title: Structure-function characterization reveals new catalytic diversity in the galactose oxidase and glyoxal oxidase family.
abstract: Alcohol oxidases, including carbohydrate oxidases, have a long history of research that has generated fundamental biological understanding and biotechnological applications. Despite a long history of study, the galactose 6-oxidase/glyoxal oxidase family of mononuclear copper-radical oxidases, Auxiliary Activity Family 5 (AA5), is currently represented by only very few characterized members. Here we report the recombinant production and detailed structure-function analyses of two homologues from the phytopathogenic fungi Colletotrichum graminicola and C. gloeosporioides, CgrAlcOx and CglAlcOx, respectively, to explore the wider biocatalytic potential in AA5. EPR spectroscopy and crystallographic analysis confirm a common active-site structure vis-à-vis the archetypal galactose 6-oxidase from Fusarium graminearum. Strikingly, however, CgrAlcOx and CglAlcOx are essentially incapable of oxidizing galactose and galactosides, but instead efficiently catalyse the oxidation of diverse aliphatic alcohols. The results highlight the significant potential of prospecting the evolutionary diversity of AA5 to reveal novel enzyme specificities, thereby informing both biology and applications.
authors.affiliation: Michael Smith Laboratories and Department of Chemistry, University of British Columbia, 2185 East Mall, Vancouver, British Columbia, Canada V6T 1Z4.
authors.forename: DeLu Tyler
authors.initials: DT
authors.lastname: Yin
authors.affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, UK.
authors.forename: Saioa
authors.initials: S
authors.lastname: Urresti
authors.affiliation: Michael Smith Laboratories and Department of Chemistry, University of British Columbia, 2185 East Mall, Vancouver, British Columbia, Canada V6T 1Z4.|Institut des Sciences Moléculaires de Marseille-Team BiosCiences UMR 7313-CNRS, Aix-Marseille University, Avenue Escadrille Normandie Niemen, Marseille 13397, France.
authors.forename: Mickael
authors.initials: M
authors.lastname: Lafond
authors.affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, UK.
authors.forename: Esther M
authors.initials: EM
authors.lastname: Johnston
authors.affiliation: Michael Smith Laboratories and Department of Chemistry, University of British Columbia, 2185 East Mall, Vancouver, British Columbia, Canada V6T 1Z4.
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Derikvand
authors.affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, UK.
authors.forename: Luisa
authors.initials: L
authors.lastname: Ciano
authors.affiliation: INRA, UMR1163 Biodiversité et Biotechnologie Fongiques Marseille F-13288, France.
authors.forename: Jean-Guy
authors.initials: JG
authors.lastname: Berrin
authors.affiliation: Architecture et Fonction des Macromolécules Biologiques, CNRS-Aix-Marseille University, 163 Avenue de Luminy, Marseille 13288, France.|INRA, USC 1408 AFMB, Marseille 13288, France.|Department of Biological Sciences, King Abdulaziz University, Jeddah, Saudi Arabia.
authors.forename: Bernard
authors.initials: B
authors.lastname: Henrissat
authors.affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, UK.
authors.forename: Paul H
authors.initials: PH
authors.lastname: Walton
authors.affiliation: Department of Chemistry, University of York, Heslington, York YO10 5DD, UK.
authors.forename: Gideon J
authors.initials: GJ
authors.lastname: Davies
authors.affiliation: Michael Smith Laboratories and Department of Chemistry, University of British Columbia, 2185 East Mall, Vancouver, British Columbia, Canada V6T 1Z4.
authors.forename: Harry
authors.initials: H
authors.lastname: Brumer
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D000438:Alcohols / Q000378:metabolism; D020134:Catalytic Domain; D020231:Colletotrichum; D003460:Crystallization; D018360:Crystallography, X-Ray; D004578:Electron Spin Resonance Spectroscopy; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism*; D005670:Fusarium; D005692:Galactose Oxidase / Q000737:chemistry / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010802:Phylogeny; D010843:Pichia; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D066244:Proton Magnetic Resonance Spectroscopy; D011994:Recombinant Proteins
chemical_list: D000438:Alcohols; D005656:Fungal Proteins; D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C052371:glyoxal oxidase; C019643:alcohol oxidase; D005692:Galactose Oxidase