RESUMO
Plasmodium falciparum mitochondrial ADP/ATP transporter or adenylate translocase (PfAdT) was previously characterised at the molecular level and intracellularly located by immuno-electromicroscopy. Inhibition of this transporter blocks parasite development in erythrocytes. In this study, PfAdT was expressed in C43 (DE3) Escherichia coli strain under isopropyl beta-d-thiogalacto-pyranoside (IPTG) induction to screen inhibitory molecules. PfAdT was integrated directly into the bacterial cytoplasmic membrane. Whereas IPTG-induced bacterial cells imported radioactively labelled ATP, non-induced cells did not. The transporter bound specifically ADP and ATP, but not AMP. IPTG-induced cells preloaded with labelled ATP exported ATP after exogenous addition of unlabelled ADP or ATP, indicating a counter exchange transport mechanism. Bongrekic acid and atractyloside, two well-known specific inhibitors of mitochondrial ADP/ATP transporter, were tested. This experimental model was evaluated using three Malagasy crude plants extracts which have shown antiplasmodial activity on in vitro parasite cultures.
Assuntos
Antimaláricos/farmacologia , Translocases Mitocondriais de ADP e ATP/antagonistas & inibidores , Translocases Mitocondriais de ADP e ATP/metabolismo , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/enzimologia , Animais , Atractilosídeo/farmacologia , Western Blotting , Ácido Bongcréquico/farmacologia , Membrana Celular/enzimologia , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Eritrócitos/parasitologia , Escherichia coli/enzimologia , Escherichia coli/genética , Regulação Enzimológica da Expressão Gênica , Translocases Mitocondriais de ADP e ATP/genética , Extratos Vegetais/farmacologia , Plasmodium falciparum/crescimento & desenvolvimento , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Especificidade por SubstratoRESUMO
Two new sesquiterpene lactones, wedelolides A (1) and B (2), were isolated by bioassay-guided fractionation from the leaves of Wedelia trilobata, together with known trilobolides 6-O-isobutyrate (3) and 6-O-methacrylate (4). The compounds 1 and 2 were the first examples of an unprecedented framework: a novel sesquiterpene delta-lactone, (9R)-eudesman-9,12-olide. The structures of the antimalarial wedelolides A (1) and B (2) were determined on the basis of MS and 2D NMR spectral analysis. The absolute configuration of eight carbon stereocenters of compounds 1 and 2 was determined to be 1S,4S,5S,6R,7S,8S,9R,10S by mean of auxiliary chiral MTPA derivatives.