Identification of RON tyrosine kinase receptor binding peptides using phage display method and molecular modeling of their binding mode

Abstract

Introduction: The Current efforts to combat cancer have been focused on molecular targeted therapies and RON tyrosine kinase receptor has gained much attention as a promising target for cancer treatment. Aims: The aims of this study were (i) to identify RON tyrosine kinase receptor binding peptides using phage display technology, (ii) studying the binding modes of interactions between RON and identified peptides by docking and molecular dynamics simulation methods and, (iii) identification of hot spot of RON-MSP complex by in silico alanine scanning mutagenesis. Methods: A 12-mer peptide phage library was utilized to perform biopanning against RON. The ability of the selected peptide displaying phage to RON and their possible binding sites were examined by ELISA. The binding mode and affinity of the selected peptides to RON were also calculated by docking and molecular dynamics simulation. Furthermore the important residues at the interface of RON-MSPβ complex were identified by computational alanine scanning mutagenesis. Results: The results of ELISA showed that P6 peptide displaying phage has higher affinity to RON compared to other phage and its binding location is out of ligand binding site. Docking and molecular dynamics simulation results also indicated the higher affinity of P6 to RON as well as its exosite-binding feature. The alanine scanning mutagenesis results revealed that Gln193, Arg220, Glu287, Pro288, Glu289 and His424 residues from RON and Arg521, His528, Ser565, Glu658, and Arg683 form MSPβ may play important roles in interaction between RON and MSPβ. Conclusions: Our data suggest a capacity for P6 peptide (FEHSLYKEMTHL) to be utilized as RON binding agent, and hence can be used for various purposes, including design of drug delivery systems for transferring cytotoxic agents to RON positive cancer cells, interfering with RON signaling, peptidomimetics design, and diagnostic imaging. Also identification of RON and MSP hotspots, direct the process of rational drug design for developing RON and MSP antagonizing agents.