Resumen: The temperature dependence of hydride transfer from the substrate to the N5 of the FAD cofactor during the reductive half-reaction of Pleurotus eryngii aryl-alcohol oxidase (AAO) is assessed here. Kinetic isotope effects on both the pre-steady state reduction of the enzyme and its steady-state kinetics, with differently deuterated substrates, suggest an environmentally-coupled quantum-mechanical tunnelling process. Moreover, those kinetic data, along with the crystallographic structure of the enzyme in complex with a substrate analogue, indicate that AAO shows a pre-organized active site that would only require the approaching of the hydride donor and acceptor for the tunnelled transfer to take place. Modification of the enzyme''s active-site architecture by replacement of Tyr92, a residue establishing hydrophobic interactions with the substrate analogue in the crystal structure, in the Y92F, Y92L and Y92W variants resulted in different temperature dependence patterns that indicated a role of this residue in modulating the transfer reaction. Idioma: Inglés DOI: 10.1039/C7CP05904C Año: 2020 Publicado en: Physical Chemistry Chemical Physics 19 (2020), 28666 ISSN: 1463-9076 Factor impacto JCR: 3.676 (2020) Categ. JCR: PHYSICS, ATOMIC, MOLECULAR & CHEMICAL rank: 8 / 37 = 0.216 (2020) - Q1 - T1 Categ. JCR: CHEMISTRY, PHYSICAL rank: 77 / 162 = 0.475 (2020) - Q2 - T2 Factor impacto SCIMAGO: 1.052 - Physics and Astronomy (miscellaneous) (Q1) - Physical and Theoretical Chemistry (Q1)