Synthesis of a Chemical Toolbox Investigating Mitochondrial Protein Degradation

Aslam, Sariya (2024) Synthesis of a Chemical Toolbox Investigating Mitochondrial Protein Degradation. Masters by Research thesis (MSc), Manchester Metropolitan University.

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Abstract

Targeted protein degradation is a key area of interest for protein regulation, with the emergence of Proteolysis Targeting Chimeras (PROTACs) being a significant tool in selective protein degradation. These small, heterobifunctional compounds allow for the drugging of proteins which were previously considered undruggable, with a moiety that binds to the protein of interest (POI) and one which binds to an E3 ligase, with both components being bound together through a linker. Previous research has seen the development of PROTACs within the eukaryotic genome through the ubiquitin proteasome system (UPS), with advancements also seeing proof of concept PROTACs applied in prokaryotic cells. Here, Caseinolytic Protease Proteolytic Unit (ClpP) has been investigated as a potential novel protease for mitochondrial based protein degradation. Previous uses of ClpP type PROTACs in prokaryotic systems have been exhibited, indicating the potential of similar tools to work within the mitochondria. Therefore, this project sees the synthesis of a novel mitochondrial based degradation toolbox utilising the HaloTag degradation strategy to further understand mitochondrial based protein degradation, potentially leading to treatment of associated diseases. We synthesised 4 HaloPROTAC compounds, which can act as useful tools for biologists in understanding the proteindynamics within the mitochondria. Designed for the HaloTag system, these compounds consist of a chloroalkane chain and the ClpP binder, which are bound together by polyethylene glycol (PEG) linkers of varying lengths. Synthesised compounds were completed with yields ranging from 39 – 60% and LCMS providing mass confirmation.