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Discovery of a fungal copper radical oxidase with high catalytic efficiency towards 5-hydroxymethylfurfural and benzyl alcohols for bioprocessing

DOI: 10.1021/acscatal.9b04727 DOI Help

Authors: Yann Mathieu (University of British Columbia) , Wendy A. Offen (University of York) , Stephanie M. Forget (University of British Columbia) , Luisa Ciano (University of British Columbia) , Alexander Holm Viborg (University of British Columbia) , Elena Blagova (University of British Columbia) , Bernard Henrissat (CNRS, Aix-Marseille University; INRA, USC1408 Architecture et Fonction des Macromolécules Biologiques (AFMB)) , Paul H. Walton (University of British Columbia) , Gideon J. Davies (University of British Columbia) , Harry Brumer (University of British Columbia; University of York)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Acs Catalysis

State: Published (Approved)
Published: February 2020
Diamond Proposal Number(s): 13587 , 18598

Abstract: Alternatives to petroleum-based chemicals are highly sought-after for on-going efforts to reduce the damaging effects of human activity on the environment. Copper radical oxidases from Auxiliary Activity Family 5/Subfamily 2 (AA5_2) are attractive biocatalysts because they oxidize primary alcohols in a chemo-selective manner without complex organic cofactors. However, despite numerous studies on canonical galactose oxidases (GalOx, EC and engineered variants, and the recent discovery of a Colletotrichum graminicola copper radical alcohol oxidase (AlcOx, EC, the catalytic potentials of very few AA5_2 members have been characterized. Guided by sequence similarity network and phylogenetic analyses, in this study we targeted a distinct paralog from the fungus C. graminicola as a representative member of a large uncharacterized subgroup of AA5_2. Through recombinant production and detailed kinetic analysis, we demonstrated that this enzyme is weakly active towards carbohydrates, but efficiently catalyzes the oxidation of aryl alcohols to the corresponding aldehydes. As such, this represents the initial characterization of a demonstrable aryl alcohol oxidase (AAO, EC in AA5, an activity which is classically associated with flavin-dependent glucose-methanol-choline (GMC) oxidoreductases of Auxiliary Activity Family 3 (AA3). X-ray crystallography revealed a distinct multidomain architecture comprising an N-terminal PAN domain abutting a canonical AA5 seven-bladed propeller catalytic domain. Of direct relevance to biomass processing, the wild-type enzyme exhibits the highest activity on the primary alcohol of 5-hydroxymethylfurfural (HMF), a product of significant interest in the lignocellulosic bio-refinery concept. Thus, the chemoselective oxidation of HMF to 2,5-diformylfuran (DFF) by C. graminicola aryl alcohol oxidase (CgrAAO) from AA5 provides a fundamental building block for chemistry via biotechnology.

Journal Keywords: oxidoreductases; enzyme kinetics; structural biology; biocatalysis; bioproducts; EC; EC

Subject Areas: Chemistry

Instruments: I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography