Engineering the methylerythritol phosphate pathway in cyanobacteria for photosynthetic isoprene production from CO2

Isoprene, a key building block of synthetic rubber, is currently produced entirely from petrochemical sources. Previous metabolic engineering efforts, centered on heterologous expression of the mevalonate pathway, have resulted in isoprene production by microbial fermentation of sugars. To produce isoprene directly from CO2, we engineered the isoprene biosynthetic pathway in the cyanobacterium Synechococcus elongatus, with guidance provided by dynamic flux analysis and metabolite profiling. The methylerythritol phosphate (MEP) pathway was selected for cyanobacterial isoprene synthesis based on comparison of carbon efficiency and the precursor driving force between the MEP pathway and the mevalonate pathway. Plant-derived isoprene synthases with high activities were introduced, followed by increasing the ratio of dimethylallyl pyrophosphate to isopentenyl pyrophosphate (IPP) by overexpression of IPP isomerase (IDI), further improvement in isoprene production activity by direct fusion of IDI and isoprene synthase, and relieving an MEP pathway bottleneck identified by kinetic flux profiling. The engineered strain directed about 40% of photosynthetically fixed carbon toward the isoprene biosynthetic pathway, resulting in the production of 1.26 g L-1 of isoprene from CO2, which is a significant increase for terpenoid production by photoautotrophic microorganisms. The strains developed in this study can serve as platform hosts for photosynthetic production of diverse terpenoids from CO2.