One approach to achieve continuous overproduction of lipids in microalgal "cell factories" relies upon depletion or removal of nutrients that act as competing electron sinks (e. g., nitrate and sulfate). However, this strategy can only be effective for bioenergy applications if lipid is synthesized primarily de novo (from CO 2 fixation) rather than from the breakdown and interconversion of essential cellular components. In the marine diatom, Phaeodactylum tricornutum, it was determined, using 13C-bicarbonate, that cell growth in nitrate (NO 3 -)-deprived cultures resulted predominantly in de novo lipid synthesis (60 % over 3 days), and this new lipid consisted primarily of triacylglycerides (TAGs). Nearly complete preservation of 12C occurred in all previously existing TAGs in NO 3 --deprived cultures and thus, further TAG accumulation would not be expected from inhibition of TAG lipolysis. In contrast, both high turnover and depletion of membrane lipids, phosphatidylcholines (PCs), were observed in NO 3 --deprived cultures (both the headgroups and fatty acid chains), while less turnover was observed in NO 3 - replete cultures. Liquid chromatography-tandem mass spectrometry mass spectra and 13C labeling patterns of PC headgroups provided insight into lipid synthesis in marine diatoms, including suggestion of an internal pool of glycine betaine that feeds choline synthesis. It was also observed that 16C fatty acid chains incorporated into TAGs and PCs contained an average of 14 13C carbons, indicating substantial incorporation of 13C-bicarbonate into fatty acid chains under both nutrient states.
Bibliographical noteFunding Information:
Acknowledgments This work was funded by the Air Force Office of Scientific Research, grant # FA9550-05-1-0365. The LC/MS instrument was obtained through National Center for Research Resources (NIH) (grant # RR021120). EHB was additionally funded by the Busch-Waksman Postdoctoral Fellowship, and NBB was funded by the Sustainable Fuels NSF-IGERT program (award #0903675). MF was funded by a gift from James Gibson. We would also like to thank Char Fuller and Kevin Wyman. Elemental analysis was performed by Marshall Otter at the Marine Biological Laboratory Ecosystems Center, Woods Hole, MA, USA.
- De novo lipid biosynthesis
- Fatty acid metabolism
- Phaeodactylum tricornutum