Fig. 4.

BB-Cl-amidine directly targets STING. (A and B) Structure of BB-Cl-Yne and BB-F-Yne alkyne probes. (C) ELISA analysis of IFN-β in conditioned medium from BMDMs pretreated with vehicle control (DMSO), BB-Cl-amidine (1 μM), BB-Cl-Yne (1 μM), or BB-F-Yne (1 μM) for 1 h followed by treatment with 500 nM diABZI-4 for 24 h. (D) Log₁₀ fold change enrichment of proteins from copper-clicked lysates from cells treated with BB-Cl-Yne (1 μM) for 1 h. (E) Representative mass spectrometry spectra of STING identified from streptavidin pull downs of clicked lysates from cells treated with BB-Cl-Yne or cells cotreated with BB-Cl-amidine and BB-Cl-Yne. (F) Peptide coverage analysis of peptides identified in streptavidin bead pull downs from clicked lysate cells treated with BB-Cl-Yne or cells cotreated with BB-Cl-amidine and BB-Cl-Yne. (G) Immunoblot of STING and strep-IR-dye in streptavidin bead pull downs and input from clicked lysates of cells treated with or without BB-Cl-Yne and BB-Cl-amidine. (C) pooled data from two independent experiments performed in triplicate. (D and E) performed in triplicate. (G) representative data of two independent experiments.