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Aldehyde oxidation by pleurotus aryl-alcohol oxidase involved in lignin biodegradation

AuthorsFerreira, Patricia ; Hernández-Ortega, Aitor ; Herguedas, Beatriz ; Rencoret, Jorge ; Gutiérrez Suárez, Ana ; Martínez Hernández, María Jesús; Jiménez-Barbero, Jesús ; Medina, Milagros; Martínez, Ángel T.
Issue Date2010
CitationLignobiotech One Symposium (2010)
AbstractAryl-alcohol oxidase (AAO) is an extracellular flavoenzyme providing H2O2 for fungal degradation of lignin. AAO is active on different benzyl alcohols that are oxidized to the corresponding aldehydes. However, the H2O2 formed from some of them was more than stoichiometric with respect to the aldehyde detected. This was due to a double reaction that involves aryl-aldehyde oxidase activity, which was investigated here using different benzylic aldehydes. Formation of the corresponding acids was demonstrated by gas chromatography-mass spectrometry. The chromatographic results, together with the steady and pre-steady state kinetics, revealed a strong influence of electron withdrawing/donating substrate substituents on activity, being the highest on p-nitrobenzaldehyde and halogenated aldehydes and the lowest on methoxylated aldehydes. The presence of these substituents was correlated to the aldehyde hydration rates estimated by 1H nuclear magnetic resonance. These findings, together with the absence in the AAO active site of a residue able to drive oxidation via an aldehyde thiohemiacetal, suggested that oxidation mainly proceeds via the hydrated (gem-diol) species. The reaction mechanism (with solvent isotope effect of D2Okred ~1.5) would be analogous to that described for alcohols, the rate-limiting reductive half-reaction involving concerted hydride transfer from the substrate α-carbon to the cofactor isoalloxazine ring, and proton abstraction from one of the gem-diol hydroxyls by a catalytic base. The existence of two steps of opposite polar requirements (hydration and hydride transfer) explains some aspects of aldehyde oxidation by AAO. Site-directed mutagenesis identified two histidines strongly involved in gem-diol oxidation and, unexpectedly, suggested that an active-site tyrosine could facilitate the oxidation of some aldehydes showing no detectable hydration. It seems that AAO has evolved to use differently substituted benzylic metabolites for H2O2 production. Depending of the compound available, AAO will behave basically as an alcohol oxidase (e.g. in the presence of veratrylic and anisylic metabolites), as an aldehyde oxidase (e.g. in the presence of p-nitrobenzylic metabolites) or as a double oxidase being able to transform alcohols into the corresponding acids (e.g. in the presence of 3-chloro-p-anisylic metabolites). This versatility would help AAO to provide maximal H2O2 supply under variable environmental conditions. The study has been supported by the BIORENEW EU-project (NMP2-CT-2006-026456).
DescriptionPóster presentado en el Topic 1: Fundamentals of lignocellulosic enzymes and transformation mechanisms (EF), en el citado simposio, celebrado del 28 de marzo al 1 de abril de 2010, en Reims, Francia.
Appears in Collections:(IRNAS) Comunicaciones congresos
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