Mutational Landscape of KRAS Substitution at 12Th Position - A Systematic and Bioinformatics Approach

Posted: 7 Feb 2020

See all articles by Udhaya Kumar S

Udhaya Kumar S

Vellore Institute of Technology (VIT)

Thirumal Kumar D

Vellore Institute of Technology

Bithia R

Vellore Institute of Technology

Srivarshini Sankar

Vellore Institute of Technology

Siva R

Vellore Institute of Technology

George Priya Doss C

Vellore Institute of Technology

Hatem Zayed

Qatar University - Department of Biomedical Sciences

Date Written: December 31, 2019

Abstract

Background and Aims: KRAS is a small GTPase that functions as a cellular signaling mediator for specific receptors. By recruiting GEFs and GAPs, KRAS alternates between inert GDP-bound and dynamic GTP-bound state. Mutation in amino acid position 12 is reported as one of the causes of this perturbation. Mutant KRAS protein is persistent in its dynamic GTP-bound state, prolonging the oncogenic cellular growth signaling. The present examination intends to comprehend the conformational changes induced by the oncogenic mutations causing the impairment of GAP intervened GTP hydrolysis.

Methods: A total of 6 variants G12A, G12C, G12D, G12R, G12S, and G12V reported in UniProt were analyzed in the current study. Distinct computational tools were used to evaluate the effect of the substitution of amino acid on the stability of native KRAS protein. iStable predicted the stability of mutated KRAS protein using sequential input. FoldX algorithm of the SNPeffect tool was used for the characterization and annotation of the disease at the molecular level based on DDG values. DynaMut enabled rapid analysis of protein's dynamics and stability resulting from vibrational entropy changes. CUPSAT was used for predicting differences in the free energy of unfolding between native and mutant proteins. As the interaction of GTP with KRAS is significant docking analysis was carried out using AutoDock4.2 by retrieving the macromolecule structures from RCSB PDB. The structure of GTP was obtained from PubChem and used as a ligand. The provision of active sites for docking was based on literature.

Results: iStable results presented a decrease in protein stability upon aminoacid substitution of all the 6 variants. FoldX calculated the difference in free energy of the mutation, where an increment in DDG value depicts the destabilization of structure. Protein stability was slightly enhanced in the case of G12R, while the other mutations had no effect on protein stability. The ΔΔG prediction of DynaMut reported all the mutations as destabilizing. CUPSAT reported G12C and G12V as stabilizing with ΔΔG values of 4.21 kcal/mol and 0.64 kcal/mol, respectively. On docking of native KRAS with GTP, the binding energy of -8.23 kcal/mol was obtained. Docking analysis of mutant KRAS with GTP yielded lesser binding energies of -7.47 kcal/mol, -6.87 kcal/mol, -6.24 kcal/mol, -6.10 kcal/mol, -5.93 kcal/mol, -5.91 kcal/mol in G12A, G12V, G12D, G12S, G12R, and G12C mutations, respectively. Furthermore, on a comparison of binding energies between the native KRAS and mutants, we can elucidate that the mutations G12A and G12V have a better binding efficiency among the other variants.

Conclusion: KRAS remains an important therapeutic target for human cancers. Malignancies with KRAS mutations are impervious to standard treatments and are often excluded from targeted therapies. It has been reported that the mutant KRAS protein exists predominantly in GTP-bound active state. Our ongoing investigation on molecular dynamics simulations of the mutant KRAS protein may benefit in selectively targeting the active GTP-bound KRAS.

Keywords: KRAS, Molecular docking, Molecular dynamics simulation, Cancer, Mutation

Suggested Citation

S, Udhaya Kumar and D, Thirumal Kumar and R, Bithia and Sankar, Srivarshini and R, Siva and C, George Priya Doss and Zayed, Hatem, Mutational Landscape of KRAS Substitution at 12Th Position - A Systematic and Bioinformatics Approach (December 31, 2019). Proceedings of International Conference on Drug Discovery (ICDD) 2020, Available at SSRN: https://ssrn.com/abstract=3532370

Udhaya Kumar S

Vellore Institute of Technology (VIT) ( email )

Gorbachev Rd
Tamil Nadu
Vellore, IN Tamil Nadu 632014
India

Thirumal Kumar D

Vellore Institute of Technology ( email )

08056295915 (Phone)

Bithia R

Vellore Institute of Technology ( email )

Gorbachev Rd
Tamil Nadu
Vellore, IN Tamil Nadu 632014
India

Srivarshini Sankar

Vellore Institute of Technology ( email )

Gorbachev Rd
Tamil Nadu
Vellore, IN Tamil Nadu 632014
India

Siva R

Vellore Institute of Technology ( email )

Gorbachev Rd
Tamil Nadu
Vellore, IN Tamil Nadu 632014
India

George Priya Doss C (Contact Author)

Vellore Institute of Technology ( email )

IN

Hatem Zayed

Qatar University - Department of Biomedical Sciences ( email )

Doha, 2713
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