Trypanosoma cruzi PLA2: In silico study of enzyme-substrate interaction
The struggle against Chagas disease prompted the study of the parasite Trypanosoma cruzi's metabolism and, in particular, enzyme-substrate interactions to find pharmacological targets necessary to eliminate the parasite without compromising the host. An enzyme that could meet thes...
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| Formato: | Artículo revista |
| Lenguaje: | Español |
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Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología
2019
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| Acceso en línea: | https://revistas.unc.edu.ar/index.php/med/article/view/25783 |
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| Sumario: | The struggle against Chagas disease prompted the study of the parasite Trypanosoma cruzi's metabolism and, in particular, enzyme-substrate interactions to find pharmacological targets necessary to eliminate the parasite without compromising the host. An enzyme that could meet these requirements is T. cruzi phospholipase A2 (PLA2) whose existence as an individual protein has been suggested experimentally but has not yet been obtained and purified.
In vitro: PLA2 activity was measured by hydrolysis of the bis-pyrenoyl phosphatidylcholine fluorescent substrate (10 mol%) incorporated in small unilamellar liposomes, SUVs, of dipalmitoylphosphatidylcholine (DPPC) in 10 mM HEPES buffer pH 7.2. 50 ml of filtered plasma from mice with different parasitemia were tested. The enzyme was activated with 20 mM calcium. Emission spectra were obtained before and after each kinetic determination. In silico: the Tc00.1047053510743.50 DNA sequence was translated into amino acids and modeled by homology on the phyre2 server. The highest scoring model was selected and resulted in a PLA2-like molecule with similarities to the human platelet activating factor (PAF) acetylhydrolases (PAF-hydrolase). The interaction of the structural model with possible substrates and with GM1 monosialoganglioside (inhibitor of PLA2 from other sources) was analyzed. The gene was identified by the presence of the amino acids HIS, SER, ASP (GLU) conforming the active site, the triad responsible for the nucleophilic attack on the carbon atom of the carbon ester bond, an activity characteristic of PAF-hydrolases. Ligand binding was originally studied by Linear Interaction Energies. The enzyme-substrate complexes (PAF or DPPC as ligands) were obtained with the PatchDock program. The simulations of bound and unbound structures in implicit solvent (GBIS) were developed with NAMD. The results were visualized with VMD and Chimera.
The electrostatic contribution increases approximately 20kcal / mol in the bound state, but VdW interactions decrease 28kcal / mol, which makes the term favorable for the bound state. The binding of GM1, a hypothetical inhibitor, was more favorable than for PAF, which would allow us to propose GM1 as a competitive inhibitor.
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