By continuing to use this site, you agree to the use of cookies in accordance with our privacy policy.

Science and Mathematics

Chemistry Colloquium: Dr. Daniel Kraut

September 19, 2023 at 4:00pm5:00pm EDT

Center for Science and Technology (CST), 1-132

This event has already occurred. The information may no longer be valid.

The Department of Chemistry in the College of Arts and Sciences is pleased to welcome Dr. Daniel Kraut, Professor and Chair of Chemistry at Villanova University.

Title: How the Proteasome Degrades GFP: Roles of Targeting, Grip & Substrate Stability


The ubiquitin–proteasome system is the canonical pathway for protein degradation in eukaryotic cells. The proteasome uses a hexameric ATPase ring to unfold and translocate target proteins into the 20S protease core particle, where they are hydrolyzed into small peptides, Green fluorescent protein (GFP) is frequently used as a reporter in proteasomal degradation assays, yet we still do not fully understand how the proteasome unfolds and degrades this highly stable protein. Herein we investigate how the fate of GFP variants of differing intrinsic stabilities is determined by the mode of targeting to the proteasome and by the proteasomal motor protein pore loops that grip the substrate. We compared two targeting systems: linear Ub4 degrons and the UBL domain from yeast Rad23, both of which are commonly used in degradation experiments. Surprisingly, the UBL degron allows for degradation of the most stable sGFP-containing substrates, whereas the Ub4 degron does not. Destabilizing the GFP by circular permutation allows degradation with either targeting signal, indicating that domain stability and mode of targeting combine to determine substrate fate. By making mutations to one or more of the ATPase pore loops that contact the substrate, we further show that multiple functional pore loops are required to unfold the more stable GFP variants but not variants with lesser stability. Thus, the fate of GFP at the proteasome depends on a combination of targeting, substrate stability, and grip strength.

This event was first published on September 8, 2023 and last updated on September 11, 2023.

Event Details