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Type: Thesis
Title: Probing a Promiscuous Binding Pocket of the Proteasome
Author: Turner, Dion Joel Lammas
Issue Date: 2019
School/Discipline: School of Physical Sciences
Abstract: The proteasome is a multi-catalytic protease complex responsible for the degradation of unneeded, damaged or misfolded proteins. The proteasome is a validated target for the treatment of haematological malignancies such as multiple myeloma and mantle cell lymphoma, as demonstrated by the FDA approved proteasome inhibitors bortezomib, carfilzomib and ixazomib. These inhibitors, especially bortezomib, suffer from poor specificity and relatively high prevalence of resistance, therefore new inhibitors should be designed such that these characteristics improve. This thesis details probing of the relatively unexplored primed site binding channel of the β5 subunit of the proteasome with P2 extended proteasome inhibitors. This work indicates the primed site binding channel as a promiscuous target for interaction which may aid in increasing the specificity of proteasome inhibitors Chapter 1 introduces the structure and activity of the proteasome and its implications and relevance to human diseases. Inhibition of the proteasome by small molecule inhibitors is discussed, including the main classes, exemplary inhibitors, their mechanisms and applications. The primed site binding channel is then identified via examination of the crystal structure of the proteasome as a pocket which provides potential for new inhibitor-enzyme interactions. Chapter 2 details the design, synthesis and evaluation of inhibitors 2.01-2.04 which probe the extent of the promiscuity of the primed site binding channel. The collection of published inhibitors which are known to, or are likely to, occupy the primed binding sites demonstrate the primed site binding channel as promiscuous regarding the substituents it accepts. The P2 residue of bortezomib was identified as providing an access point to the primed binding sites. Imidazolyl and phenyl substituents were demonstrated to be accommodated by the primed site binding channel, with greater potency found for longer extensions into the pocket, or inhibitors with a phenyl substituent within the pocket. Chapter 3 describes further probing of the primed site binding channel with the azobenzene-containing proteasome inhibitor 3.01, which can be converted between cis- and trans-enriched isomeric states using light. The azobenzene substituent was placed at the P2 position of a bortezomib-inspired inhibitor and allowed probing of the primed binding sites with greater conformation predictability. Remarkably, despite significant change in conformation between the cis and trans isomers, there is little difference between the low nanomolar range potencies of the isomeric states. This further indicates the significant promiscuity of the primed site binding channel. Chapter 4 presents the evaluation of inhibitors 2.01-2.04 and the thermally adapted state of 3.01 alongside bortezomib against bovine α-chymotrypsin to examine the specificity of such inhibitors. Primed site-occupying inhibitors 2.01 and 2.04 demonstrate more than 2.5 times greater specificity towards the β5 subunit of the proteasome over α-chymotrypsin. This result indicates occupying the primed site binding channel as an effective strategy of improving proteasome inhibitor specificity, which may be critical in improving upon the currently available proteasome
Advisor: Abell, Andrew
Avery, Thomas
Dissertation Note: Thesis (MPhil) -- University of Adelaide, School of Physical Sciences, 2020
Keywords: Proteasome
Azobenzene multiple myeloma
drug development
medicinal chemistry
organic chemistry
Provenance: This electronic version is made publicly available by the University of Adelaide in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material you wish to be removed from this electronic version, please complete the take down form located at:
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