Production of the antimalarial drug precursor artemisinic acid in engineered yeast

Abstract

Drug-resistant strains of the malaria parasite are widespread, and as a result mortality due to malaria has increased significantly in recent years. Artemisinin, isolated from the herb Artemisia annua (sweet wormwood), is one drug that shows a high efficacy in killing multi-resistant strains of the parasite. The drug is extremely expensive, and high demand has led to a shortage of artemisinin, available only by extraction from the plant source. Ro et al. now report the development of a yeast strain engineered to carry a cytochrome P450 monooxygenase from A. annua that can produce the drug precursor, artemisinic acid. Artemisinin can be synthesized from this precursor. If the efficiency of this process can be improved, this engineered yeast strain has the potential to alleviate the drug shortage. Through the bio-engineering of Saccharomyces cerevisiae high titres of artemisinic acid were produced using a novel cytochrome P450 monooxygenase. Optimization of this process on an industrial scale may significantly reduce the cost of artemisinin, which could then be used to combat malaria in resource-poor settings. Malaria is a global health problem that threatens 300–500 million people and kills more than one million people annually1. Disease control is hampered by the occurrence of multi-drug-resistant strains of the malaria parasite Plasmodium falciparum2,3. Synthetic antimalarial drugs and malarial vaccines are currently being developed, but their efficacy against malaria awaits rigorous clinical testing4,5. Artemisinin, a sesquiterpene lactone endoperoxide extracted from Artemisia annua L (family Asteraceae; commonly known as sweet wormwood), is highly effective against multi-drug-resistant Plasmodium spp., but is in short supply and unaffordable to most malaria sufferers6. Although total synthesis of artemisinin is difficult and costly7, the semi-synthesis of artemisinin or any derivative from microbially sourced artemisinic acid, its immediate precursor, could be a cost-effective, environmentally friendly, high-quality and reliable source of artemisinin8,9. Here we report the engineering of Saccharomyces cerevisiae to produce high titres (up to 100 mg l-1) of artemisinic acid using an engineered mevalonate pathway, amorphadiene synthase, and a novel cytochrome P450 monooxygenase (CYP71AV1) from A. annua that performs a three-step oxidation of amorpha-4,11-diene to artemisinic acid. The synthesized artemisinic acid is transported out and retained on the outside of the engineered yeast, meaning that a simple and inexpensive purification process can be used to obtain the desired product. Although the engineered yeast is already capable of producing artemisinic acid at a significantly higher specific productivity than A. annua, yield optimization and industrial scale-up will be required to raise artemisinic acid production to a level high enough to reduce artemisinin combination therapies to significantly below their current prices.