Malaria is a worldwide disease caused by Plasmodium parasites. A sesquiterpene endoperoxide artemisinin isolated from Artemisia annua was discovered and has been accepted for its use in artemisinin-based combinatorial therapies, as the most effective current antimalarial treatment. However, the quantity of this compound produced from the A. annua plant is very low, and the availability of artemisinin is insufficient to treat all infected patients. In addition, the emergence of artemisinin-resistant Plasmodium has been reported recently. Several techniques have been applied to enhance artemisinin availability, and studies related to its mode of action and the mechanism of resistance of malaria-causing parasites are ongoing. In this review, we summarize the application of modern technologies to improve the production of artemisinin, including our ongoing research on artemisinin biosynthetic genes in other Artemisia species. The current understanding of the mode of action of artemisinin as well as the mechanism of resistance against this compound in Plasmodium parasites is also presented. Finally, the current situation of malaria infection and the future direction of antimalarial drug development are discussed.
The production of artemisinin, the most effective antimalarial compound, is limited to Artemisia annua. Enzymes involved in artemisinin biosynthesis include amorpha-4,11-diene synthase (ADS), amorpha-4,11-diene 12-monooxygenase (CYP71AV1) and artemisinic aldehyde Δ(11)13 reductase (DBR2). Although artemisinin and its specific intermediates are not detected in other Artemisia species, we reported previously that CYP71AV1 and DBR2 homologs were expressed in some non-artemisinin-producing Artemisia plants. These homologous enzymes showed similar functions to their counterparts in A. annua and can convert fed intermediates into the following products along the artemisinin biosynthesis in planta These findings suggested a partial artemisinin-producing ability in those species. In this study, we examined genes highly homologous to ADS, the first committed gene in the pathway, in 13 Artemisia species. We detected ADS homologs in A. absinthium, A. kurramensis and A. maritima. We analyzed the enzymatic functions of all of the ADS homologs after obtaining their cDNA. We found that the ADS homolog from A. absinthium exhibited novel activity in the cyclization of farnesyl pyrophosphate (FPP) to koidzumiol, a rare natural sesquiterpenoid. Those from A. kurramensis and A. maritima showed similar, but novel, activities in the cyclization of FPP to (+)-α-bisabolol. The unique functions of the novel sesquiterpene synthases highly homologous to ADS found in this study could provide insight into the molecular basis of the exceptional artemisinin-producing ability in A. annua.
Artemisinin is the most effective antimalarial compound isolated from Artemisia annua. Artemisinic aldehyde Δ11(13)-reductase (DBR2) catalyzes the reduction of artemisinic aldehyde into dihydroartemisinic aldehyde, switching the pathway towards artemisinin production. Although other Artemisia species cannot produce artemisinin, we found a putative DBR2 ortholog expressed in A. absinthium (abDBR2). We examined the catalytic activity of abDBR2 in vitro and found that it shows comparable activity to that of DBR2 based on the reduction of artemisinic aldehyde into dihydroartemisinic aldehyde. Furthermore, we found that dihydroartemisinic aldehyde was detected in the extract of A. absinthium leaves fed with artemisinic aldehyde, suggesting the presence of active abDBR2 in planta. Our results indicate that A. absinthium may be a potential host for the production of artemisinin through metabolic engineering.
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