Substrate promiscuity of enzymes from the sesquiterpene biosynthetic pathways from Artemisia annua and Tanacetum parthenium allows for novel combinatorial sesquiterpene production

Abstract

The therapeutic properties of complex terpenes often depend on the stereochemistry of their functional groups. However, stereospecific chemical synthesis of terpenes is challenging. To overcome this challenge, metabolic engineering can be employed using enzymes with suitable stereospecific catalytic activity. Here we used a combinatorial metabolic engineering approach to explore the stereospecific modification activity of the Artemisia annua artemisinic aldehyde Δ11(13) double bond reductase2 (AaDBR2) on products of the feverfew sesquiterpene biosynthesis pathway (GAS, GAO, COS and PTS). This allowed us to produce dihydrocostunolide and dihydroparthenolide. For dihydroparthenolide we demonstrate that the preferred order of biosynthesis of dihydroparthenolide is by reduction of the exocyclic methylene of parthenolide, rather than through C4-C5 epoxidation of dihydrocostunolide. Moreover, we demonstrate a promiscuous activity of feverfew CYP71CB1 on dihydrocostunolide and dihydroparthenolide for the production of 3β-hydroxy-dihydrocostunolide and 3β-hydroxy-dihydroparthenolide, respectively. Combined, these results offer new opportunities for engineering novel sesquiterpene lactones with potentially improved medicinal value.

Synthetic biology strategies for microbial biosynthesis of plant natural products

“…Neither of these two products had previously been identified in the native plant host, P. somniferum 65 . The combinatorial space around terpene biosynthetic pathways has been similarly probed using transient expression in Nicotiana benthamiana to generate novel sesquiterpenoids derived from the parthenolide biosynthetic pathway from feverfew ( Tanacetum parthenium ) 83 . This space can be further expanded through the introduction of analogues of the native pathway enzymes.…”

Artemisinins in Combating Viral Infections Like SARS-CoV-2, Inflammation and Cancers and Options to Meet Increased Global Demand

“… A schematic representation of the artemisinin biosynthetic pathway in Artemisia annua L. The precursors for artemisinin biosynthesis (DMAPP and IPP) are produced in the cytosolic Mevalonate (MVA) pathway and the plastidal 2-C-Methylerythritol 4-phosphate pathway (MEP) pathway, respectively ( Dewick, 2009 ; Vranová et al, 2013 ; Beyraghdar Kashkooli et al, 2019 ). IPP+DMAPP are converted to FPP, which is the general precursor for sesquiterpenes ( Fuentes et al, 2016 ).…”

“…Artemisinin is an oxygenated sesquiterpene lactone, mostly produced in glandular trichomes (GTs) of the medicinal plant Artemisia annua L. ( Tang et al, 2014 ; Wang et al, 2016 ; Beyraghdar Kashkooli et al, 2018 , 2019 ). Artemisinin and related compounds derived from the biosynthetic pathway ( Figure 1 ) have been shown to be effective against malaria caused by the Plasmodium spp.…”

Synthetic biology of plant natural products: From pathway elucidation to engineered biosynthesis in plant cells

“…benthamiana , many of which had not been reported previously ( Reed et al., 2017 ). Kashkooli et al. (2019 ) used a combinatorial metabolic engineering approach to generate costunolide and parthenolide derivatives.…”