Rbin-1

Rbin-1 is a potent, reversible, and specific chemical inhibitor of eukaryotic ribosome biogenesis. Rbin-1 inhibits the ATPase with GI50 of 136±7 nM.

Signaling Pathways >> Others >> Others

Research Areas >> Metabolic Disease

GI50: 136±7 nM (ATPase)[1]

Rbin-1 is a potent and reversible triazinoindole-based inhibitors of eukaryotic ribosome biogenesis. Rbin-1 inhibits recombinant full-length Mdn1’s ATPase activity. Two of the active analogs (Rbin-1 and Rbin-2) inhibit the ATPase activity by 40% at 1 uM. In particular, an analog (Rbin-2) with a bromine substituent at postion-7 is 10-fold more active than Rbin-1 (GI50=14±1 nM (Rbin-2); 136±7 nM (Rbin-1), n=4, mean±SD)[1].

Radioactive γ-P32-ATP is added to 600 mM MgATP (pH=7) solutions at volume ratios of 1:1000-1:300, depending on the lifetime of the radioactive reagent. The total volume of each reaction is 12 mL, including 6 ml of protein from size exclusion chromatography fractions (final concentration 0-50 nM for different fractions, peak fractions are used for Rbin-1 and AMPPNP inhibition), 4 mL FPLC SEC buffer with 0.6 mM Na2SO4 and 2 mL MgATP (final concentration=100 mM). The reactions are then incubated at room temperature for 30 or 60 min before quenching with 12 mL 0.2 M EDTA. 1 mL from each reaction mixture is spotted on to TLC PEI cellulose F plates. The TLC buffer contained 0.15 M formic acid and 0.15 M lithium chloride. The TLC plates are then imaged using the Typhoon Scanner 9400. ImageJ is used to calculate the densitometric ratio of the spots corresponding to radioactive free phosphate and ATP to determine the percent of ATP hydrolyzed[1].

[1]. Kawashima SA, et al. Potent, Reversible, and Specific Chemical Inhibitors of Eukaryotic Ribosome Biogenesis. Cell. 2016 Oct 6;167(2):512-524.e14.

Captisol | Cyclosporin A | H2DCFDA | 0MPTP hydrochloride | GW4869 | Etomoxir | TD139 | Mitoquinone mesylate | GSK2795039 | JC-1 | BAPTA-AM | AP 20187 | Setanaxib (GKT137831) | D-Luciferin | Crotaline