Engineering Cyanobacteria for High-Yield Photosynthetic Isoprene Production With Long-Term Phenotypic Stability.

Abstract

In light of the looming climate crisis, a key cornerstone for a sustainable bioeconomy is photosynthetic production of chemicals and fuels from CO2 and water, powered by sunlight. Isoprene is a five-carbon volatile hydrocarbon with industrial use as a feedstock for rubber production and chemical synthesis and is, at present, generated from crude oil sources. It is, however, possible to produce isoprene using photoautotrophic microorganisms such as cyanobacteria, heterologously expressing the plant enzyme isoprene synthase. Here, we have employed diverse metabolic engineering strategies to develop new strains of the unicellular cyanobacterium Synechocystis sp. PCC 6803 capable of high-level isoprene production from CO2, and have characterised the resulting strains regarding growth, stability and productivity. The new isoprene-producing strains address several challenges in large-scale photobiotechnological production such as genetic and metabolic stability, biosafety and thermotolerance. Moreover, we tested photosynthetic terpenoid production in photobioreactors under process-relevant conditions, achieving the highest volumetric productivities reported so far for a terpene or terpenoid product in cyanobacteria, reaching 148 mg L-1 day-1. Furthermore, we identified and discussed process limitations, laying the foundation for further strain and process engineering towards highly efficient and stable cyanobacterial hydrocarbon production at large scale without selection pressure.