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Title [sv]
Undersökande av bakteriella toxiner substratspecificitet via Reaktivt Protein - Proteom Profilering RP3
Title [en]
Exploring bacterial toxin substrate specificity via Reactive Protein - Proteome Profiling “ RP3 ”
Abstract [en]
This project has the purposeto conceptualize Reactive Protein - Proteome Profiling“ RP3”.The goal is to create a robust method for absolute substrate profiling of bacterial toxins against cellular target proteins employing nucleotide co-substrates resulting in covalent ternary complexes. During infection, pathogenic bacteria tweak their eukaryotic hosts by translocating toxins. Toxins display catalytic activity against host cell proteins, targeting key functions, often using a metabolite as co-substrate. No tools exist for the proteomic evaluation of the absolute substrate profile of a given toxin. Non-covalent affinity enrichment gives biased results depending on the reagent and relative target abundance. We will develop a new concept based on covalent (and subsequently cleavable) capture of the target proteins by the reactive co-substrate of the toxin. We have demonstrated proof of concept for the RP3workflow. Mutants (X to C) in the nucleotide-binding pocket of the toxin IbpA (H. somni) were modified with ATP-analogues carrying an electrophile. The binary complex forms the ternary covalent complex with the IbpA-substrate cdc42 in vitro. For ADP-ribosylation, Pertussis toxin captures Giain vitrousing NAD+analogues. Altogether, this suggests that the RP3concept is valid and practicable. The project is highly significant for the infection biology field in terms of profiling toxins, but also to the chemical biology community, by moving ABPP to macromolecules via RP3
Principal InvestigatorHedberg, Christian
Coordinating organisation
Umeå University
Funder
Period
2020-01-01 － 2023-12-31
National Category
Organic Chemistry
Identifiers
DiVA, id: project:2147Project, id: 2019-05384_VR

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