Designed artificial protein heterodimers with coupled functions constructed using bio-orthogonal chemistry

Johnson, Rachel L., Blaber, Hayley G., Evans, Tomas, Worthy, Harley L., Pope, Jacob R. ORCID: https://orcid.org/0000-0001-6478-5378 and Jones, D. Dafydd ORCID: https://orcid.org/0000-0001-7709-3995 2021. Designed artificial protein heterodimers with coupled functions constructed using bio-orthogonal chemistry. Frontiers in Chemistry 9 , 733550. 10.3389/fchem.2021.733550

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Abstract

The formation of protein complexes is central to biology, with oligomeric proteins more prevalent than monomers. The coupling of functionally and even structurally distinct protein units can lead to new functional properties not accessible by monomeric proteins alone. While such complexes are driven by evolutionally needs in biology, the ability to link normally functionally and structurally disparate proteins can lead to new emergent properties for use in synthetic biology and the nanosciences. Here we demonstrate how two disparate proteins, the haem binding helical bundle protein cytochrome b562 and the β-barrel green fluorescent protein can be combined to form a heterodimer linked together by an unnatural triazole linkage. The complex was designed using computational docking approaches to predict compatible interfaces between the two proteins. Models of the complexes where then used to engineer residue coupling sites in each protein to link them together. Genetic code expansion was used to incorporate azide chemistry in cytochrome b562 and alkyne chemistry in GFP so that a permanent triazole covalent linkage can be made between the two proteins. Two linkage sites with respect to GFP were sampled. Spectral analysis of the new heterodimer revealed that haem binding and fluorescent protein chromophore properties were retained. Functional coupling was confirmed through changes in GFP absorbance and fluorescence, with linkage site determining the extent of communication between the two proteins. We have thus shown here that is possible to design and build heterodimeric proteins that couple structurally and functionally disparate proteins to form a new complex with new functional properties.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Biosciences Advanced Research Computing @ Cardiff (ARCCA)
Publisher:	Frontiers Media
ISSN:	2296-2646
Funders:	BBSRC
Date of First Compliant Deposit:	9 August 2021
Date of Acceptance:	22 July 2021
Last Modified:	05 Oct 2023 21:52
URI:	https://orca.cardiff.ac.uk/id/eprint/143239

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