TROSY-based NMR evidence for a novel class of 20S proteasome inhibitors.

The proteasome plays a central role in maintaining cellular homeostasis, in controlling the cell cycle, in removing misfolded proteins that can be toxic, and in regulating the immune system. It is also an important target for novel anticancer drugs, such as bortezomib, a potent inhibitor that has been used successfully in the treatment of multiple myeloma. Here, we show that the antimalaria drug chloroquine inhibits proteasome function in eukaryotic cell extracts and in preparations of purified 20S archaeal proteasome from Thermoplasma acidophilium. Methyl-TROSY-based NMR spectroscopy experiments conducted with the 670 kDa 20S proteasome localize chloroquine binding to regions between the α and β subunits of the α−β−β−α barrel-like structure, ~20 A from the proteolytic active sites in this 7-fold symmetric molecule. Complementary amide TROSY experiments that provide further probes of proteasome−inhibitor interactions were performed on a novel 180 kDa single-ring construct containing only α subunits, the p...