Exploring and mapping the functional chemical space of amorphadiene synthase with non-canonical farnesyl diphosphate analogues

Malaria is a threat to approximately 40% of the world’s population. Artemisinin and its derivatives in combination therapy are the world’s number one treatment against malaria. Agricultural production remains the primary source of artemisinin, however alternative methods are constantly being seeked to combat the unreliable and fluctuating processes present today. In addition, reported cases of parasitic resistance to the combination therapies emphasise the urgent need to produce novel and active analogues to replace the existing first line treatments. Artemisinin is a terpenoid extracted from Artemisia annua. Terpenes constitute one of the largest families of natural products that boast a substantial degree of chemical diversity and functionality. The broad diversity of biological applications of terpenes has found several uses, particularly in medicine. Terpene synthases convert linear isoprenyl diphosphates into complex hydrocarbon structures that exhibit exquisite stereochemistry. Amorphadiene synthase converts farnesyl diphosphate to amorpha-4,11-diene. This bicyclic frame, with four stereocenters is the first precursor found in the biosynthesis of artemisinin. Terpene synthases are studied extensively by scientists, because of their ability to generate complex structures that require several synthetic steps to reproduce the same complexity. This project focuses on studying the promiscuity of amorphadiene synthase. By using the recombinant protein in vitro gives the advantage of feeding the enzyme with novel FDP analogues, in attempt to introduce new functionalities in the resulting amorphadiene structure. These added functional groups then serve as platforms to carry out further chemical steps or can be carried forward to the respective artemisinin derivative, and consequently tested for antimalarial activity. A library of FDP analogues were designed, synthesized and incubated with ADS to test for enzymatic activity. In addition to the search for new amorphadiene derivatives, one of the synthesized FDP analogues, 12-hydroxy FDP, was converted to dihydroartemisinic aldehyde. Dihydroartemisinic aldehyde is a precursor found further down the schematic pathway to artemisinin. A three-step chemo-enzymatic approach to producing dihydroartemisinic aldehyde was established, providing an alternative method to sourcing artemisinin

Concise synthesis of artemisinin from a farnesyl diphosphate analogue

Artemisinin is one of the most potent anti-malaria drugs and many often-lengthy routes have been developed for its synthesis. Amorphadiene synthase, a key enzyme in the biosynthetic pathway of artemisinin, is able to convert an oxygenated farnesyl diphosphate analogue directly to dihydroartemisinic aldehyde, which can be converted to artemisinin in only four chemical steps, resulting in an efficient synthetic route to the anti-malaria drug

Effiziente chemoenzymatische Synthese von dhydroartemisinaldehyd

Artemisinin aus der Pflanze Artemisia annua ist das wirkungsvollste Arzneimittel zur Behandlung von Malaria. Die Sesquiterpen-Cyclase Amorphadien-Synthase, ein Cytochrom-abhängiges CYP450 und eine Aldehyd-Reduktase wandeln in der Pflanze Farnesyl-Diphosphat (FDP) in Dihydroartemisinaldehyd (DHAAl) um, welches ein Schlüsselzwischenprodukt in der Biosynthese von Artemisinin und eine halbsynthetische Vorstufe in der chemischen Synthese des Arzneimittels ist. Hier berichten wir über einen chemoenzymatischen Prozess, der in der Lage ist, DHAAl nur mithilfe der Sesquiterpen-Synthase aus einem gezielt synthetisierten, hydroxylierten FDP-Derivat herzustellen. Dieser Prozess, der die natürliche Sequenz aus Cyclisierung von FDP und Oxidation des Kohlenwasserstoffs umkehrt, stellt eine wesentliche Verbesserung der DHAAl-Synthese dar und zeigt das Potenzial neuer Substrate in der Terpen-Synthase-katalysierten Synthese hochwertiger Naturstoffe auf

Harnessing enzyme plasticity for the synthesis of oxygenated sesquiterpenoids

8-Methoxy-γ-humulene, (E)-8-methoxy-β-farnesene, 12-methoxy-β-sesquiphellandrene and 12-methoxyzingiberene can be synthesised in amorphadiene synthase-catalysed reactions from 8- and 12-methoxyfarnesyl diphosphates due to the highly plastic yet tightly controlled carbocationic chemistry of this sesquiterpene cyclase