A manuscript by Leila Pazouki, Hamid Rajabi Memari, Astrid Kännaste, Rudolf Bichele and Ülo Niinemets characterizing a sesquiterpene synthase, germacrene A synthase, has been accepted for publication in a broad spectrum plant science journal Frontiers in Plant Science. This study is the first paper from our team reporting genetic engineering of terpenoid synthases, and thus, constitutes an important milestone of our recent efforts to link production and emission of plant volatiles to gene-level controls. This line of work was started when Prof. Rajabi Memari from Shahid Chamran University, Ahvaz, Iran visited our team for three months back in 2012, and initiated the cloning of key terpenoid synthases as well as provided a lot of stimulating insight into possible engineering of plant secondary metabolism.
Sesquiterpenes are widely distributed volatile terpenoids consisting of 3 isoprene residues (C15). They are characteristically emitted from flowers and serve as attractants to pollinators as well as repellents against nectar thieves. In many plant species, leaf sesquiterpene emissions play major roles in direct and indirect chemical defense against pathogens and herbivores. Furthermore, oxygenated sesquiterpene derivatives can serve as important non-volatile defenses. There is overall limited information on sesquiterpenoid synthases, the enzymes that are responsible for formation of the rich class of C15 terpenoids in plants. Among sesquiterpene synthases, particularly scarce is the information on germacrene synthases. Volatile germacrenes themselves fulfill relevant roles as infochemicals in plant-to-plant and plant-to-insect communications, but they also play a central role in the formation of different sesquiterpene derivatives, in particular, sesquiterpene lactones. Many of these lactones are characterized by antifungal and antibacterial properties and can protect against pests and herbivores. Due to their medicinal properties, they are considered a promising class of compounds for pharmacological industry. We have used the important medicinal plant yarrow (Achillea millefolium) for this analysis, and therefore, we believe that in addition to improving the basic knowledge of terpenoid synthesis, the results of this study could also have a major practical value in engineering terpenoid metabolism in medicinal plants.
This paper focuses primarily on the isolation of germacrene A synthase (GAS) gene from A. millefolium, synthesis of recombinant protein from this gene and in vitro enzyme functional characterization. The results of this study demonstrate that A. millefolium GAS enzyme is a multi-substrate cytosolic enzyme catalyzing formation of germacrene A when C15 substrate is available, but also acyclic and cyclic monoterpenes (C10) when C10 substrate is available. Although sesquiterpenes are typically formed in cytosol and monoterpenes in plastids, this finding suggests a hugely exciting possibility of monoterpene synthesis in cytosol. This can be relevant under physiological conditions in aromatic plants that are known to accumulate volatiles in their foliar storage structures. Such an appealing opportunity of cytosolic monoterpene synthesis will await further experimental testing in vivo. To our knowledge, this is the first report of gene cloning and functional characterization of germacrene A synthase gene in A. millefolium, and multi-substrate use of GAS enzymes.
Full citation: Pazouki, L., Rajabi Memari, H., Kännaste, A., Bichele, R., & Niinemets, Ü. (2015). Germacrene A Synthase in Yarrow (Achillea millefolium) Is an Enzyme with Mixed Substrate Specificity: Gene Cloning, Functional Characterization and Expression Analysis. Frontiers in Plant Science, 6, 111. (link to full text; open access)