Preliminary studies of biosynthesis of heme O and heme A in intraerythrocytic stages of P. falciparum

RAQUEL SIMAO-GURGE; JULIA A CRICCO; GERHARD WUNDERLICH; EMILIA KIMURA; TOMOKO TOYAMA; ALEJANDRO M KATZIN

Ciudad de Mexico

Congreso; ICOPA XIII- International Congress of Parasitology; 2014

Background: Intermediates of the isoprenoid metabolism can bind to heme groups, or take part in the biosynthesis of gibberellin in plants. Gibberillin-precursor inhibitors, Inabenfide and Uniconazole-P, reduce P. falciparum growth in vitro. Since a number of plant-exclusive isoprenic metabolites have also been found in P. falciparum, we set out to characterize the biosynthesis of gibberellins. Like others antifungals they can interfere in the Heme biosynthetic pathway. For this, it is important to characterize this pathway in P. falciparum entirely, starting with farnesylation of the heme group by ?heme O synthase? (HOS or Cox10) and ?heme A synthase? (HAS or Cox15). Method: We monitored gibberellin synthesis by metabolic labeling of parasites with [3H]GGPP, organic extraction and analyses by RP-HPLC. Growth inhibition assays were conducted using Inabenfide or Uniconazole-P and recovered with giberellins. The plasmodial orthologs of HOS and HAS genes were identified by their homology HOS and HAS enzymes form other organisms. We then genetically tagged both genes with GFP and HA coding sequences and visualized transfectants by fluorescence microscopy. Results: We observed no production of gibberellin in blood stage parasites and supplementation with giberellins did not recover growth in drug treated parasites. Visualization of tagged of PfCox10 and PfCox15 revealed that they were most probably localized in the mitochondria although PfCox10 presented an unusual extension at its N-terminus. Conclusion: We conclude that the parasites do not produce significant amounts of gibberellins. Given that the two drugs affect parasite growth, it would be interesting to see if these like others antifungals will interfere in the heme pathway, since azoles can remove heme from histidine rich peptide-heme complex. PfCox10 PfCox15 are fairly similar to HOS and HAS from other organisms, showing several conserved residues reported essential for these type of enzymes in S. cerevisae