Improved Bioproduction of 1-Octanol Using Engineered Synechocystis sp. PCC 6803

1-Octanol has gained interest as a chemical precursor for both high and low value commodities including fuel, solvents, surfactants, and fragrances. By harnessing the power from sunlight and CO₂ as carbon source, cyanobacteria has recently been engineered for renewable production of 1-octanol. The productivity, however, remained low. In the present work, we report efforts to further improve the 1-octanol productivity. Different N-terminal truncations were evaluated on three thioesterases from different plant species, resulting in several candidate thioesterases with improved activity and selectivity toward octanoyl-ACP. The structure/function trials suggest that current knowledge and/or state-of-the art computational tools are insufficient to determine the most appropriate cleavage site for thioesterases in Synechocystis. Additionally, by tuning the inducer concentration and light intensity, we further improved the 1-octanol productivity, reaching up to 35% (w/w) carbon partitioning and a titer of 526 ± 5 mg/L 1-octanol in 12 days. Long-term cultivation experiments demonstrated that the improved strain can be stably maintained for at least 30 days and/or over ten times serial dilution. Surprisingly, the improved strain was genetically stable in contrast to earlier strains having lower productivity (and hence a reduced chance of reaching toxic product concentrations). Altogether, improved enzymes and environmental conditions (e.g., inducer concentration and light intensity) substantially increased the 1-octanol productivity. When cultured under continuous conditions, the bioproduction system reached an accumulative titer of >3.5 g/L 1-octanol over close to 180 days.