Multi-omic elucidation of aromatic catabolism in adaptively evolved Rhodococcus opacus

• Published in: Metabolic engineering 2018-09, Vol.49 (C), p.69-83
• Main Authors: Henson, William R., Campbell, Tayte, DeLorenzo, Drew M., Gao, Yu, Berla, Bertram, Kim, Soo Ji, Foston, Marcus, Moon, Tae Seok, Dantas, Gautam
• Format: Article
• Language: English
• Subjects: Adaptation; Aromatic tolerance; bacteria; Bacterial Proteins - biosynthesis; Bacterial Proteins - genetics; biorefining; Biotechnology & Applied Microbiology; biotransformation; byproducts; depolymerization; Directed Molecular Evolution; enzyme activity; Funneling pathway; gene expression regulation; gene targeting; genes; Genomics; guaiacol; Hydrocarbons, Aromatic - metabolism; Lignin; lignocellulose; metabolism; microbial growth; mutants; mutation; phenol; profitability; Redox metabolism; redox reactions; Rhodococcus - genetics; Rhodococcus - metabolism; Rhodococcus opacus; sequence analysis; superoxide dismutase; transcriptomics
• ISSN: 1096-7176; 1096-7184
• DOI: 10.1016/j.ymben.2018.06.009

Lignin utilization has been identified as a key factor in biorefinery profitability. However, lignin depolymerization generates heterogeneous aromatic mixtures that inhibit microbial growth and the conversion of lignocellulose to biochemicals. Rhodococcus opacus is a promising aromatic-catabolizing, oleaginous bacterium, but mechanisms for its aromatic tolerance and utilization remain undercharacterized. To better understand these mechanisms, we adaptively evolved R. opacus for improved utilization of 32 combinations of diverse aromatic compounds. Evolved R. opacus mutants showed up to 1900% growth improvement in the utilization of phenol, guaiacol, 4-hydroxybenzoate, vanillate, and benzoate compared to the wild-type strain. Whole genome sequencing revealed several redox-related genes with mutations shared across multiple adapted mutants. PVHG6, the mutant with the most improved growth on a mixture of multiple aromatic compounds, showed 56% lower superoxide dismutase activity than the wild-type strain, suggesting that redox reactions are important for aromatic tolerance and utilization. Comparative transcriptomics revealed by-product detoxification pathways and five aromatic funneling pathways that were upregulated in response to specific aromatic compounds. Gene knockout experiments confirmed the two degradation routes of the β-ketoadipate pathway for five aromatic compounds. These results provide an improved understanding of aromatic bioconversion and facilitate development of R. opacus as a biorefinery host. •Generation of adapted Rhodococcus opacus strains for increased aromatic tolerance and utilization.•Comparative genomics of adapted R. opacus strains to identify mutations linked to aromatic tolerance.•Transcriptomic elucidation of aromatic utilization pathways in R. opacus.