Figure 5.

Structural basis for the inhibition of FGFR4K^V550L gate-keeper mutant by FIIN-2. (A) The chemical structure of FIIN-2. (B) The LC-MS/MS spectra of the kinase peptide (Pro42-Arg53) from FGFR4K^WT and FGFR4K^V550L with and without FIIN-2. The reacting Cys477 from the glycine-rich loop of the kinase is highlighted in red color. (C) Ribbon diagram of the FGFR4K^V550L–FIIN-2 cocrystal structure. (D) The close-up view of the main interactions between FGFR4K^V550L and FIIN-2. The hydrogen bonds are indicated as black dashed lines, and the hydrophobic interactions are shown as surface. (E) Close-up view of the DFG motif conformation in the FGFR4K^WT (in orange), FGFR4K^V550L (in teal), and FGFR4K^V550L–FIIN-2 (in blue) structures following superimposition of the three structures. Note that FIIN-2 also binds to the ATP-binding site of FGFR4K in DFG-out mode. The phenylalanines in the DFG region of FGFR4K^WT, FGFR4K^V550L, and the FGFR4K^V550L–FIIN-2 complex are rendered as orange, teal, and blue sticks and labeled in black and blue, respectively. (F) Superimposition of the FGFR4K^WT–FIIN-2 complex structure onto the FGFR4K^V550L–FIIN-2 structure. Note that rotational freedom around the single bond linking the scaffold and dimethoxyphenyl of FIIN-2 allows for small structural adjustments to bypass any potential steric clash with the bulkier side chain of L550. The V550 in FGFR4K^WT and L550 in FGFR4K^V550L are shown in orange and yellow sticks, and labeled in orange and red, respectively. (G) The distances between L550 of the FGFR4K^V550L gate-keeper mutant and FIIN-2 are shown as dashed lines and labeled in a black color. In all of the structures, the FIIN-2 is rendered as sticks and labeled in black.