RATIONAL DESIGN OF PEPTIDES AGAINST Pseudomonas aeruginosa TOLB-PAL INTERACTION

TUMAS IGNACIO NICOLAS; MIGUEL VIRGINIA; MONTI MARIELA ROXANA

Chapadmalal

Congreso; XVIII Congreso de la Sociedad Argentina de Microbiología General; 2023

Sociedad Argentina de Microbiología General

Pseudomonas aeruginosa (PA) is an opportunistic pathogen capable of producinginfections in immunocompromised patients. This Gram-negative bacterium has a markedtendency to acquire resistance to antibiotics for clinical use. Therefore, according to theWHO, it is among the main bacteria with priority for the development of new antimicrobials.In this work, we have focused on the periplasmic protein TolB as a potential antimicrobialtarget. It participates in the division and transport of other proteins across the outermembrane of PA. TolB essentiality has been demonstrated and there are several crystallinestructures of its homologue in Escherichia coli alone or interacting with one of its interactionpairs, such as PAL protein and colicin fragments. We propose as a strategy for thedevelopment of TolB inhibitors, the in silico design of peptides whose mechanism of actionconsists of interrupting TolB interaction with partners. With this aim, we generated a TolB3D model of PA TolB using the MODELLER software, employing a multi-template homologymodeling approach. Subsequently, the model was meticulously compared with the modelproduced by AlphaFold, a state-of-the-art protein structure prediction tool. This comparativeanalysis was conducted to ensure there are no significant discrepancies between the twomodels, thus reinforcing the accuracy and reliability of our constructed TolB model. TheTolB model was used to build four different TOLB-partners complexes using PAL protein,and the three colicin fragments as partners, resulting in 4 different protein-protein andprotein-peptide structures. These complexes were used as starting points for 30 nsmolecular dynamics simulations (MD) in order to evaluate complexes stability. Also, wedetermined the total interaction energy and energetic contribution of the residues of theligands in the interaction with TolB using GROMACS and gmx_MMPBSA. Complexesshowed low RMSD values and high affinity energies. We selected the PAL and colicinresidues that have the highest interaction energy with TolB. Using this information, we wereable to design several peptides of 10 to 16 amino acid length that mimicked the interaction of PAL and colicins combining the residues that showed the stronger affinity with TolB ofPA. Finally, we evaluated their performance using MD simulations and interaction energyanalysis.