Computational protocol: Structural Basis of the Interaction of a Trypanosoma cruzi Surface Molecule Implicated in Oral Infection with Host Cells and Gastric Mucin

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Protocol publication

[…] For the modeling of gp82 protein, we selected as a template the high resolution crystal structure of inhibitor-bound Trypanosoma rangeli sialidase (PDB 1N1T), which is closely related T. cruzi trans-sialidase, . The gp82 sequence (Genbank L14824), which exhibited >39% identity when aligned with T. rangeli sialidase, was modeled using YASARA software (www.yasara.org) based on T. rangeli sialidase structure obtained from the Protein Data Base (www.rcsb.org). The best model was prepared for energy minimization and all the hydrogen atoms and other missing atoms from the model were created. Parameters for the force field were obtained from YAMBER3 , the pKa values for Asp, Glu, His and Lys residues were predicted. Based on the pH 7.0, the protonation states were assigned according to convention: Asp and Glu were protonated if the predicted pKa was higher than the pH, His was protonated if the predicted pKa was higher than the pH and it did not accept a hydrogen bond, otherwise it was deprotonated, Cys was protonated, Lys was deprotonated if the predicted pKa was lower than the pH, Tyr and Arg were not modified (www.yasara.org). A simulation box was defined at 15 Å around all atoms of each macromolecular complexes, then it was filled with water molecules and Na/Cl counter ions, that were placed at the locations of the lowest/highest electrostatic potential, until the cell neutralization, and the requested NaCl concentration reached 0.9%. A short molecular dynamics (MD) simulation was performed for the solvent adjust, and water molecules were subsequently deleted until the water density reached 0.997 g/ml. A short steepest descent energy minimization was carried out until the maximum atom speed dropped below 2,200 m/s. Then 500 steps of simulated annealing were performed with a temperature 0 K. Finally, a 45 nanosecond simulation at 298 K and a non-bonded cutoff of 7.86 A was carried out. A snapshot was saved every 7.5 picosecond. The graphical analysis was carried out using Visual Molecular Dynamics (VMD) software . The average structure based on the last 20 nanoseconds was submitted to an energy minimization and used in all analyses. Validation procedure using the program PROCHECK demonstrated that the final 3D structure agreed with the distance restrains and offered good geometry and side chain packing. The residues’ exposure for the solvent was analyzed using the program DSSP . […]

Pipeline specifications

Software tools YASARA, VMD, PROCHECK
Applications Drug design, Protein structure analysis
Organisms Trypanosoma cruzi
Diseases Infection, Chagas Disease