| Solution Information | help | |
| Enzyme: | Nuclear receptor subfamily 4 group A member 1 | |
| inhibitor: | BDBM42116 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Bcl-B fluorescence polarization assay materials: 1)GST-Bcl-B protein was expressed in E. coli and purified at the Prof. John Reed's laboratory (Sanford-Burnham Medical Research Institute, San Diego, CA). a)Bcl-B is a GST-fusion protein lacking their C-terminal transmembrane domains (~~20 amino acids) b)FITC-TR3-R8 is FITC-Ahx-FSRSLHSLL-GX-R8 peptide, where Ahx is an N-aminohexanoic acid linker; X is N-aminocaproic acid; and R8 is 8 residues of D-arginine 2)Assay Buffer: 25 mM HEPES-KOH,20 mM N-glycerophosphate, 0.005% Tween-20, pH 7.5, 1 mM TCEP. 3)Bcl-B working solution: 44.4 nM GST-Bcl-B in the assay buffer, freshly prepared and kept on ice prior to use. 4)FITC-TR3 working solution: 44.4 nM FITC-TR3-R8 peptide in the assay buffer Bcl-B dose-response confirmation protocol: 1)Dose-response curves contained 10 concentrations of compounds obtained using 2-fold serial dilution. Compounds were serially diluted in 100% DMSO, and then diluted with water to 10% final DMSO | |
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