Encapsulin engineering for metal nanoparticle production

Abstract

The COVID-19 pandemic has highlighted the importance of technology in our society and emphasised the fragility of supply lines. Governments have identified metal supply chains as a security concern, highlighted by the World Trade Organisation. At the same time, the shift towards a greener economy, which places a significant emphasis on metals, has led to an increase in prices, with some metals reaching ten-year highs. To move towards a circular economy, it is essential to develop a sustainable method for recovering metal ions from waste streams. This study aims to achieve this by using synthetic biology to recover and upcycle metal ions as metal nanoparticles through protein nanocompartments called encapsulins.

Encapsulins can limit the size and shape of nanoparticles, increase their stability, and be directly functionalised.

To test encapsulins with different metal binding specificities, a modular DNA assembly kit was modified for rapid cloning. Each variant was expressed in vivo and purified for in vitro experiments. Escherichia coli cells had enhanced resistance to silver ions and improved metal recovery for silver and gold ions in vivo when expressing the encapsulin variants. In vitro experiments and electron microscopy confirmed that purified encapsulins could assemble and precipitate metal ions as nanoparticles. Molecular dynamics simulations were used to understand how ions pass through encapsulin pores and how they were distributed within the encapsulin. Point mutations were made to the encapsulin pore and screened in silico with the aim of occluding metal ions based on size. A DNA library was constructed to screen the encapsulin mutants, including the in silico mutants, for their ability to occlude metal ions in vivo.