Improved lipid production via fatty acid biosynthesis and free fatty acid recycling in engineered sp. PCC 6803.

BACKGROUND

Cyanobacteria are potential sources for third generation biofuels. Their capacity for biofuel production has been widely improved using metabolically engineered strains. In this study, we employed metabolic engineering design with target genes involved in selected processes including the fatty acid synthesis (a cassette of , , and encoding acetyl-CoA carboxylase, ACC), phospholipid hydrolysis ( encoding lipase A), alkane synthesis ( encoding acyl-ACP reductase, AAR), and recycling of free fatty acid (FFA) ( encoding acyl-acyl carrier protein synthetase, AAS) in the unicellular cyanobacterium sp. PCC 6803.

RESULTS

To enhance lipid production, engineered strains were successfully obtained including an -overexpressing strain (OX), an -overexpressing strain with knockout (OX/KO), and an -overexpressing strain with knockout (OX/KO). All engineered strains grew slightly slower than wild-type (WT), as well as with reduced levels of intracellular pigment levels of chlorophyll and carotenoids. A higher lipid content was noted in all the engineered strains compared to WT cells, especially in OX, with maximal content and production rate of 34.5% w/DCW and 41.4 mg/L/day, respectively, during growth phase at day 4. The OX/KO strain, with an impediment of phospholipid hydrolysis to FFA, also showed a similarly high content of total lipid of about 32.5% w/DCW but a lower production rate of 31.5 mg/L/day due to a reduced cell growth. The knockout interruptions generated, upon a downstream flow from intermediate fatty acyl-ACP, an induced unsaturated lipid production as observed in OX/KO and OX/KO strains with 5.4% and 3.1% w/DCW, respectively.

CONCLUSIONS

Among the three metabolically engineered strains, the OX with enhanced free fatty acid recycling had the highest efficiency to increase lipid production.