Computational prediction of the conformational diversity impact on Rac1 and it?s novel inhibitor 1A-116 interaction

CHINESTRAD, PATRICIO; NAZARENO GONZÁLEZ; LORENZANO MENNA, PABLO

Congreso; The A2B2C 10th Argentinian Congress of Bioinformatics and Computational Biology (10CAB2C).; 2019

BACKGROUND: The Ras-related C3 botulinum toxin substrate 1 (Rac1) protein is among the most studied proteins in the Rho GTPase family and a key regulators in the actin cytoskeleton reorganization, as it has effects on endocitosis, vesicular trafficking, cell cycle progression and cellular migration and adhesion. It is been shown that Rac1 is overactivated in a wide range of tumor types. In this context, the regulatory activity of Rac1 regarding the cytoskeleton restructuration impacts on several key process for the course of the disease, such as the invasion, migration and metastasis of tumor cells. In this work, we show proof of 1A-116?s inhibition mechanism. RESULTS: We evaluated the stability of Rac1?s interaction with its inhibitor 1A-116, taking into account said proteins conformational diversity. For further analysis, we evaluated the impact of mutations in the Trp 56 in 1A-116 binding to Rac1. For this assay, we evaluated the binding energy of 1A-116 to Rac1 W56F mutant, generated with the I-TASSER software. We observed a decrease in the binding energy of 1A-116 to the Rac1 W56F mutant in comparison to the wild type (wt) (-5.59 ± 0.0139 kcal/mol and -6.09 ± 0.00994 kcal/mol, respectively). Through point mutations in CDC42 (Rac1 holomogous protein) we observed an increase in the binding energy of 1A-116 to the CDC42 F56W mutant in comparison to the wt (-5.69 ± 0.0170 kcal/mol and -6.09 ± 0.00994 kcal/mol respectively), establishing the role of the Trp 56 in the interaction with the 1A-116 inhibitor. In order to evaluate these results, in vitro assays with the SRE-Luc reporter were performed. Similar tendencies were observed, where 1A-116 inhibitory activity was lost for the Rac1 W56F mutant but was acquired for the CDC42 F56W mutant. CONCLUSIONS: Our results not only deepen the knowledge of 1A-116 inhibition of Rac1, emphasizing the key role of Trp 56 in the interaction of 1A-116 with the protein, but also show that redocking analysis together with in silico mutation models could be a proper way of identifying drug resistant single-point mutations for more effective drug designs.