DNAbling Protein Partners

The proteasome, the multiprotein complex that is responsible for a large portion of protein degradation in the cell, has captured the interest of scientists from diverse areas, including evolutionary biology, the biochemistry of proteolysis, and drug discovery. Indeed, the recent approval of the proteasome inhibitor bortezomib for the treatment of multiple myeloma underscores the value of understanding the structure and function of this extraordinary protease. Activity-based inhibitors that become specifically and covalently attached to the proteasome have contributed greatly to its characterization, but to date no compounds have been developed that are simultaneously specific, irreversible, sensitive, and cell-permeable and that can enable visualization in live cells. Now, Verdoes et al. (Chem. Biol. 2006, 13, 1217-1226) present the synthesis and biological characterization of MV151, a fluorescent, cell-permeable vinyl sulfone-based inhibitor that can label proteasomes both in vitro and in vivo. MV151, synthesized with standard Fmoc-based solid-phase peptide chemistry, exhibited potent and specific activity against the trypsin-like, chymotrypsin-like, and peptidylglutamyl peptide hydrolytic proteasomal activities. The utility of MV151 was first demonstrated in proteasome profiling experiments in which cells were treated with known proteasome inhibitors, lysed, and then incubated with MV151. With a fluorescence scanner, direct ingel visualization of proteaseome activity not blocked by the known inhibitors enabled elucidation of their specificity. Next, fluorescence microscopy analysis of live cells exposed to MV151 demonstrated that the inhibitor was indeed cell-permeable and colocalized with the proteasome. Finally, tissue analysis of mice treated with MV151 revealed information about the bioavailability of the inhibitor. The authors observed that MV151 accumulates in the liver and pancreas and that the proteasome was labeled as expected in these tissues. The ability of this new molecular tool to facilitate characterization of proteasome inhibitors for both biochemical and medical applications will contribute significantly to progress in proteasome research. Eva J. Gordon, Ph.D.