Engineering volatile isoprene production from the polyextremophilic red microalga Cyanidioschyzon merolae

Abstract

Isoprene is a 5-carbon volatile chemical monomer used in the production of various industrial products. Its current sourcing is primarily through petrochemical-based processes, but there is growing interest in developing more sustainable and bio-based production methods. This has led to the exploration of genetic engineering techniques to enable isoprene biosynthesis in photosynthetic microorganisms that convert carbon dioxide into their biomass and concomitantly, isoprene. This work focused on the use of the polyextremophilic red microalga, Cyanidioschyzon merolae strain 10D, as a candidate for heterologous isoprene synthesis. Through synthetic biology-enabled custom designed plasmid transformation and foreign transgene expression, a heterologous isoprene synthase (IspS) from sweet potato was successfully expressed from the nuclear genome of C. merolae and the translated protein was targeted to the algal plastid. In this work, transformant C. merolae strains were grown in a co-cultivation with related Galdieria sulphuraria, a mixotrophic Cyanidiophyceae, here fed with glucose to deliver respiratory CO2 to C. merolae when grown together in closed gas-chromatography vials. Through this strategy, the heterologous generation of isoprene could be quantified from transformants. It was determined that IspS-YFP fusions produced a higher isoprene yield compared to IspS-chloramphenicol selection marker fusions. The best isoprene-producing transformant was used to monitor the isoprene yield over 9-days of cultivation, reaching 231 mg/L culture on d9. This work represents a first-of-its-kind genetic engineering in red microalgae. The successful production of isoprene from this microalga could pave the way for the development of new and sustainable industrial applications for this commodity chemical.