Associate Professor, Department of Molecular Biosciences, Center for Bioinformatics
University of Kansas
Characterizing and developing inhibitors of proteasome assembly (2017-18)
The proteasome is a large macromolecular machine that serves as the proteolytic component of the major protein degradation pathways in eukaryotes, archaea, and actinomycete bacteria. As a result, the proteasome plays a key role in both protein homeostasis and the regulation of protein function in a variety of organisms. It has also emerged as a major drug target in a number of diseases, particularly in the treatment of cancer and tuberculosis infections. Cells do not synthesize the proteasome as a single, active unit, but rather as a set of protein subunits that must be assembled into a specific structure in order to function. This has lead to the proposal that proteasome assembly inhibitors could represent a novel therapeutic approach to tuberculosis and other diseases, offering an avenue to overcoming both drug resistance and the toxic offtarget effects that characterize traditional active site inhibitors of proteasome function.
Despite the clear promise of targeting assembly, there are currently no examples of proteasome assembly inhibitors in the literature. To overcome this problem, we have focused on discovering inhibitors of the assembly of the proteasome Core Particle (CP). The CP is the catalytic component of the proteasome, and only becomes active after the last step in assembly, making it a prime target for an assembly inhibitor.
We have developed a set of in vitro self-assembly assays using the well-established Rhodoccocus erythropolis CP as a model system. Its two subunits (α and β) can be expressed and purified separately as monomers; these monomers spontaneously self-assemble into active CPs when mixed. Using computational screens, we recently discovered a small molecule that represents the first known inhibitor of CP assembly. The first goal of this proposal is to work with the Biomolecular NMR and Protein Production core labs to validate that this molecule is binding to the β subunit of the erythropolis CP. The second aim of this proposal focuses on developing self-assembly assays for the tuberculosis CP, so that we can test if our existing inhibitor, and hits from ongoing computational screens, can inhibit assembly in tuberculosis.
If successful, this work will provide validation of the first known example of a proteasome assembly inhibitor. It will also allow us to determine if this approach can be used to target tuberculosis. Compounds discovered and characterized through this work will serve as the starting point for the eventual development of a novel class of therapeutics targeting CP assembly in the treatment of tuberculosis and other diseases.