Abstract
Mutant RAS genes play an important role in regulating tumors through lysine residue 104 to impair GEF-induced nucleotide exchange, but the regulatory role of KRAS K104 modification on the KRASG12D mutant remains unclear. Therefore, we simulated the acetylation site on the KRASG12D three-dimensional protein structure, including KRASG12D, KRASG12D/K104A and KRASG12D/K104Q, and determined their trajectories and binding free energy with GEF. KRASG12D/K104Q induced structural changes in the α2- and α3-helices, promoted KRAS instability and hampered GEF binding (ΔΔG = 6.14 kJ/mol). We found decreased binding to the Raf1 RBD by KRASG12D/K104Q and reduced cell growth, invasion and migration. Based on whole-genome cDNA microarray analysis, KRASG12D/K104Q decreased expression of NPIPA2, DUSP1 and IL6 in lung and ovarian cancer cells. This study reports computational and experimental analyses of Lys104 of KRASG12D and GEF, and the findings provide a target for exploration for future treatment.
Highlights
Mutant RAS genes play an important role in regulating tumors through lysine residue 104 to impair guanine nucleotide-exchange factors (GEFs)-induced nucleotide exchange, but the regulatory role of KRAS K104 modification on the KRASG12D mutant remains unclear
To better understand how the K104 modification alters the intrinsic biochemical properties of KRASG12D, we focused on the KRAS/GEF protein complex structure and investigated its structural dynamics properties, including conformational changes, atomic fluctuations, and binding free energy differences between KRAS and GEF with KRASG12D/K104A and K RASG12D/K104Q mutants
The results suggest that more structural changes of the α2- and α3-helices were prevalent in the G12D + K104Q KRAS/GEF complex, which is consistent with the root mean square deviation (RMSD) results (Fig. 2A)
Summary
Mutant RAS genes play an important role in regulating tumors through lysine residue 104 to impair GEF-induced nucleotide exchange, but the regulatory role of KRAS K104 modification on the KRASG12D mutant remains unclear. The switch between the two forms is controlled by guanine nucleotide-exchange factors (GEFs) that induce activation by binding to Ras family proteins, including KRAS (and Rho-family GTPases), and catalyzing the release of GDP1–3. To better understand how the K104 modification alters the intrinsic biochemical properties of KRASG12D, we focused on the KRAS/GEF protein complex structure and investigated its structural dynamics properties, including conformational changes, atomic fluctuations, and binding free energy differences between KRAS and GEF with KRASG12D/K104A and K RASG12D/K104Q mutants. This study reveals a novel pathway through which K104Q of KRASG12D may be useful for human cancer therapy
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