Maria Cleveland1,2,3, Mickael Lafond1,4, Fan Roderick Xia1,2,3, Ryan Chung2, Paul Mulyk2, Jason E Hein2, Harry Brumer5,6,7,8,9. 1. Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, BC, V6T 1Z4, Canada. 2. Department of Chemistry, University of British Columbia, 2036 Main Mall, Vancouver, BC, V6T 1Z1, Canada. 3. BioProducts Institute, University of British Columbia, 2385 East Mall, Vancouver, BC, V6T 1Z4, Canada. 4. Aix Marseille Univ, CNRS, Centrale Marseille, iSm2, Marseille, France. 5. Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, BC, V6T 1Z4, Canada. brumer@msl.ubc.ca. 6. Department of Chemistry, University of British Columbia, 2036 Main Mall, Vancouver, BC, V6T 1Z1, Canada. brumer@msl.ubc.ca. 7. BioProducts Institute, University of British Columbia, 2385 East Mall, Vancouver, BC, V6T 1Z4, Canada. brumer@msl.ubc.ca. 8. Department of Biochemistry and Molecular Biology, University of British Columbia, 2350 Health Sciences Mall, Vancouver, BC, V6T 1Z3, Canada. brumer@msl.ubc.ca. 9. Department of Botany, University of British Columbia, 3200 University Boulevard, Vancouver, BC, V6T 1Z4, Canada. brumer@msl.ubc.ca.
Abstract
BACKGROUND: Biomass valorization has been suggested as a sustainable alternative to petroleum-based energy and commodities. In this context, the copper radical oxidases (CROs) from Auxiliary Activity Family 5/Subfamily 2 (AA5_2) are attractive biocatalysts for the selective oxidation of primary alcohols to aldehydes. Originally defined by the archetypal galactose 6-oxidase from Fusarium graminearum, fungal AA5_2 members have recently been shown to comprise a wide range of specificities for aromatic, aliphatic and furan-based alcohols. This suggests a broader substrate scope of native CROs for applications. However, only 10% of the annotated AA5_2 members have been characterized to date. RESULTS: Here, we define two homologues from the filamentous fungi Fusarium graminearum and F. oxysporum as predominant aryl alcohol oxidases (AAOs) through recombinant production in Pichia pastoris, detailed kinetic characterization, and enzyme product analysis. Despite possessing generally similar active-site architectures to the archetypal FgrGalOx, FgrAAO and FoxAAO have weak activity on carbohydrates, but instead efficiently oxidize specific aryl alcohols. Notably, both FgrAAO and FoxAAO oxidize hydroxymethyl furfural (HMF) directly to 5-formyl-2-furoic acid (FFCA), and desymmetrize the bioproduct glycerol to the uncommon L-isomer of glyceraldehyde. CONCLUSIONS: This work expands understanding of the catalytic diversity of CRO from AA5_2 to include unique representatives from Fusarium species that depart from the well-known galactose 6-oxidase activity of this family. Detailed enzymological analysis highlights the potential biotechnological applications of these orthologs in the production of renewable plastic polymer precursors and other chemicals.
BACKGROUND: Biomass valorization has been suggested as a sustainable alternative to petroleum-based energy and commodities. In this context, the copper radical oxidases (CROs) from Auxiliary Activity Family 5/Subfamily 2 (AA5_2) are attractive biocatalysts for the selective oxidation of primary alcohols to aldehydes. Originally defined by the archetypal galactose 6-oxidase from Fusarium graminearum, fungal AA5_2 members have recently been shown to comprise a wide range of specificities for aromatic, aliphatic and furan-based alcohols. This suggests a broader substrate scope of native CROs for applications. However, only 10% of the annotated AA5_2 members have been characterized to date. RESULTS: Here, we define two homologues from the filamentous fungi Fusarium graminearum and F. oxysporum as predominant aryl alcohol oxidases (AAOs) through recombinant production in Pichia pastoris, detailed kinetic characterization, and enzyme product analysis. Despite possessing generally similar active-site architectures to the archetypal FgrGalOx, FgrAAO and FoxAAO have weak activity on carbohydrates, but instead efficiently oxidize specific aryl alcohols. Notably, both FgrAAO and FoxAAO oxidize hydroxymethyl furfural (HMF) directly to 5-formyl-2-furoic acid (FFCA), and desymmetrize the bioproduct glycerol to the uncommon L-isomer of glyceraldehyde. CONCLUSIONS: This work expands understanding of the catalytic diversity of CRO from AA5_2 to include unique representatives from Fusarium species that depart from the well-known galactose 6-oxidase activity of this family. Detailed enzymological analysis highlights the potential biotechnological applications of these orthologs in the production of renewable plastic polymer precursors and other chemicals.
Authors: Christina M Payne; Brandon C Knott; Heather B Mayes; Henrik Hansson; Michael E Himmel; Mats Sandgren; Jerry Ståhlberg; Gregg T Beckham Journal: Chem Rev Date: 2015-01-28 Impact factor: 60.622
Authors: S Venkata Mohan; G N Nikhil; P Chiranjeevi; C Nagendranatha Reddy; M V Rohit; A Naresh Kumar; Omprakash Sarkar Journal: Bioresour Technol Date: 2016-03-29 Impact factor: 9.642
Authors: Anthony Levasseur; Elodie Drula; Vincent Lombard; Pedro M Coutinho; Bernard Henrissat Journal: Biotechnol Biofuels Date: 2013-03-21 Impact factor: 6.040
Authors: Maria E Cleveland; Yann Mathieu; David Ribeaucourt; Mireille Haon; Paul Mulyk; Jason E Hein; Mickael Lafond; Jean-Guy Berrin; Harry Brumer Journal: Cell Mol Life Sci Date: 2021-11-05 Impact factor: 9.261