Abstract
Valorization of all major lignocellulose components, including lignin, cellulose, and hemicellulose is critical for an economically viable bioeconomy. In most biochemical conversion approaches, the standard process separately upgrades sugar hydrolysates and lignin. Here, we present a new process concept based on an engineered microbe that could enable simultaneous upgrading of all lignocellulose streams, which has the ultimate potential to reduce capital cost and enable new metabolic engineering strategies. Pseudomonas putida is a robust microorganism capable of natively catabolizing aromatics, organic acids, and D-glucose. We engineered this strain to utilize D-xylose by tuning expression of a heterologous D-xylose transporter, catabolic genes xylAB, and pentose phosphate pathway (PPP) genes tal-tkt. We further engineered L-arabinose utilization via the PPP or an oxidative pathway. This resulted in a growth rate on xylose and arabinose of 0.32 h−1 and 0.38 h−1, respectively. Using the oxidative L-arabinose pathway with the PPP xylose pathway enabled D-glucose, D-xylose, and L-arabinose co-utilization in minimal medium using model compounds as well as real corn stover hydrolysate, with a maximum hydrolysate sugar consumption rate of 3.3 g/L/h. After modifying catabolite repression, our engineered P. putida simultaneously co-utilized five representative compounds from cellulose (D-glucose), hemicellulose (D-xylose, L-arabinose, and acetic acid), and lignin-related compounds (p-coumarate), demonstrating the feasibility of simultaneously upgrading total lignocellulosic biomass to value-added chemicals.
| Original language | English |
|---|---|
| Pages (from-to) | 62-71 |
| Number of pages | 10 |
| Journal | Metabolic Engineering |
| Volume | 62 |
| DOIs | |
| State | Published - Nov 2020 |
Bibliographical note
Publisher Copyright:© 2020 The Authors
Funding
This manuscript has been authored by UT-Battelle, LLC under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, world-wide license to publish or reproduce the published form of this manuscript, or allow others to do so, for United States Government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/doe-public-access-plan ). Oak Ridge National Laboratory is managed by UT-Battelle, LLC, for the U.S. Department of Energy under contract DE-AC05-00OR22725. This work was also partially authored by Alliance for Sustainable Energy, LLC, the manager and operator of the National Renewable Energy Laboratory for the U.S. Department of Energy under Contract No. DE-AC36-08GO28308, and Battelle, the manager and operator of the Pacific Northwest National Laboratory for the U.S. Department of Energy under Contract No DE-AC05-76RLO1830. Research was in part sponsored by the Laboratory Directed Research and Development Program of Oak Ridge National Laboratory , Project ID 7866 . We also acknowledge funding from the U.S. Department of Energy Office of Energy Efficiency and Renewable Energy the Bioenergy Technologies Office via the Agile BioFoundry project, Metabolic Engineering for Lignin Conversion project, and the Biological Lignin Valorization project. This work was supported in part by the U.S. Department of Energy, Office of Science, Office of Workforce Development for Teachers and Scientists (WDTS) under the Science Undergraduate Laboratory Internship (SULI) program to KG. We thank Annette De Capite and E. Anne Hatmaker for helpful discussions on HPLC analysis of sugars and analysis of genome resequencing, respectively. We thank Austin Carroll for determining the ratio of optical densities between the plate reader and Biochrom spectrophotometer. Oak Ridge National Laboratory is managed by UT-Battelle, LLC, for the U.S. Department of Energy under contract DE-AC05-00OR22725. This work was also partially authored by Alliance for Sustainable Energy, LLC, the manager and operator of the National Renewable Energy Laboratory for the U.S. Department of Energy under Contract No. DE-AC36-08GO28308, and Battelle, the manager and operator of the Pacific Northwest National Laboratory for the U.S. Department of Energy under Contract No DE-AC05-76RLO1830. Research was in part sponsored by the Laboratory Directed Research and Development Program of Oak Ridge National Laboratory, Project ID 7866. We also acknowledge funding from the U.S. Department of Energy Office of Energy Efficiency and Renewable Energy the Bioenergy Technologies Office via the Agile BioFoundry project, Metabolic Engineering for Lignin Conversion project, and the Biological Lignin Valorization project. This work was supported in part by the U.S. Department of Energy, Office of Science, Office of Workforce Development for Teachers and Scientists (WDTS) under the Science Undergraduate Laboratory Internship (SULI) program to KG. We thank Annette De Capite and E. Anne Hatmaker for helpful discussions on HPLC analysis of sugars and analysis of genome resequencing, respectively. We thank Austin Carroll for determining the ratio of optical densities between the plate reader and Biochrom spectrophotometer.
| Funders | Funder number |
|---|---|
| Office of Workforce Development for Teachers | |
| U.S. Department of Energy Office of Energy Efficiency and Renewable Energy the Bioenergy Technologies Office | |
| U.S. Department of Energy | DE-AC05-00OR22725 |
| Battelle | DE-AC05-76RLO1830 |
| Office of Science | |
| Office of Energy Efficiency and Renewable Energy | |
| Workforce Development for Teachers and Scientists | |
| Oak Ridge National Laboratory | |
| National Renewable Energy Laboratory | DE-AC36-08GO28308 |
Keywords
- Biomass valorization
- Hydrolysate
- Lignin
- Lignocellulose
- Metabolic engineering
- Pseudomonas putida
- Substrate co-utilization
