Control of protein degradation is required for healthy cellular function. Loss of control can lead to cancer and neurodegenerative diseases. One central control system for protein degradation is referred to as UPS (ubiquitin-proteasome system). The activity of the UPS is highly variable across individual cells in a population. It is important therefore to be able to measure activity on a single cell basis. This project will develop fluorescent sensors to measure the UPS activity of individual cells. These sensors could enhance efforts to develop new drugs and optimal treatment strategies on a patient by patient basis. Hands-on learning modules will also be developed and presented to middle school students in the Baton Rouge area. Undergraduates and high school students will also be recruited to participate in extended research experiences in an effort to develop and populate a robust biomanufacturing workforce.
The ubiquitin-proteasome system (UPS) governs DNA repair and protein degradation. Its specificity is governed by E3 ubiquitin ligases, which recognize select degradation sequences, or degrons, on target proteins. Unfortunately, our understanding of E3 ligase and proteasome activity is incomplete. This translates into an inability to obtain a complete picture of proteasome activity across a population of cells. The UPS could be harnessed to inhibit proteins by directing them to the proteasome for degradation using proteolysis targeting chimeras (PROTACs). While effective, current PROTACs suffer from reduced potency due to low cell permeability and high protease susceptibility. Thus, there is a need for long-lived, cell permeable peptides to be incorporated into the next generation of PROTACs. Our overarching hypothesis is that degron-based substrates can be utilized as (1) a next generation method to directly quantify proteasome activity and (2) a novel PROTAC with increased lifetime and sensitivity. The proposed work will evaluate peptides containing a β-hairpin motif as a new approach for quantifying proteasome activity in intact cells and degrading difficult to inhibit targets. We will also investigate the use of highly stable, non-natural amino acid sequences as a new class of degron-based substrates.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.