%0 Journal Article
%T Engineering Limonene and Bisabolene Production in Wild Type and a Glycogen-Deficient Mutant of Synechococcus sp. PCC 7002
%A Fiona K. Davies
%A Alexander S. Beliaev
%A Matthew C. Posewitz
%J Frontiers in Bioengineering and Biotechnology
%D 2014
%I Frontiers Media
%R 10.3389/fbioe.2014.00021
%X The plant terpenoids limonene (C10H16) and ¦Á-bisabolene (C15H24) are hydrocarbon precursors to a range of industrially relevant chemicals. High-titer microbial synthesis of limonene and ¦Á-bisabolene could pave the way for advances in in vivo engineering of tailor-made hydrocarbons, and production at commercial scale. We have engineered the fast-growing unicellular euryhaline cyanobacterium Synechococcus sp. PCC 7002 to produce yields of 4 mg L£¿1 limonene and 0.6 mg L£¿1 ¦Á-bisabolene through heterologous expression of the Mentha spicata l-limonene synthase or the Abies grandis (E)-¦Á-bisabolene synthase genes, respectively. Titers were significantly higher when a dodecane overlay was applied during culturing, suggesting either that dodecane traps large quantities of volatile limonene or ¦Á-bisabolene that would otherwise be lost to evaporation, and/or that continuous product removal in dodecane alleviates product feedback inhibition to promote higher rates of synthesis. We also investigate limonene and bisabolene production in the ¦¤glgC genetic background, where carbon partitioning is redirected at the expense of glycogen biosynthesis. The Synechococcus sp. PCC 7002 ¦¤glgC mutant excreted a suite of overflow metabolites (¦Á-ketoisocaproate, pyruvate, ¦Á-ketoglutarate, succinate, and acetate) during nitrogen-deprivation, and also at the onset of stationary growth in nutrient-replete media. None of the excreted metabolites, however, appeared to be effectively utilized for terpenoid metabolism. Interestingly, we observed a 1.6- to 2.5-fold increase in the extracellular concentration of most excreted organic acids when the ¦¤glgC mutant was conferred with the ability to produce limonene. Overall, Synechococcus sp. PCC 7002 provides a highly promising platform for terpenoid biosynthetic and metabolic engineering efforts.
%K terpenoids
%K limonene
%K bisabolene
%K cyanobacteria
%K glycogen
%K glgC
%K metabolic sink
%U http://www.frontiersin.org/Journal/10.3389/fbioe.2014.00021/abstract