RESUMO
The cDNA encoding the Schistosoma mansoni dolichol phosphate mannose synthase was completely sequenced, displaying the highest homology with Cricetulus griseus and Saccharomyces pombe genes. The Schistosome enzyme had a K(m) of 0.127 microM, a value that is within the range of those reported for several other species. Thin-layer chromatography of the radiolabelled schistosome lipid intermediate showed it was identical to dolichol-phosphate (C80-C105). Expression of dolichol phosphate mannose synthase of S. mansoni (SmDPMS) was analysed by Northern blot and quantified by semi-quantitative RT-PCR with cDNA from mature and immature male and female worms. Northern blot analysis revealed a single 1-kb band. Both approaches confirmed a higher level of expression in mature female worms, as compared to immature and male worms.
Assuntos
Fosfatos de Dolicol/metabolismo , Manosiltransferases/metabolismo , Schistosoma mansoni/enzimologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Northern Blotting , Cromatografia em Camada Fina , Primers do DNA/química , Feminino , Biblioteca Gênica , Masculino , Manosiltransferases/genética , Dados de Sequência Molecular , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de AminoácidosRESUMO
Complementary DNA, encoding the mitochondrial enzyme NADH-ubiquinone oxidoreductase subunit 5 (SmND5) of the human parasite Schistosoma mansoni was isolated by screening a S. mansoni cDNA library with a human androgen receptor (hAR) cDNA probe. The complete nucleotide and deduced aminoacid sequences of SmND5 were determined. Southern blot analysis revealed the occurrence of a single copy gene for SmND5 and by means of RT-PCR, it was shown that sex- and stage-specific expression of SmND5 occurred. In order to establish a functional relationship between the mitochondrial enzyme and the androgen receptor, the effects of testosterone were compared to those of classical respiratory chain inhibitors, using adult schistosome and beef heart submitochondrial particles. Physiological concentrations of testosterone were able to inhibit the maintenance of proton gradient across the mitochondrial membranes, as well as ATP synthesis. The steroid was found to be cytotoxic to the larvae, but not to adult schistosomes. A model is proposed to explain the observed in vivo testosterone-related differences in worm burdens, in experimental chronic infections.
