant position within the active-site cavity. Moreover, removal of aromaticity at the 397 position impairs the oxygen-reduction activity of the enzyme. Experimental and computational ndings agree very well in the timing of product release from AAO, and the simulations help to understand the experimental results. This highlights the potential of adaptive-PELE to provide answers to the questions raised by the empirical results in the study of enzyme mechanisms. info:eu-repo/grantAgreement/ES/MINECO/NOESIS-BIO2014-56388-R info:eu-repo/grantAgreement/ES/MINECO/CTQ2016-79138-R info:eu-repo/grantAgreement/ES/MINECO/BIO2016-75183-P info:eu-repo/grantAgreement/ES/MEC/FPU-AP2012-2041 This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No H2020 720297-EnzOx2 info:eu-repo/grantAgreement/EC/H2020/720297/EU/New enzymatic oxidation/oxyfunctionalization technologies for added value bio-based products/EnzOx2 info:eu-repo/grantAgreement/EC/FP7/613549/EU/Optimized oxidoreductases for medium and large scale industrial biotransformations/INDOX info:eu-repo/semantics/openAccess by http://creativecommons.org/licenses/by/3.0/es/ 4.011 2018 MULTIDISCIPLINARY SCIENCES 14 / 69 = 0.203 2018 Q1 T1 1.414 2018 Multidisciplinary 2018 Q1 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Amengual-Rigo, Pep Ferran, Sancho (orcid)0000-0001-8743-0182 Medina, Milagros Universidad de Zaragoza Guallar, Victor (orcid)0000-0003-4076-6118 Ferreira, Patricia Universidad de Zaragoza Martínez, T. Angel 1002 060 Universidad de Zaragoza Dpto. Bioq.Biolog.Mol. Celular Área Bioquímica y Biolog.Mole. 8 (2018), 8121 [12 pp.] Sci. rep. Scientific Reports 2045-2322 2541375 http://zaguan.unizar.es/record/71218/files/texto_completo.pdf Versión publicada 116311 http://zaguan.unizar.es/record/71218/files/texto_completo.jpg?subformat=icon icon Versión publicada oai:zaguan.unizar.es:71218 articulos driver 2020-11-30-07:57:32 ARTICLE