Abstract
BackgroundLignin is a recalcitrant aromatic polymer that is a potential feedstock for renewable fuel and chemical production. Rhodococcus opacus PD630 is a promising strain for the biological upgrading of lignin due to its ability to tolerate and utilize lignin-derived aromatic compounds. To enhance its aromatic tolerance, we recently applied adaptive evolution using phenol as a sole carbon source and characterized a phenol-adapted R. opacus strain (evol40) and the wild-type (WT) strain by whole genome and RNA sequencing. While this effort increased our understanding of the aromatic tolerance, the tolerance mechanisms were not completely elucidated.ResultsWe hypothesize that the composition of lipids plays an important role in phenol tolerance. To test this hypothesis, we applied high-resolution mass spectrometry analysis to lipid samples obtained from the WT and evol40 strains grown in 1 g/L glucose (glucose), 0.75 g/L phenol (low phenol), or 1.5 g/L phenol (high phenol, evol40 only) as a sole carbon source. This analysis identified > 100 lipid species of mycolic acids, phosphatidylethanolamines (PEs), phosphatidylinositols (PIs), and triacylglycerols. In both strains, mycolic acids had fewer double bond numbers in phenol conditions than the glucose condition, and evol40 had significantly shorter mycolic acid chain lengths than the WT strain in phenol conditions. These results indicate that phenol adaptation affected mycolic acid membrane composition. In addition, the percentage of unsaturated phospholipids decreased for both strains in phenol conditions compared to the glucose condition. Moreover, the PI content increased for both strains in the low phenol condition compared to the glucose condition, and the PI content increased further for evol40 in the high phenol condition relative to the low phenol condition.ConclusionsThis work represents the first comprehensive lipidomic study on the membrane of R. opacus grown using phenol as a sole carbon source. Our results suggest that the alteration of the mycolic acid and phospholipid membrane composition may be a strategy of R. opacus for phenol tolerance.
Highlights
Lignin is a recalcitrant aromatic polymer that is a potential feedstock for renewable fuel and chemical production
R. opacus is a promising host for the biological conversion of lignin-derived aromatic compounds into valuable products, but the role of its lipid metabolism in aromatic tolerance is not well understood
Mycolic acids had fewer double bonds during growth using phenol compared to glucose for both strains, and mycolic acid chain lengths in evol40, but not in the WT strain, were significantly shorter in phenol conditions compared to the glucose condition, showing the similarity and difference in phenol responses between the two strains
Summary
Lignin is a recalcitrant aromatic polymer that is a potential feedstock for renewable fuel and chemical production. To enhance its aromatic tolerance, we recently applied adap‐ tive evolution using phenol as a sole carbon source and characterized a phenol-adapted R. opacus strain (evol40) and the wild-type (WT) strain by whole genome and RNA sequencing. While this effort increased our understanding of the aromatic tolerance, the tolerance mechanisms were not completely elucidated. Efforts are underway to improve the aromatic tolerance of microbes used for fermentation of lignocellulose-derived sugars and to develop strains that have aromatic degradation pathways to convert ligninderived aromatics into valuable products [14,15,16,17,18]. While many studies have focused on characterizing the lipid metabolism of R. opacus for the goal of improving TAG accumulation [29,30,31,32], it is unknown whether, or how, the lipid metabolism plays a role in the tolerance of aromatic compounds
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