Obtaining and biochemical characterization of recombinant protozoan metacaspases of clinical interest

• Grant number: 22/06394-0
• Support Opportunities: Regular Research Grants
• Start date: February 01, 2023
• End date: January 31, 2025
• Field of knowledge: Biological Sciences - Biochemistry - Enzymology
• Principal Investigator: Mauricio Ferreira Marcondes Machado
• Grantee: Mauricio Ferreira Marcondes Machado
• Host Institution: Pró-Reitoria Acadêmica. Universidade de Mogi das Cruzes (UMC). Campus da Sede Mogi das Cruzes. Mogi das Cruzes , SP, Brazil
• Associated researchers: Marcelo Ferreira Marcondes Machado ; Vitor Marcelo Silveira Bueno Brandão de Oliveira ; Wagner Alves de Souza Júdice
• Associated scholarship(s): 24/05526-5 - Obtaining metacaspase (ScMCA-IA) from Saccharomyces cerevisiae containing site-directed alterations of the calcium interaction site, BP.IC

Abstract

The present research project is a continuation of the previous projects carried out by our research group, in which we produced yeast metacaspases in an expression system in E. coli, in their wild-type forms and containing site-directed mutations. Interestingly, yeast metacaspases are classified as belonging to group I, which has the characteristic of presenting its N-terminal region constituted by apolar residues, contained a lot of residues of asparagines, glutamines and prolines. The biochemical characterization of these enzymes is compromised due to their insolubility, making it necessary to produce them without their N-terminal portion. Interestingly, protozoan metacaspases belongs to group II of these enzymes, which have the characteristic of not presenting the apolar portion at its N-terminal, being soluble and active in its monomeric form. In previous works by our group, we have already cloned, produced enzymes with site-directed mutagenesis and biochemically characterized the T.brucei metacaspase-2 (TbMCA-IIb). In this research project, we intend to obtain recombinant metacaspases from Trypanosoma cruzei, Leishmania amazonenses and Plasmodium falciparum, and with these recombinant enzymes we intend to evaluate their physicochemical characteristics, evaluating the interaction of calcium ions that are essential for the activation and activity of metacaspases, we carry out the complete biochemical characterization of these enzymes as an assay with synthetic substrates, study of pH and temperature. Such biochemical characterization will be important so that in future experiments with collaborators we can test and evaluate the action of molecules with the capacity to modulate the activity of mecaspases, being of great interest for the development of new drugs against these neglected diseases. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

ARAUJO, LAURA HELENA; CHAGAS, THAYNAN APARECIDA BUENO; REIS, TAIZ; BORBA, JOAO RICARDO BUENO DE MORAIS; TRUJILHO, MARIANA NASCIMENTO ROMERO; DALZOTO, LAURA DE AZEVEDO MAFFEIS; MARCONDES, MARCELO FERREIRA; JULIANO, MARIA APARECIDA; JUDICE, WAGNER ALVES DE SOUZA; VELOSO, MARCIA PARANHO; et al. Oximic compounds as potential inhibitors of metacaspase-2 (TbMCA2) of Trypanosoma brucei. Biochemical and Biophysical Research Communications, v. 735, p. 12-pg., 2024-09-11. (22/06394-0, 19/27813-8, 21/01503-2)

DALZOTO, LAURA DE AZEVEDO MAFFEIS; TRUJILHO, MARIANA NASCIMENTO ROMERO; SANTOS, TAIZ DOS REIS; COSTA, JOAO PEDRO MARTINS SILVA; DUARTE, ANE CAROLINE MOREIRA; JUDICE, WAGNER ALVES DE SOUZA; MARCONDES, MARCELO FERREIRA; MACHADO, MAURICIO FERREIRA MARCONDES. Metacaspase of Saccharomyces cerevisiae (ScMCA-Ia) presents different catalytic cysteine in a processed and non-processed form. Biochemical and Biophysical Research Communications, v. 687, p. 5-pg., 2023-11-10. (22/06394-0, 19/27813-8, 21/01503-2)