Understanding and engineering protein oligomers

Ahmed, Rochelle 2022. Understanding and engineering protein oligomers. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

Protein oligomerisation is a fascinating topic concerning the association of proteins to form a single multimeric complex, either naturally or artificially. The importance of protein association is highlighted by their prevalence, where protein oligomers are more prevalent in nature than monomers. They are critical for a wide range of biological processes such as providing diversity and specificity to physiological pathways, mediating gene expression, and the activity of enzymes and ion channels. They also play a key role in regulation of apoptosis and tumour formation. Thus, unveiling the structural and functional aspects of protein oligomerisation has acquired increasing importance. The first portion of this thesis examines the oligomerisation state of Bcl3- p50, a IkB - NF-kB protein complex evidenced to play a key role in breast cancer metastasis. A range of in silico, structural and biophysical techniques were used to understand the binding and stability of PPIs governing the protein complex (Chapter 3). A novel small molecule inhibitor of Bcl3 was also tested for its ability to block interactions between Bcl3 and p50. One key method that is widely used to study biomolecular interactions within cells is FRET (or Förster resonance energy transfer). However, the dependence of two fluorescent proteins limits this technique to monitoring heterodimerisation. Thus, the second aspect of this thesis sets out to engineer superfolder Green Fluorescent Protein (sfGFP) for use as a homodimer biosensor. This work was divided into separate sections. First, the design and incorporation of mutations into desired residue positions of sfGFP to alter the spectral properties (Chapter 4). Next, establishing PPIs between sfGFP subunits and detection of homodimerisation through resulting fluorescent changes (Chapter 5). Finally, fusing the engineered sfGFP to p50 to evidence its ability to detect homodimerisation (Chapter 6).

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Biosciences
Subjects:	Q Science > Q Science (General)
Date of First Compliant Deposit:	23 May 2023
Last Modified:	06 Jan 2024 04:21
URI:	https://orca.cardiff.ac.uk/id/eprint/159886

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