Fig. 7: Conformational changes induced by NRAS mutants alter BRAF binding affinity.

a Representative conformations of NRAS^61R, NRAS^61K, and NRAS^61P extracted from their highly populated replica-exchange molecular dynamics (REMD) structural ensembles. Interactions with the codon 61 sidechain are listed below each structure. b Binding orientation of NRAS^61R and NRAS^61P with the BRAF-RBDCRD as generated using Hex molecular docking simulations. The average conformation representing highly populated structural ensembles extracted from each NRAS codon 61 mutant trajectory was docked against the BRAF-RBDCRD. In the cartoon representation, the NRAS codon 61 mutant and bound nucleotide are shown in licorice, the BRAF-RBDCRD in gray and polar interactions for each mutant, and its surrounding residues are indicated by blue dashed lines. Comparisons of the interaction energy and the number of contacts between the BRAF-RBDCRD and each NRAS mutant suggest that highly melanomagenic NRAS mutants (NRAS^61R, NRAS^61K) bind BRAF with higher affinity than NRAS^61H, NRAS^61L, and NRAS^61P. The number of autoinhibitory contacts relieved by NRAS mutant binding is listed in parentheses. BRET protein–protein interaction data from Venus-tagged NRAS mutant and Rluc8-tagged BRAF (c) or CRAF (d) constructs co-transfected into 293T cells at increasing receptor to donor ratios. The data shown are representative of two replicates. Best fit BRET₅₀ values (binding affinity) and standard error, determined by non-linear regression, are shown for each mutant. Bolded values indicate statistically significant values as compared to both NRAS^61H and NRAS^61P. p-values determined by t-tests with 20 degrees of freedom representing the number of measures per curve. Source data are provided as a Source Data file.