D-protein and D-peptide technology in applied research

Lander, Alexander 2022. D-protein and D-peptide technology in applied research. PhD Thesis, Cardiff University. Item availability restricted.

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Abstract

Chemical protein synthesis provides access to entire D-protein enantiomers, with unique utility in applied research. Firstly, proteins may be crystallized from a racemic mixture of L- and Denantiomers, known as racemic protein crystallography, providing facile access to high resolution X-ray structures. In this thesis, to understand how membrane-active bacteriocins have acquired their antibacterial properties, an analysis based on racemic protein crystallography (0.9–1.2 Å) of two key representatives, aureocin a53 (AucA) and lacticin Q (LnqQ) was conducted. Through structural analysis and systematic residue substitutions, conserved surface tryptophans appeared to play important roles in coordination of lipid phosphate and were critical for antibacterial activity. Additional tryptophans in AucA were also involved in forming an oligomeric assembly for stability enhancement. Collectively, racemic protein crystallography shed light on the molecular interactions of tryptophans, demonstrating how a bacteriocin delivers its antibacterial properties. Secondly, D-enantiomers of protein drug targets can be used in mirror-image phage display (MIPD), allowing discovery of non-proteolytic D-peptide ligands as lead candidates. Development of a D-peptide capable of targeting tumor necrosis factor receptor I (TNFR1) activation could be a beneficial solution in controlling inflammation caused by cytokine storm. However, containing 144 residues and 12 disulfide bonds, synthesis of the D-enantiomer for MIPD would be a significant challenge. Here, isolation of the mid-chain, cytokine binding domain (TNRCD2) is reported, and its enantiomer enabled MIPD to discover a cyclic peptide binder with low micromolar affinity. Additionally, the use of computational methods to develop D-protein ligands based only on the target structure was explored.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Chemistry
Date of First Compliant Deposit:	28 March 2023
Last Modified:	28 Mar 2024 02:30
URI:	https://orca.cardiff.ac.uk/id/eprint/158140

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