Fig. 5: Bufalin facilitates ERα degradation by enhancing interaction between ERα and the E3 ligase STUB1.

a The binding poses of ERα and STUB1 with or without Bufalin. The magenta structure represents the result of protein-protein docking, the green structure corresponds to the clustered conformation after a 200 ns MD simulation with Bufalin, and the blue structure represents the clustered conformation after a 200 ns MD simulation without Bufalin. Bufalin is shown as yellow sticks in the structural representation. b The conformational changes of the ERα-Bufalin-STUB1 complex at 200 ns. c RMSD plots of the ERα-STUB1 complex over 200 ns of MD simulation with or without Bufalin. d The binding surface area analysis between ERα and STUB1 with or without Bufalin. e RMSF (root-mean-square fluctuation) profiles of ERα and STUB1 with or without Bufalin. f Changes in the number of H-bonds throughout the simulation between ERα and STUB1 with or without Bufalin. g MCF-7 and T47D cell was transfected with nontargeting siRNA or STUB1-targeted siRNA, the expression of ERα and STUB1 were measured by western blot, the samples derive from the same experiment and that gels/blots were processed in parallel. h 293 T cells transfected with Flag-ERα and HA-STUB1 plasmid were treated by Bufalin, then lysed and lysates were subjected immunoprecipitation with anti-Flag antibodies. Proteins retained on sepharose were blotted with the indicated antibodies. The input samples derive from the same experiment, and that gels/blots were processed in parallel. i MCF-7 and T47D cells were transfected with nontargeting siRNA or STUB1-targeted siRNA followed by treatment with Bufalin for 48 h. The ERα and STUB1 protein levels were measured by Western blot.