Blog
About

88
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: not found

      Production of the antimalarial drug precursor artemisinic acid in engineered yeast.

      Nature

      metabolism, chemistry, Sesquiterpenes, genetics, Saccharomyces cerevisiae, Plasmodium falciparum, Molecular Sequence Data, Mevalonic Acid, economics, drug therapy, Malaria, Falciparum, Genetic Engineering, Gas Chromatography-Mass Spectrometry, Fermentation, trends, Drug Costs, Cytochrome P-450 Enzyme System, Bioreactors, Artemisinins, enzymology, Artemisia annua, Antimalarials, Animals

      Read this article at

      ScienceOpenPublisherPubMed
      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Malaria is a global health problem that threatens 300-500 million people and kills more than one million people annually. Disease control is hampered by the occurrence of multi-drug-resistant strains of the malaria parasite Plasmodium falciparum. Synthetic antimalarial drugs and malarial vaccines are currently being developed, but their efficacy against malaria awaits rigorous clinical testing. 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 sufferers. Although total synthesis of artemisinin is difficult and costly, 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 artemisinin. 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.

          Related collections

          Most cited references 21

          • Record: found
          • Abstract: not found
          • Article: not found

          Identification of an antimalarial synthetic trioxolane drug development candidate.

          The discovery of artemisinin more than 30 years ago provided a completely new antimalarial structural prototype; that is, a molecule with a pharmacophoric peroxide bond in a unique 1,2,4-trioxane heterocycle. Available evidence suggests that artemisinin and related peroxidic antimalarial drugs exert their parasiticidal activity subsequent to reductive activation by haem, released as a result of haemoglobin digestion by the malaria-causing parasite. This irreversible redox reaction produces carbon-centred free radicals, leading to alkylation of haem and proteins (enzymes), one of which--the sarcoplasmic-endoplasmic reticulum ATPase PfATP6 (ref. 7)--may be critical to parasite survival. Notably, there is no evidence of drug resistance to any member of the artemisinin family of drugs. The chemotherapy of malaria has benefited greatly from the semi-synthetic artemisinins artemether and artesunate as they rapidly reduce parasite burden, have good therapeutic indices and provide for successful treatment outcomes. However, as a drug class, the artemisinins suffer from chemical (semi-synthetic availability, purity and cost), biopharmaceutical (poor bioavailability and limiting pharmacokinetics) and treatment (non-compliance with long treatment regimens and recrudescence) issues that limit their therapeutic potential. Here we describe how a synthetic peroxide antimalarial drug development candidate was identified in a collaborative drug discovery project.
            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            Measurement of trends in childhood malaria mortality in Africa: an assessment of progress toward targets based on verbal autopsy.

            Reduction of deaths associated with malaria in children is a primary goal of malaria control programmes in Africa, but there has been little discussion about how changes in mortality will be measured. This paper assesses recent historical changes in the contribution of malaria to child survival in Africa by examining data from demographic surveillance systems (DSS) in 25 mainly rural settings. The data were adjusted for the varying sensitivity and specificity of verbal autopsies (VA) in different ranges of malaria mortality and for varying parasite prevalences. Average malaria mortality in the DSS sites in west Africa was 7.8 per 1000 child-years between 1982 and 1998; the rate did not change significantly over this period. In the sites in east and southern Africa combined, malaria mortality was 6.5 per 1000 child-years between 1982 and 1989, but it increased to 11.9 per 1000 child-years between 1990 and 1998. All-cause child mortality and non-malaria mortality, by contrast, decreased significantly over time in both regions; consequently, the proportion of deaths due to malaria rose from 18% to 23% in west African sites and from 18% to 37% in east and southern African sites between 1982-89 and 1990-98. If malaria mortality fell at a rate consistent with the Roll Back Malaria target of halving malaria mortality by the year 2010, an individual DSS of a total population of 63 500 could with adequate VA adjustment detect this reduction after 7 years.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: not found

              Molecular cloning, expression, and characterization of amorpha-4,11-diene synthase, a key enzyme of artemisinin biosynthesis in Artemisia annua L.

              In plants, sesquiterpenes of different structural types are biosynthesized from the isoprenoid intermediate farnesyl diphosphate. The initial reaction of the biosynthesis is catalyzed by sesquiterpene cyclases (synthases). In Artemisia annua L. (annual wormwood), a number of such sesquiterpene cyclases are active. We have isolated a cDNA clone encoding one of these, amorpha-4,11-diene synthase, a putative key enzyme of artemisinin biosynthesis. This clone contains a 1641-bp open reading frame coding for 546 amino acids (63.9 kDa), a 12-bp 5'-untranslated end, and a 427-bp 3'-untranslated sequence. The deduced amino acid sequence is 32 to 51% identical with the sequence of other known sesquiterpene cyclases from angiosperms. When expressed in Escherichia coli, the recombinant enzyme catalyzed the formation of both olefinic (97.5%) and oxygenated (2.5%) sesquiterpenes from farnesyl diphosphate. GC-MS analysis identified the olefins as (E)-beta-farnesene (0.8%), amorpha-4,11diene (91.2%), amorpha-4,7(11)-diene (3.7%), gamma-humulene (1.0%), beta-sesquiphellandrene (0.5%), and an unknown olefin (0.2%) and the oxygenated sesquiterpenes as amorpha-4-en-11-ol (0.2%) (tentatively), amorpha-4-en-7-ol (2.1%), and alpha-bisabolol (0.3%) (tentatively). Using geranyl diphosphate as substrate, amorpha-4,11-diene synthase did not produce any monoterpenes. The recombinant enzyme has a broad pH optimum between 7.5 and 9.0 and the Km values for farnesyl diphosphate, Mg2+, and Mn2+ are 0.9, 70, and 13 microM, respectively, at pH 7.5. A putative reaction mechanism for amorpha-4,11-diene synthase is suggested.
                Bookmark

                Author and article information

                Journal
                10.1038/nature04640
                16612385

                Comments

                Comment on this article