## 2 Results and Discussion
### Effect of A59E and K117R on Dynamics of HRAS  
DCCMs offer valuable insight into the alterations in the internal dynamics of targets resulting from point mutations and ligand associations, which are crucial for drug design toward these targets. In this study, DCCMs were computed using the coordinates of the C\(\alpha\) atoms in HRAS to investigate the impact of mutations on the internal dynamics of HRAS (Figures 3 and S5). Our findings demonstrate that A59E and K117R mutations lead to substantial disruptions in correlated motions between structural regions of the GTP-bound active and GDP-bound inactive forms of HRAS. For the GTP- and GDP-bound WT HRAS (Figure 3A,B), the regions R1 and R2 show the anti-correlated motions of the P-loop relative to the switch domains SW1 and SW2, respectively. The region R3 describes the positively correlated motions of the loop L1 and the helix \(\alpha\)3 relative to the P-loop. In contrast, the region R4 characterizes the anti-correlated movements of the loop L1 and the helix \(\alpha\)3 relative to the SW2 (Figures 1A and 3A,B). In addition, the region R5 shows the anti-correlated motion between the L3 loop and the SW2 (Figures 1A and 3A,B). Compared to the GTP-bound WT HRAS, A59E weakens the anti-correlated motion between the P-loop and the SW1, but K117R strengthens the anti-correlated motion of this region (Figures 3C,E and S5A,C). K117R not only leads to the disappearance of the anti-correlated motion of the P-loop relative to the SW2, but also induces the appearance of positively correlated motion between the P-loop and the SW2 by referencing the GTP-bound WT HRAS (Figures 3C,E and S5A,C). On the contrary, A59E hardly affects the anti-correlated movement in the R2 or the positively correlated motion of the R3 in the GTP-bound active HRAS (Figure 3B). Differently, K119R weakens the positively correlated motion of the region R3 relative to the GTP-bound WT HRAS (Figure 3C and S5A). Although A59E and K117R scarcely affect the anti-correlated movement of the region R4, these two mutations reduce the anti-correlated motion of the region R5 compared to the GTP-bound WT HRAS (Figures 3C,E and S5A,C). Regarding the GDP-bound HRAS, K117R hardly changes the anti-correlated movements of the P-loop relative to SW1 and SW2 or the positively correlated motion of the region R3 by comparison with the GDP-bound WT HRAS (Figures 3D,F and S5B,D). However, A59E highly weakens the anti-correlated movements of the P-loop relative to the SW1 and SW2. It slightly reduces the positively correlated motion of the region R3 by referencing the GDP-bound WT HRAS (Figures 3D,F and S5B,D). By comparison with the GDP-bound WT HRAS, A59E significantly weakens the anti-correlated movement of the region R4 (Figures 3D and S5B), and K117R slightly abates the anti-correlated motion of this region (Figures 3D,F and S5B,D). In addition, A59E and K117R almost lead to the disappearance of anti-correlated movement in the region R5 relative to the GDP-bound WT HRAS (Figures 3D,F and S5B,D). In the regions above, the R1 and R2 correspond to the switch domains SW1 and SW2, respectively, while the R3, R4, and R5 are involved in allosteric regulation sites on the activity of HRAS. Thus, the changes in the internal dynamics of the regions R1-R5 caused by A59E and K117R can significantly impact the binding of HRAS to its effectors and allosteric regulations on the activity of HRAS. With the expectation of deeply probing the mutation-mediated effect on the dynamics behavior of HRAS, PCA was carried out through diagonalization of a covariance matrix constructed using the coordinates of the C\(\alpha\) atoms. The function of eigenvalues vs. eigenvectors arising from PCA was calculated (Figure S6). The results suggest that A59E and K117R strengthen structural fluctuation along the first eigenvector relative to the GTP-bound WT HRAS (Figure S6A).