Biochemical characterization of caspase-like involved in a cell cycle of simple eukariotic.

Abstract

The cell cycle is basically determined by the processes of cell division, differentiation and death. In recent a year's caspase family enzymes has been described which are involved in programmed cell death in a simple eukaryotic organisms such as fungi and protozoa (metacaspases) and in metazoan and amoeboid Dictyostelium sp (paracaspases), however, the biochemical characterization of these enzymes has not been described. In present project we intend to achieve the biochemical studies to determine kinetic parameters (KM, kcat e kcat/KM) that metacaspases of Trypanossoma brucei (TbMCA2) and Saccharomyces cerevisiae (YCA1) and that paracaspase by Dictyostelium discoideum, in order to evaluate the specificity study, evaluate the effect of salt, pH and temperature on the proteolytic activity of these proteases. Recently was describe the importance of calcium on the function of that proteases, we intend realized a kinetics assays with these enzymes wild type and contained a site direct mutagenesis, changing the concentration of calcium and established the importance of this ion in the structure-activity relationship of these proteases, evaluating the calcium effect in physical-chemical parameters of metacaspases and paracaspases. We pretend implementing in our laboratory a Dictyostelium discoideum culture in order to realize a future experiments of the actions that caspase-like proteases (metacaspases and paracaspases) on the cell cycle and the cell communication on the cell model. (AU)