Abstract
Lignocellulosic biofuels are promising as sustainable alternative fuels, but lignin inhibits access of enzymes to cellulose, and by-products of lignin degradation can be toxic to cells. The fast growth, high efficiency and specificity of enzymes employed in the anaerobic litter deconstruction carried out by tropical soil bacteria make these organisms useful templates for improving biofuel production. The facultative anaerobe Enterobacter lignolyticus SCF1 was initially cultivated from Cloud Forest soils in the Luquillo Experimental Forest in Puerto Rico, based on anaerobic growth on lignin as sole carbon source. The source of the isolate was tropical forest soils that decompose litter rapidly with low and fluctuating redox potentials, where bacteria using oxygen-independent enzymes likely play an important role in decomposition. We have used transcriptomics and proteomics to examine the observed increased growth of SCF1 grown on media amended with lignin compared to unamended growth. Proteomics suggested accelerated xylose uptake and metabolism under lignin-amended growth, with up-regulation of proteins involved in lignin degradation via the 4-hydroxyphenylacetate degradation pathway, catalase/peroxidase enzymes, and the glutathione biosynthesis and glutathione S-transferase (GST) proteins. We also observed increased production of NADH-quinone oxidoreductase, other electron transport chain proteins, and ATP synthase and ATP-binding cassette (ABC) transporters. This suggested the use of lignin as terminal electron acceptor. We detected significant lignin degradation over time by absorbance, and also used metabolomics to demonstrate moderately significant decreased xylose concentrations as well as increased metabolic products acetate and formate in stationary phase in lignin-amended compared to unamended growth conditions. Our data show the advantages of a multi-omics approach toward providing insights as to how lignin may be used in nature by microorganisms coping with poor carbon availability.
Highlights
Lignocellulose is the most abundant biopolymer on earth, and a recent joint analysis by the DOE and USDA shows that there is sufficient national supply to make lignocellulosic biofuels technically feasible (Perlack, 2005)
Genetic, metabolic and proteomic analysis of lignin degradation is performed by comparing ligninamended xylose minimal media to unamended xylose minimal media, and lignin degradation mechanisms and pathways are inferred by differential gene expression and protein production
We further examined proteins and pathways likely associated with xylose degradation, lignin degradation, and dissimilatory lignin reduction to explore the ways in which SCF1 might be gaining a growth advantage in ligninamended compared to unamended cultivation conditions
Summary
Lignocellulose is the most abundant biopolymer on earth, and a recent joint analysis by the DOE and USDA shows that there is sufficient national supply to make lignocellulosic biofuels technically feasible (Perlack, 2005). Development of renewable, sustainable biofuels from plant feedstock material has emerged as a key goal of the US Department of Energy. Our primary goal is to improve biofuel production through better saccharification of pretreated feedstock (switchgrass) from pathways and enzymes of anaerobic bacterial lignin degraders. By characterizing anaerobic lignin degradation in the bacterium Enterobacter lignolyticus SCF1, we may be able to incorporate these enzymes and pathways into metabolic engineering of biofuel- and biodiesel-producing bacteria. These discoveries promise to provide insight to the natural processes of bacterial lignin decomposition
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