Protein dynamics promote hydride tunnelling in substrate oxidation by aryl-alcohol oxidase

Carro, Juan ; Martínez-Julve, Marta (Universidad de Zaragoza) ; Medina, Milagros (Universidad de Zaragoza) ; Martínez, Angel T. ; Ferreira, Patricia (Universidad de Zaragoza)

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)

Financiación: info:eu-repo/grantAgreement/EC/FP7/613549/EU/Optimized oxidoreductases for medium and large scale industrial biotransformations/INDOX
Financiación: info:eu-repo/grantAgreement/EC/H2020/720297/EU/New enzymatic oxidation/oxyfunctionalization technologies for added value bio-based products/EnzOx2
Financiación: info:eu-repo/grantAgreement/ES/MINECO/BIO2016-75183-P
Financiación: info:eu-repo/grantAgreement/ES/MINECO/NOESIS-BIO2014-56388-R

Tipo y forma: Artículo (PostPrint)

Área (Departamento): Área Bioquímica y Biolog.Mole. (Dpto. Bioq.Biolog.Mol. Celular)