Engineering protein cages with synthetic biology

Nanotechnology has the potential to revolutionise every facet of human life. One particularly exciting branch of nanotechnology involves the construction of nanodevices using protein cages. Protein cages are spherically shaped structures with large internal cavities. The research described in this thesis was conducted with the aim of rationalising the design and fabrication of protein cage-based nanodevices. Protein-based nanodevices are typically constructed by re-engineering naturally occurring protein chassis (e.g. ferritin). To rationalise the process of chassis selection, an online registry of protein cages, rings and tubes was designed and populated by computationally mining the Protein Data Bank. The resulting registry was made publically available to the research community through the website – www.nanodevice.build. The functionality of protein cage-based nanodevices can be augmented by packaging inorganic nanoparticles inside their internal cavities. The methods currently used to achieve this typically involve exposure to harsh conditions, which can cause irreversible damage to the protein cage. To address this, a strategy for efficiently packaging inorganic nanoparticles into protein cages under mild conditions was formulated and tested. These experiments were conducted using gold nanoparticles and a number of different protein cages (e.g. Bfr, FtnH and FtnL). Cholangiocarcinoma (CCA) is a deadly liver cancer for which current treatment options are limited. Therefore a CCA-targeting protein cage-based nanodevice was designed, constructed and experimentally evaluated. CCA-targeting was achieved in the context of the CCA cell line TFK-1 using an anti-mesothelin antibody as a targeting agent. Collectively, these three outputs provide a rational framework for selecting a protein cage chassis, loading it with a pre-fabricated inorganic nanoparticle and targeting the resulting device to a particular cell-type. It is hoped that by leveraging these three tools, synthetic biologists will be able to engineer a new generation of nanodevices.Open Acces