Abstract
Ras proto-oncogene encodes for small GTPases, downstream of epidermal growth factor receptor (EGFR) in the RAS/RAF/MAPK pathway. Wild type KRAS is associated with EGFR-signalling activation. In normal physiological conditions, it is activated by upstream signals when the GTP is exchanged with GDP. Playing an important role in regulation of differentiation and cell growth, RasGTPases behave as genetic switches. This transient process of GAP-mediated GTP hydrolysis becomes altered when the Kras gene is mutated. The most common Kras mutations are found in codon 12 and 13 and 61. Some other noncanonical mutations have been reported in codon 11, 14, 15, 17, 18, 19, 20, 22, 27, 30, 31, 117, 146 and 154. We aimed to demonstrate the conformational changes induced in two novel K RAS variants, p.E31K and p.G138V, identified in two CRC patients, which may account for transformative capacity by biochemical and signalling readouts in these patients. Dynamical implications and functional impact of variants were determined by in silico analysis and molecular docking of variants with GTP. MutationTaster was used for functional analysis of genetic variants and three-dimensional structure of mutant proteins were built by Swiss-Model and were further subjected to structural alignment and stability studies by I-Mutant Suite and DUET server. Both variants were predicted as 'disease causing' and protein stability analysis revealed p.G138V to be more destabilizing variant than p.E31K. When three-dimensional structures of variants were subjected to molecular docking with GTP, the mutated KRAS showed low binding affinity to the GTP as compared to the wild-type KRAS protein.