Installing functional metal-binding sites into designer proteins

This project will involve the computational design of alpha-helical bundle and barrel proteins as scaffolds to accommodate catalytically active metal sites. The aim is to use these to catalyse chemical reactions for greener synthetic chemistry. This will help to address a longstanding problem in protein design, as designing protein structure is well advanced, but designing functions that rival natural proteins is challenging. The proposed work will involve using de novo coiled-coil proteins because they are stable, well understood, and robust to mutation. Gold will be the initial target metal for incorporation, because it is able to catalyse reactions under mild reaction conditions and considered non-toxic compared with alternatives such as palladium.

This research will begin with the analysis of natural proteins in the Protein Data Bank and studying the way in which these bind metals of interest. Following this, computational design of protein scaffolds and metal-binding sites will be conducted, facilitated by AlphaFold3 for generating and visualising the protein models. Next, in silico evaluations of metal binding will predict and examine suitable binding sites in detail. After the computational design and metal-binding studies, synthetic genes for candidate protein designs will be expressed in bacteria. The resulting proteins will be screened for metal binding. This will be monitored using analytical techniques such as spectroscopy and mass spectrometry. Ultimately, experimental structures will be determined by X-ray crystallography. After successfully binding the metals of interest, focus will turn to incorporating and testing catalytic activity for a model reactions. At each stage of the experimental work, the protein scaffolds and metal binding will be assessed and optimised as needed to improve structure, stability, and catalytic properties for the overarching goal of generating de novo metalloproteins with the utility as tools for synthetic chemistry.