Fig. 4

Protein structure-based analysis of drug-associated mutations. a The number of known drug-associated mutations that can be mapped onto PDB structures, the number of known drug-associated mutations that are found in HotSpot3D clusters, and the number of putative druggable mutations are shown, both in aggregate and for specific genes (x-axis). b Protein structure views of one HotSpot3D cluster in BRAF (PDB: 4MBJ). Known and putative druggable mutations are distinguished by different colors in mutation labels. A drug molecule in the binding pocket is indicated in blue. c Western blot for BRAF mutation cluster found in b. HEK293T cells were transiently transfected with wild-type (WT) or mutant BRAF constructs and were cultured in 0.5% calf serum for 24 h before treatment with Dabrafenib (0-1uM) for 6 h. BRAF activity was analyzed by quantifying phosphorylation changes in MEK1/2. To normalize for transfection and loading variations, pMEK levels were divided by BRAF levels and then by GAPDH levels to produce the normalized relative intensities of pMEK/BRAF/GAPDH. This was then normalized to the WT sample without drug treatment that was set as 1. The error bars represent biological replicates