Designing obligate consortia with carbon-nitrogen exchange using new-to-nature exchange molecules

The sustainable provision of nitrogen to agriculture is one of the major challenges facing humanity in the 21st century. A range of biotechnological solutions to replace the highly polluting Haber-Bosch process have been proposed. In this thesis, I introduce an approach using metabolic engineering to design novel nutrient-exchange pathways between crop plants and soil-dwelling nitrogen-fixing microbiota. A consortium of the nitrogen-fixing soil bacterium Azotobacter vinelandii with the cyanobacterium Synechocystis sp. PCC 6803, a model photoautotrophic organism, is developed both in vitro and implemented as genome-scale metabolic model in silico. In addition to the total set of compounds naturally present in either consortium partner, rationally designed pathways based on the Anabaena sp. PCC 7120 nitrogen-carbon exchange system were evaluated. A high-throughput unbiased approach utilising bioretrosynthesis was also employed to evaluate a broad nutrient-exchange compound design space spanning 1,292,154 compounds including those not native to the consortium partners. Overall, 14,574 unique nutrient-exchange pathways were identified as capable of transferring fixed nitrogen from A. vinelandii to Synechocystis 6803, and fixed carbon from Synechocystis 6803 to A. vinelandii in return while supporting growth of both partners, thus enabling the establishment of a nutrient-exchange symbiosis.