(567am) Metabolic Engineering of Cyanobacteria for Biodiesel Feedstock

Ruffing, A., Sandia National Laboratories

With rising concerns of energy sustainability and climate change, metabolic engineering strategies must be applied to advance the development of second generation biofuels. Major challenges in biodiesel production include 1) maximizing the production rate to meet the high demand of current energy consumption and 2) developing low-energy and cost-effective methods for isolation and purification of the biodiesel. Photosynthetic microorganisms offer a promising solution to these challenges, while at the same time, addressing growing environmental concerns through CO2 mitigation. Algae, natural photosynthetic oil producers, are the focus of most biodiesel research efforts, and little attention has been given to other photosynthetic microorganisms, particularly cyanobacteria. Cyanobacteria do not naturally produce oil like algae; however, there are other advantages to using cyanobacteria for biodiesel feedstock production. Unlike algae, cyanobacteria have well-established methods for genetic engineering, as evidenced by the genetic engineering of cyanobacteria for the production of first generation biofuels including ethanol and butanol. Furthermore, cyanobacteria will naturally secrete free fatty acids (FFA), a biodiesel precursor, into the extracellular media, simplifying downstream product isolation. These attributes motivate the investigation of cyanobacteria as a potential source for biodiesel feedstock.

In this study, the cyanobacterium Synechococcus elongatus PCC 7942 is engineered for the production of FFA. The metabolic engineering strategy involves the elimination of FFA metabolism, removal of feedback inhibition of the fatty acid synthesis pathway, improving carbon flux through the fatty acid and photosynthetic pathways, and elimination of competing pathways. The strategy is unique from other efforts involving E. coli and Synechocystis sp., in that, novel genes are cloned from the green alga, Chalmydomonas reinhardtii. As algae naturally produce high levels of fatty acid for triacylglyceride (TAG) production, the lipid-producing genes of C. reinhardtii are likely to have higher activity compared to the corresponding homologs in E. coli. While additional engineering is needed to develop a production strain, the results illustrate the potential of cyanobacteria for the production of second generation biofuels.