Development of a Pichia pastoris cell factory for efficient production of germacrene A: a precursor of β-elemene

Abstract

β-Elemene, an active ingredient found in medicinal plants like turmeric and zedoary, is a sesquiterpene compound with antitumor activity against various cancers. However, its current mode of production through plant extraction suffers from low efficiency and limited natural resources. Recently, there has been an increased interest in establishing microbial cell factories to produce germacrene A, which can be converted to β-elemene by a one-step reaction in vitro. In this study, we constructed an engineered Pichia pastoris cell factory for producing germacrene A. We rerouted the fluxes towards germacrene A biosynthesis through the optimization of the linker sequences between germacrene A synthase (GAS) and farnesyl pyrophosphate synthase (ERG20), overexpression of important pathway genes (i.e., IDI1, tHMG1, and ACS), and multi-copy integration of related expression cassettes. In combination with medium optimization and bioprocess engineering, the final titer of germacrene A in a 1 L fermenter reached 1.9 g/L through fed-batch fermentation. This represents the first report on the production of germacrene A in P. pastoris and demonstrates its advantage in producing terpenoids and other value-added natural products.

Keywords

β-Elemene / Germacrene A / Pichia pastoris / Microbial cell factories / Metabolic engineering

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Funding

National Key Research and Development Program of China(2021YFC2103200)

China Postdoctoral Science Foundation(2022M712772)

Natural Science Foundation of Zhejiang Province(LR20B060003)

National Natural Science Foundation of China(22278361)

Fundamental Research Funds for the Central Universities(226-2023-00015)