Harnessing the Anaphase-Promoting Complex for Targeted Protein Degradation

Targeted protein degradation (TPD) is rapidly becoming a prevalent modality of therapeutics development due to its event-driven pharmacology that overcomes some major limitations of conventional small molecule inhibitors. One of the paradigms of TPD involves co-opting components of the ubiquitin-proteasome system (UPS) to label disease-associated proteins with ubiquitin, thereby tagging them for destruction by the cellular protein degradation machinery. However, a major limitation in the field of TPD is that most UPS-based TPD tools or degraders utilize only a handful of ubiquitin ligases despite the availability of over 600 such enzymes in humans. This work aims to address this gap by designing TPD tools to harness a ubiquitin ligase that is yet to be explored in the field, namely the anaphase-promoting complex or cyclosome (APC/C). The APC/C is a vital ubiquitin ligase involved in the destruction of numerous proteins including Securin, Cyclin B1 and Aurora kinase A. Most natural APC/C substrates contain multiple copies of short linear motifs (SLiMs), termed degrons, that serve as APC/C-recognition sequences. My research approach involved systematically dissecting various combinations of degrons from natural APC/C substrates, primarily using cell-based degradation assays conducted by timelapse quantitative fluorescence imaging. The study of degron combinations provides lessons on how the sequence, spacing, orientation, and number of degrons affect the rate, timing, and extent of degradation. One of the effective degron combinations identified was then used to functionalize a number of protein scaffolds to create a panel of synthetic APC/C-directing molecules which were further functionalized with target recognition motifs to generate candidate APC/C-based degraders.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2022

Author dc:creator

• Okoye, Cynthia Ngozi

Advisors dc:contributor.advisor

• Lindon, Catherine
• Itzhaki, Laura

• Anaphase-Promoting Complex
• PROTACs
• Targeted Protein Degradation

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/e1c5ed0e-3ad3-4aae-8bdb-682c6ce27d33/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.96465

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/349196