Comparative effects of oncogenic mutations G12C, G12V, G13D, and Q61H on local conformations and dynamics of K-Ras

Abstract

K-Ras is the most frequently mutated protein in human cancers. However, until very recently, its oncogenic mutants were viewed as undruggable. To develop inhibitors that directly target oncogenic K-Ras mutants, we need to understand both their mutant-specific and pan-mutant dynamics and conformations. Recently, we have investigated how the most frequently observed K-Ras mutation in cancer patients, G12D, changes its local dynamics and conformations (Vatansever et al., 2019). Here, we extend our analysis to study and compare the local effects of other frequently observed oncogenic mutations, G12C, G12V, G13D and Q61H. For this purpose, we have performed Molecular Dynamics (MD) simulations of each mutant when active (GTP-bound) and inactive (GDP-bound), analyzed their trajectories, and compared how each mutant changes local residue conformations, inter-protein distance distributions, local flexibility and residue pair correlated motions. Our results reveal that in the four active oncogenic mutants we have studied, the α2 helix moves closer to the C-terminal of the α3 helix. However, P-loop mutations cause α3 helix to move away from Loop7, and only G12 mutations change the local conformational state populations of the protein. Furthermore, the motions of coupled residues are mutant-specific: G12 mutations lead to new negative correlations between residue motions, while Q61H destroys them. Overall, our findings on the local conformational states and protein dynamics of oncogenic K-Ras mutants can provide insights for both mutant-selective and pan-mutant targeted inhibition efforts.

Keywords: K-Ras; K-Ras mutant; Local dynamics; Molecular dynamics.

Graphical abstract

Fig. 1

K-Ras protein and the most frequently mutated residues. (A) Secondary structure of K-Ras in ribbon representation. Functional regions are in the same color as in K-Ras sequence. Arrows point to mutated residues. (B) Schematic of K-Ras sequence (residues 1–169). Arrows: β-sheets, rectangles: α-helices.

Fig. 2

Alterations in active K-Ras conformations due to oncogenic mutations. Left panels: changes in pairwise residue distances ($\mathrm{\Delta }\stackrel{-}{R}$_ij) in active K-Ras due to mutation. Positive $\mathrm{\Delta }\stackrel{-}{R}$_ij values indicate divergent pairs (red); negative $\mathrm{\Delta }\stackrel{-}{R}ij$ values indicate convergent pairs (blue). Right panels: all $\mathrm{\Delta }\stackrel{-}{R}$_ij values averaged for each residue, ($\langle \mathrm{\Delta }\stackrel{-}{R}i\rangle )$. Positive values indicate that the mutation causes a residue to move away from its neighbors; negative values indicate that a residue moves close to its neighbors. The predominant behavior for all studied mutants is positive (A-B) K-Ras^G12C (C-D) K-Ras^G12V (E-F) K-Ras^G13D (G-H) K-Ras^Q61H. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig 3

Distribution of distances W(R_ij) between Cα-Cα atoms of residue pairs in SII-α3 region in wild-type and G12 mutant K-Ras proteins. GTP-bound active K-Ras (black: WT, red: mutant) (A) Q61-D92 in WT and G12C, (B) Q61-D92 in WT and G12V; GTP-bound active K-Ras (grey: WT, pink: mutant) (C) Q61-D92 in WT and G12C, (D) Q61-D92 in WT and G12V. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 4

Distribution of distances W(R_ij) between Cα-Cα atoms of residue pairs in the P loop-SII region. GTP-bound active K-Ras (black: WT, red: G12C mutant) (A) A11-Q61, (B) G12D-Q61, (C) G12D-Q61; GTP-bound active K-Ras (grey: WT, pink: G12C mutant) (D) A11-Q61, (E) G12D-Q61, (F) G12D-Q61. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 5

Distribution of distances W(R_ij) between Cα-Cα atoms of residue pairs in the α3-Loop7 region. GTP-bound active K-Ras (black: WT, red: G12C mutant) (A) A11-Q61, (B) G12D-Q61, (C) G12D-Q61; GTP-bound active K-Ras (grey: WT, pink: G12C mutant) (D) A11-Q61, (E) G12D-Q61, (F) G12D-Q61. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 6

Changes in K-Ras dynamics due to oncogenic mutations. Y-axis shows the RMSF values of residues. GTP-bound active K-Ras (black: WT, red: mutant) (A) G12C-GTP (B) G12V-GTP (C) G13D-GTP (D) Q61H-GTP; GTP-bound active K-Ras (grey: WT, pink: mutant) (E) G12C-GDP (F) G12V-GDP (G) G13D-GDP (H) Q61H-GDP. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 7

Correlated motions of residue pairs in both active and inactive K-Ras G12 mutants. Positive correlations are in red and negative correlations are in blue. Pairwise correlation coefficients are plot for (A) K-Ras^WT-GTP, (B) K-Ras^G12C-GTP, (C) K-Ras^G12V-GTP, (D) K-Ras^WT-GDP, (E) K-Ras^G12C-GDP, (F) K-Ras^G12V-GDP. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 8

Correlated motions of residue pairs in both active and inactive G13D and Q61H mutants. Positive correlations are in red and negative correlations are in blue. Pairwise correlation coefficients are plot for (A) K-Ras^G13D-GTP, (B) K-Ras^Q61H-GTP, (C) K-Ras^G13D-GDP, (D) K-Ras^Q61H-GDP. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)