Towards the development of a visible light-enabled histidine-selective protein modification strategy using diazo sulfonium-based probes

Recognition-driven labelling techniques such as affinity/activity-based probes or photoaffinity labelling (PAL) are invaluable tools for the interrogation and study of biological processes in cellulo. Among those, PAL has proven to be the most powerful method, supplemented by the temporal control of its photoactivation step. However, the UV irradiation required for reagent activation and non-selective reactivity of the generated intermediates often results in low photo cross-linking yields and numerous off-target reactions. Therefore, the development of a more selective strategy using visible-light mediated photocatalysis appears a promising alternative. This thesis describes the efforts towards the development of a novel ligand-mediated strategy using visible light-activated -diazo sulfonium-based probes for the modification of histidine residues. Histidines represent attractive amino acid targets by virtue of their naturally low abundance but recurrent presence in protein active sites and pivotal biological roles. In addition, the design of a histidine-selective strategy constitutes an ideal opportunity to provide a complementary approach to the histidine bioconjugation toolkit given the lack of robust methods. Firstly, the biocompatibility and functional group tolerance of the reaction are demonstrated on short peptidic systems. Subsequent studies towards the application of the reaction on a complex ligand-protein system in vitro are also described. Owing to the modularity of the photoreactive a-diazo sulfonium motif employed, a broad variety of probes were designed. Finally, the research performed towards the development of an analogous affinity-guided catalyst strategy using an iridium-containing ligand and small diazo sulfonium trapping reagents is reported.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2022

Author dc:creator

• Folliet, Sarah

Advisor dc:contributor.advisor

• Gaunt, Matthew

• photoaffinity labelling
• affinity/activity-based probes
• photocatalysis
• bioconjugation
• histidine
• amino acids

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/1a2df5a2-9125-4e33-8a4c-07e55e9006dd/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.76846

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/329398