Exploring the Profiles of ROS1 Tyrosine Kinase: A Structural Analysis of G2032R and D2033N Mutations

Abstract

Background: ROS1, a proto-oncogene, drives cancer through chromosomal fusions. The G2032R and D2033N mutations, common in ROS1-rearranged non-small cell lung cancer, hinder crizotinib treatment. We investigate these mutations' impact on ROS1 structure through molecular dynamics (MD) simulations, revealing destabilization. Our findings shed light on how these mutations contribute to cancer development.

Materials and methods: The crystal structure of human ROS1 (PDB ID: 7z5x) served as the template for homology modeling and further mutation insertion of G2032R and D2033N substitutions introduced using Swiss-PdbViewer. The MD simulations were conducted on the wild-type (WT) and mutant ROS1 kinase domains to explore the structural changes and interactions.

Results: The initial model of the human ROS1 crystal structure was constructed, incorporating missing loop residues and then utilized for the MD simulation studies. The examination of conformational changes in WT, G2032R, and D2033N mutant ROS1 proteins involved observing alterations in the C-alpha protein. We observed that the mutations resulted in deviations in the MD trajectory over the 500 ns period. Consequently, the MD simulations unveiled significant conformational changes induced by the G2032R and D2033N mutations, affecting protein stability and dynamics, particularly in regions such as the ATP binding and active sites.

Conclusion: Our study constructed an initial model of the human ROS1 and used it for MD simulation studies to examine the conformational changes in ROS1 mutants. Notably, our observations revealed that the mutations caused deviations in the MD trajectory. The G2032R and D2033N mutations significantly alter ROS1 structure, affecting its stability and dynamics, offering key insights into their role in cancer disease development.

Keywords: Molecular dynamics simulation; ROS1; mutation G2032R D2033N; non-small cell lung cancer.

Figure 1

A comparison of the missing loop of ROS1 in the crystal structure with the modeled structure is shown in (a), with the modeled loop depicted as an orange surface view. The mutation positions G2032R and D2033N in the mutant ROS1 structure are highlighted in (b)

Figure 2

Molecular dynamics simulation was conducted for both the native ROS1 in the crystal structure and the mutant structure with mutations at positions G2032R and D2033N. Analysis included a root-mean-square deviation plot (a) and a root-mean-square fluctuations plot (b). Different curves in the figures are denoted by distinct colors. WT: Wild-type

Figure 3

Molecular dynamics simulation results of the native ROS1 mutant structure with mutations at positions G2032R and D2033N. The analysis encompassed solvent-accessible surface area plot (a) and a radius of gyration plot (b). Various curves in the figures are distinguished by colors. WT: Wild-type

Figure 4

Free energy landscape analysis was conducted for both the wild-type ROS1 (a) and the mutant structures G2032R (b) and D2033N (c). WT: Wild-type

Figure 5

Two-dimensional projection plot by molecular dynamics simulation results were analyzed for ROS1 Wild-type and mutations at positions G2032R and D2033N in the mutant structure. The analysis included eigenvector 1 versus eigenvector 2 (a), root mean square (RMS) fluctuations 1 plot versus atom number for the entire system (b), and the RMS fluctuations 2 plot (c). All curves in the figures share color schemes. RMS: root mean square. WT: Wild-type

Figure 6

The graphical representation depicts alterations in the propensity of secondary structure elements, illustrating the time evolution of these elements in the protein at 300 K, classified using DSSP, for both the wild-type ROS1 (a) and the mutant structures G2032R (b) and D2033N (c). WT: Wild-type, DSSP: Define secondary structure of proteins

Figure 7

Density distribution analysis was graphed to comprehend atomic orientation, utilizing the densmap script for the WT and mutant structures throughout the molecular dynamics (MD) simulations. The density maps of the MD simulation trajectory analysis for ROS1 wild-type and the mutations G2032R and D2033N in the mutant structure are illustrated in panels (a-c), respectively