FUNÇÃO DO METAL BIVALENTE NA RNA TRIFOSFATASE DE TRYPANOSOMA CRUZI. / The role of the divalente metal in the RNA triphosphatase from Trypanosoma cruzi.

AUTOR(ES)
DATA DE PUBLICAÇÃO

2006

RESUMO

Cap triphosphatases act in the first step of the mRNA capping process, removing the -phosphoryl group from the 5 end of nascent RNA. Enzymes from trypanosomatids and other lower eukaryotes depend on metals for this catalysis. This contrasts with the cysteine-dependent activity of the corresponding enzymes of mammals. This difference points to these enzymes as potential targets for drug design. This dissertation describes the identification, expression, purification, enzyme kinetics, crystallization trials and the role of divalent metal in the ATPase activity of the cap triphosphatase from Trypanosoma cruzi, the agent of Chagas disease. The properties of this enzyme are compared to the homologue enzymes previously characterized from Trypanosoma brucei and Saccharomyces cerevisiae. Similarity among those sequences indicates that the triphosphatase from T. cruzi has a tunnel domain containing the divalent metal forming its active site, as the enzyme from S. cerevisiae, whose structure was solved by crystallography and X-ray diffraction. Based on data from enzyme kinetics, circular dichroism, and intrinsic fluorescence analysis, a kinetic mechanism for the ATPase activity of the T. cruzi tunnel triphosphatase is proposed. A single metal interacts with the enzyme through the formation of a productive MnATP-enzyme complex, while free ATP inhibits activity. Manganese is also required for the tunnel stability of the T. cruzi enzyme, while the T. brucei homologue remains stable in the absence of metal, as shown for other triphosphatases. These findings may be useful to devise specific inhibitors for the T. cruzi RNA triphosphatase.

ASSUNTO(S)

microbiologia 1. trypanosoma cruzi. 2. formação do cap. 3. rna trifosfatase. 4. estrutura e função de proteínas.

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