Identification of a stretch of four discontinuous amino acids involved in regulating kinase activity of IGF1R

IGF1R is pursued as a therapeutic target because of its abnormal expression in various cancers. Recently, we reported the presence of a putative allosteric inhibitor binding pocket in IGF1R that could be exploited for developing novel anti-cancer agents. In this study, we examined the role of nine highly conserved residues surrounding this binding pocket with the aim to screen compound libraries in order to develop small molecule allosteric inhibitors of IGF1R. We generated GFP fusion constructs of these mutants to analyze their impact on subcellular localization, kinase activity as well as downstream signalling of IGF1R. K1055H and E1056G were seen to completely abrogate the kinase activity of IGF1R whereas R1064K and L1065A were seen to significantly reduce the IGF1R activity as well. During molecular dynamics analysis, various structural and conformational changes were observed in different conserved regions of mutant proteins particularly in the activation loop resulting in compromising kinase activity of IGF1R. These results show that a stretch of four discontinuous residues within this newly identified binding pocket is critical for activity as well as the structural integrity of IGF1R.