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Structural and functional characterisation of multi-copper oxidase CueO from lignin-degrading bacterium Ochrobactrum sp. reveal its activity towards lignin model compounds and lignosulfonate

DOI: 10.1111/febs.14437 DOI Help

Authors: Rommel Santiago Granja-Travez (University of Warwick) , Rachael Wilkinson (University of Warwick) , Gabriela Felix Persinoti (Universidade de Sorocaba) , Fabio M. Squina (Universidade de Sorocaba) , Vilmos Fulop (University of Warwick) , Timothy D. H. Bugg (University of Warwick)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: The Febs Journal , VOL 27

State: Published (Approved)
Published: March 2018
Diamond Proposal Number(s): 14692

Abstract: The identification of enzymes responsible for oxidation of lignin in lignin‐degrading bacteria is of interest for biotechnological valorization of lignin to renewable chemical products. The genome sequences of two lignin‐degrading bacteria, Ochrobactrum sp., and Paenibacillus sp., contain no B‐type DyP peroxidases implicated in lignin degradation in other bacteria, but contain putative multicopper oxidase genes. Multi‐copper oxidase CueO from Ochrobactrum sp. was expressed and reconstituted as a recombinant laccase‐like enzyme, and kinetically characterized. Ochrobactrum CueO shows activity for oxidation of β‐aryl ether and biphenyl lignin dimer model compounds, generating oxidized dimeric products, and shows activity for oxidation of Ca‐lignosulfonate, generating vanillic acid as a low molecular weight product. The crystal structure of Ochrobactrum CueO (OcCueO) has been determined at 1.1 Å resolution (PDB: 6EVG), showing a four‐coordinate mononuclear type I copper center with ligands His495, His434 and Cys490 with Met500 as an axial ligand, similar to that of Escherichia coli CueO and bacterial azurin proteins, whereas fungal laccase enzymes contain a three‐coordinate type I copper metal center. A trinuclear type 2/3 copper cluster was modeled into the active site, showing similar structure to E. coli CueO and fungal laccases, and three solvent channels leading to the active site. Site‐directed mutagenesis was carried out on amino acid residues found in the solvent channels, indicating the importance for residues Asp102, Gly103, Arg221, Arg223, and Asp462 for catalytic activity. The work identifies a new bacterial multicopper enzyme with activity for lignin oxidation, and implicates a role for bacterial laccase‐like multicopper oxidases in some lignin‐degrading bacteria.

Diamond Keywords: Enzymes; Bacteria; Biodegradation

Subject Areas: Chemistry, Biology and Bio-materials

Instruments: I03-Macromolecular Crystallography

Added On: 05/04/2018 11:11

Discipline Tags:

Earth Sciences & Environment Biotechnology Climate Change Biochemistry Catalysis Chemistry Structural biology Engineering & Technology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)