Human peroxiredoxin 3: the shape-shifting peroxidase as a versatile protein tecton
| dc.contributor.author | Yewdall, N. Amy | |
| dc.date.accessioned | 2017-08-20T21:52:24Z | |
| dc.date.available | 2017-08-20T21:52:24Z | |
| dc.date.issued | 2017 | en |
| dc.description.abstract | The biological realm contains numerous examples of nano-scale molecules that can self-assemble into a diverse array of architectures, making them attractive building blocks (or tectons) for applications in bionanotechnology. Proteins are one such biological molecule able to assemble into various three-dimensional structures. Exploring the mechanism and conditions in which these protein structures form is not only useful for the understanding of its biological role, but is also a prerequisite for their use in rational materials design. Human peroxiredoxin 3 (HsPrx3) are ubiquitous antioxidant proteins that can form a plethora of protein architectures: from homodimers that reversibly assemble into dodecameric rings (or toroids), and rings that can further associate into protein tubes. This thesis examines the high molecular weight protein tube structure of HsPrx3 (Chapter 2) and its assembly mechanism (Chapter 3). A 2.8 Å crystal structure of HsPrx3 was elucidated for the first time and was displayed as a short tube composed of three rings. This structure, together with a cryo-electron microscopy reconstruction obtained with collaborators, enabled a novel hypothesis for the biological role of these protein tubes as having a self-associating chaperone function. Using native mass spectrometry, protein tube formation was demonstrated to be formed via a non-commutative mechanism. Protein tube formation was also shown to be reversible, increasing the appeal of HsPrx3 proteins as tectons for bionanotechnology. HsPrx3 proteins react with hydrogen peroxide and upon oxidation, the reduced dodecameric rings disassemble into oxidised homodimers. The relationship between this quaternary structural switch and peroxidase activity was investigated (Chapter 4). Point mutations at the dimer-dimer interface were generated, creating an obligate dimer (S75E HsPrx3) and a stabilised toroid (S78C HsPrx3). Intriguingly, the obligate dimer was minimally active, suggesting that the ring structure is important, but not vital, for active site positioning. This raises interesting questions as to the biological function of this redox-induced structural change. On the other hand, the stabilised toroid was crystallised and the 2.4 Å structure provided a detailed understanding of the interactions that stabilise the dimer-dimer interface. S78C HsPrx3 will be a useful tecton as componentry for future applications. Having gained a deeper understanding of HsPrx3 self-assembly, functionalisation of the protein surface with novel chemistries was explored (Chapter 5). An unnatural amino acid, p-azidophenylalanine, was chosen for in vivo incorporation into HsPrx3 via an E. coli expression system. Although, not entirely successful, this marks a promising initial venture at functionalising HsPrx3. | en |
| dc.identifier.uri | http://hdl.handle.net/10092/14135 | |
| dc.identifier.uri | http://dx.doi.org/10.26021/5705 | |
| dc.language | English | |
| dc.language.iso | en | |
| dc.publisher | University of Canterbury | en |
| dc.rights | All Rights Reserved | en |
| dc.rights.uri | https://canterbury.libguides.com/rights/theses | en |
| dc.title | Human peroxiredoxin 3: the shape-shifting peroxidase as a versatile protein tecton | en |
| dc.type | Theses / Dissertations | en |
| thesis.degree.discipline | Biochemistry | en |
| thesis.degree.grantor | University of Canterbury | en |
| thesis.degree.level | Doctoral | en |
| thesis.degree.name | Doctor of Philosophy | en |
| uc.bibnumber | 2519602 | |
| uc.college | Faculty of Science | en |