Identification and Characterization of a Novel Plasmid-Encoded Laccase-Like Multicopper Oxidase from Ochrobactrum sp. BF15 Isolated from an On-Farm Bio-Purification System
1IBBM - Instituto de Biotecnología y Biología Molecular, CONICET - Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, Calles 47 y 115 (1900) La Plata, Argentina
2Department of Biotechnology and Life Sciences, University of Insubria, via J.H. Dunant 3, 21100 Varese, Italy
3Department of Food Science and Technology, Biotechnical Faculty, University of Ljubljana, Vecna pot 111, 1000 Ljubljana, Slovenia
Received: 25 March 2021
Accepted: 20 October 2021
laccase-like multicopper oxidases, Ochrobactrum, biopurification system, plasmid, biodegradation, heterologous expression
Research background. In recent decades, laccases (p-diphenol-dioxygen oxidoreductases; EC 184.108.40.206) have attracted the attention of researchers due to their wide range of biotechnological and industrial applications. Laccases can oxidize a variety of organic and inorganic compounds, making them suitable as biocatalysts in biotechnological processes. Even though the most traditionally used laccases in the industry are of fungal origin, bacterial laccases have shown an enormous potential given their ability to act on several substrates and in multiple conditions. The present study aims to characterize a plasmid-encoded laccase-like multicopper oxidase (LMCO) from Ochrobactrum sp. BF15, a bacterial strain previously isolated from polluted soil.
Experimental approach. We used in silico profile hidden Markov models to identify novel laccase-like genes in Ochrobactrum sp. BF15. For laccase characterization, we performed heterologous expression in Escherichia coli, purification and activity measurement on typical laccase substrates.
Results and conclusions. Profile hidden Markov models allowed us to identify a novel LMCO, named Lac80. In silico analysis of Lac80 revealed the presence of three conserved copper oxidase domains characteristic of three-domain laccases. We successfully expressed Lac80 heterologously in E. coli, allowing us to purify the protein for further activity evaluation. Of thirteen typical laccase substrates tested, Lac80 showed lower activity on 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), pyrocatechol, pyrogallol and vanillic acid, and higher activity on 2,6-dimethoxyphenol.
Novelty and scientific contribution. Our results show Lac80 as a promising laccase for use in industrial applications. The present work shows the relevance of bacterial laccases and highlights the importance of environmental plasmids as valuable sources of new genes encoding enzymes with potential use in biotechnological processes.
|*Corresponding author:||+542214250497 ext. 31|
|+42214223409 ext. 56|
#These authors contributed equally to this study.