Technologies for Enhanced Photosynthetic Butanol Production

Synechocystis sp. PCC 6803 ( Synechocystis ) has been successfully established as a carbon-neutral platform for butanol production through extensive metabolic engineering strategies. Building on these advancements, this chapter highlights four innovative technologies developed and applied to further elucidate Synechocystis metabolism and enhance butanol productivity. First, a fast-growing cyanobacterium, Synechococcus sp. PCC 11901, was introduced as a novel platform for 3-methyl-1-butanol production. Although the achieved titers were lower compared to those obtained with Synechocystis , this finding underscores the potential for further optimization and improvement. Second, rather than exclusively targeting the 2-keto acid pathway, a CRISPRi system employing catalytically inactive dCas12a was successfully established to reprogram the broader cellular metabolic network. This approach aimed to identify and mitigate competing pathways that divert carbon flux away from the 2-keto acid pathway, thereby enhancing isobutanol and 3-methyl-1-butanol biosynthesis. Repression of 9 out of 15 targeted genes led to a significant increase in isobutanol and/or 3-methyl-1-butanol production, demonstrating the effectiveness of this strategy in metabolic pathway optimization. In addition to the molecular approaches to increase production, various processing techniques applied to growth cultures, increased volume removal and adsorption filters, resulted in significantly enhanced yields.