Further engineering of R. toruloides for the production of terpenes from lignocellulosic biomass
Autor: | Anna Fuhrmann, Yuqian Gao, Junko Yaegashi, Jeffrey M. Skerker, Katharina Seibel, Jan-Philip Prahl, Gina M. Geiselman, Bobbie-Jo M. Webb-Robertson, Jeremy Zucker, Mary Bao Tran-Gyamfi, Deepti Tanjore, James Kirby, Veronica T. Benites, Carrie D. Nicora, Nathalie Munoz, Kristin E. Burnum-Johnson, Edward E. K. Baidoo, Xun Zhuang, Eric R. Sundstrom, Jon K. Magnuson, John M. Gladden, Joonhoon Kim |
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Rok vydání: | 2021 |
Předmět: |
Mevalonate pathway
Α Geranylgeranyl pyrophosphate Phosphomevalonate kinase 1 8-Cineole Rhodosporidium toruloides Lignocellulosic biomass Management Monitoring Policy and Law Applied Microbiology and Biotechnology Hydrolysate Industrial Biotechnology Metabolic engineering 03 medical and health sciences chemistry.chemical_compound TP315-360 bisabolene 030304 developmental biology 8-Cineole 0303 health sciences Eucalyptol biology 030306 microbiology Renewable Energy Sustainability and the Environment Chemistry Research Α-bisabolene Rhodotorula Mevalonate kinase Chemical Engineering Fuel Isoprenoids biology.organism_classification General Energy Biochemistry biology.protein TP248.13-248.65 Biotechnology |
Zdroj: | Biotechnology for biofuels, vol 14, iss 1 Biotechnology for Biofuels, Vol 14, Iss 1, Pp 1-16 (2021) Biotechnology for Biofuels |
ISSN: | 1754-6834 |
Popis: | Background Mitigation of climate change requires that new routes for the production of fuels and chemicals be as oil-independent as possible. The microbial conversion of lignocellulosic feedstocks into terpene-based biofuels and bioproducts represents one such route. This work builds upon previous demonstrations that the single-celled carotenogenic basidiomycete, Rhodosporidium toruloides, is a promising host for the production of terpenes from lignocellulosic hydrolysates. Results This study focuses on the optimization of production of the monoterpene 1,8-cineole and the sesquiterpene α-bisabolene in R. toruloides. The α-bisabolene titer attained in R. toruloides was found to be proportional to the copy number of the bisabolene synthase (BIS) expression cassette, which in turn influenced the expression level of several native mevalonate pathway genes. The addition of more copies of BIS under a stronger promoter resulted in production of α-bisabolene at 2.2 g/L from lignocellulosic hydrolysate in a 2-L fermenter. Production of 1,8-cineole was found to be limited by availability of the precursor geranylgeranyl pyrophosphate (GPP) and expression of an appropriate GPP synthase increased the monoterpene titer fourfold to 143 mg/L at bench scale. Targeted mevalonate pathway metabolite analysis suggested that 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGR), mevalonate kinase (MK) and phosphomevalonate kinase (PMK) may be pathway bottlenecks are were therefore selected as targets for overexpression. Expression of HMGR, MK, and PMK orthologs and growth in an optimized lignocellulosic hydrolysate medium increased the 1,8-cineole titer an additional tenfold to 1.4 g/L. Expression of the same mevalonate pathway genes did not have as large an impact on α-bisabolene production, although the final titer was higher at 2.6 g/L. Furthermore, mevalonate pathway intermediates accumulated in the mevalonate-engineered strains, suggesting room for further improvement. Conclusions This work brings R. toruloides closer to being able to make industrially relevant quantities of terpene from lignocellulosic biomass. |
Databáze: | OpenAIRE |
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