Wilkins, Laura Elizabeth (2018) Bioorthogonal nanomaterials to probe adhesive protein function. PhD thesis, University of Warwick.

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Abstract

Tools to understand the mechanisms of protein binding to cell surface proteins are of interest to enable the manipulation of such proteins, such as for therapy and drug delivery. In particular, bioorthogonal techniques are of interest as they can be used to probe protein structure and function in their native environment with minimal structural perturbation, as described in Chapter 1. This work seeks to develop such bioorthogonally-designed materials (polymers and nanoparticles) in order to study adhesive proteins.

In Chapter 2, using controlled radical polymerisation and “click”-like chemistries, doubly-functionalised glycopolymers with sequential variation in carbohydrate density, side chain length and secondary functionality were synthesized to probe the binding activity of an adhesive pathogenic toxin (Vibrio cholerae toxin subunit B). This revealed a new approach whereby sterically large secondary units enabled selectivity towards a toxin to be introduced.

In chapter 3, a small (<5 nm diameter) gold nanoparticle (AuNP) scaffold was used as the basis for the development of a dendrimeric ice binding protein (antifreeze protein type III). Two different bioorthogonal immobilization strategies were developed to probe AFP capture to NP surfaces. Optimum AFP activity was observed only for covalent SNAP-tag conjugates. Furthermore, this was critically compared to a fullysynthetic AFP mimic; poly(vinyl alcohol) coated gold nanoparticles.

Chapter 4 reports a detailed investigation into the function of adhesive protein fibres from an insect pathogen, using the nanoparticle tools developed here. Recombinant his-tagged fibres were immobilized and evaluated for whole-cell binding to a range of cell lines, and also lipid and glycan arrays. Furthermore, proteomics used to identify insect and mammalian cell protein targets. This resulted in the first ever description of Pvc13 tail fibre protein function, which may help elucidate its pathogenicity and a potential application in drug delivery.

In summary, novel nanomaterials were developed in order to probe adhesive protein structure and function. These materials must be delicately designed to avoid an impact on native protein function, and to exploit the material properties to their full effect.

Item Type:	Thesis (PhD)
Subjects:	Q Science > QD Chemistry Q Science > QP Physiology
Library of Congress Subject Headings (LCSH):	Nanostructured materials -- Synthesis, Bioconjugates, Protein binding, Glycoproteins -- Synthesis, Dendrimers
Official Date:	September 2018
Dates:	Date Event September 2018 Published
Institution:	University of Warwick
Theses Department:	Department of Chemistry
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Gibson, Matthew I.; Waterfield, Nicholas R.
Format of File:	pdf
Extent:	xxxii, 284 leaves: illustrations, charts
Language:	eng

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