1Synthetic and Systems Biology Innovation Hub, Texas A&M University, College Station, Texas 77843, USA
2Department of Plant Pathology and Microbiology, Texas A&M University, College Station, Texas 77843, USA
3Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, State Key Laboratory of Applied Microbiology Southern China, Institute of Microbiology, Guangdong Academy of Science, China
4Section of Molecular Biology, University of California San Diego, La Jolla, CA 92093, USA
5Department of Microbiology, Miami University, Oxford, Ohio 45056, USA
| Received 04 Nov 2021 |
Accepted 11 Jan 2022 |
Published 09 Feb 2022 |
Photosynthetic terpene production represents one of the most carbon and energy-efficient routes for converting CO2 into hydrocarbon. In photosynthetic organisms, metabolic engineering has led to limited success in enhancing terpene productivity, partially due to the low carbon partitioning. In this study, we employed systems biology analysis to reveal the strong competition for carbon substrates between primary metabolism (e.g., sucrose, glycogen, and protein synthesis) and terpene biosynthesis in Synechococcus elongatus PCC 7942. We then engineered key “source” and “sink” enzymes. The “source” limitation was overcome by knocking out either sucrose or glycogen biosynthesis to significantly enhance limonene production via altered carbon partitioning. Moreover, a fusion enzyme complex with geranyl diphosphate synthase (GPPS) and limonene synthase (LS) was designed to further improve pathway kinetics and substrate channeling. The synergy between “source” and “sink” achieved a limonene titer of 21.0 mg/L. Overall, the study demonstrates that balancing carbon flux between primary and secondary metabolism can be an effective approach to enhance terpene bioproduction in cyanobacteria. The design of “source” and “sink” synergy has significant potential in improving natural product yield in photosynthetic species.
