Anaerobic butanol production driven by oxygen-evolving photosynthesis using the heterocyst-forming multicellular...

Cyanobacteria are oxygen-evolving photosynthetic bacteria. Established genetic manipulation methods and recently developed gene-regulation tools have enabled the photosynthetic conversion of carbon dioxide to biofuels and valuable chemicals in cyanobacteria, especially in unicellular cyanobacteria. However, the oxygen sensitivity of enzyme(s) introduced into cyanobacteria hampers productivity in some cases. Anabaena sp. PCC 7120 is a filamentous cyanobacterium consisting of a few hundred of vegetative cells, which perform oxygenic photosynthesis. Upon nitrogen deprivation, heterocysts, which are specialized cells for nitrogen fixation, are differentiated from vegetative cells at semiregular intervals. The micro-oxic environment within heterocysts protects oxygen-labile nitrogenase from oxygen. This study aimed to repurpose the heterocyst as a host for the production of chemicals with oxygen-sensitive enzymes under photosynthetic conditions. Herein, Anabaena strains expressing enzymes of 1-butanol synthetic pathway from the anaerobe Clostridium acetobutylicum within heterocysts were created. A strain that expressed a highly oxygen-sensitive Bcd/EtfAB complex produced 1-butanol even under photosynthetic conditions. Furthermore, the 1-butanol production per heterocyst cell of a butanol-producing Anabaena strain was fivefold higher than that per cell of unicellular cyanobacterium with the same set of 1-butanol synthetic pathway genes. Thus, our study showed the usefulness of Anabaena heterocysts as a chassis for anaerobic production driven by oxygen-evolving photosynthesis.