The self-assembly of nanoarchitectures via protein-ligand interactions

Nature has evolved proteins that can spontaneously self-assemble to create complex structures such as virus particles and molecular motors. The fields of bionanoscience and synthetic biology are based on the concept that by combining biological building blocks with synthetic molecules it will be possible to construct novel nanoarchitectures and machines that can do useful work. This thesis describes strategies that have been developed using protein-ligand interactions to construct nanoscale assemblies.
The B-subunit of cholera toxin (CTB) has been site-specifically modified at the N-teminus, by oxime ligation, with carbohydrate ligands and the assembly of these modified proteins was observed. A W88E non-binding mutant of CTB was made and modified with GM1os ligands. The interaction of this pentavalent protein-based ligand with wild-type CTB has been investigated and showed the formation of a protein heterodimer.
The CTA2-subunit of the cholera toxin AB5 complex was also modified at the N-terminus enzymatically with depsipeptides via sortase ligation and by oxime ligation. Biotin ligands have been covalently attached and the formation of a 2:1 complex with streptavidin was observed.
The structures and assemblies demonstrated herein have been analysed and characterised by a range of analytical and biophysical techniques including dynamic light scattering, analytical ultracentrifugation and isothermal titration calorimetry.