Fig. 4: Unexpected molecular mechanism of CU-CPD107.

a Crystal structure of the TLR8/CU-CPD107 complex (PDB: 7CRF). Front (left) and side (right) views of the complex. TLR8 and its dimerization partner, TLR8*, are colored green and cyan, respectively. The ligand molecules are illustrated by space-filling representations. The C, O, and N atoms of the ligands are colored yellow, red, and blue, respectively. b and c are the close-up views and schematic representation of interactions between CU-CPD107 and the TLR8 protein, respectively. The hydrophobic pocket and hydrogen bonds are shown as dashed gray arcs and dashed red lines, respectively. d Computational molecular modeling of CU-CPD107 docked to TLR8/ssRNA40 (PDB: 4R08) suggesting that the dimeric interface is the most likely target region for CU-CPD107. e and f are the close-up views and schematic representation of interactions between the CU-CPD107 with TLR8/ssRNA40 complex. g In silico alanine scanning of the residues within the CU-CPD107-binding site. The relative binding energy change (∆∆G) of each mutant over the wild type was calculated using the Prime-MM/GBSA method. h Activities of CU-CPD107 in the mutants and wild-type hTLR8, stimulated by 5 μg/mL ORN06. The NF-κB induction fold of the mutants and wild-type TLR8 were analyzed by a SEAP assay using HEK-Blue Null1 cells. Data are mean ± s.d.; n = 3 independent experiments. A one-way analysis of variance with Bonferroni’s multiple comparisons test for multiple comparisons was used for statistical analysis. Statistical significance of the data is indicated as follows: *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; ns not significant. Source data are provided as a Source data file.