A desthiobiotin labelled NAD + analogue to uncover Poly(ADP-ribose) polymerase 1 protein targets.

Autor: Rieth S; Department of Chemistry, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany., Spliesgar D; Department of Chemistry, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany., Orth J; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Department of Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany., Lehner M; Department of Chemistry, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany., Kasprzyk R; Department of Chemistry, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany., Stengel F; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Department of Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany., Marx A; Department of Chemistry, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.; Konstanz Research School Chemical Biology, University of Konstanz, Universitätsstraße10, 78457, Konstanz, Germany.
Jazyk: angličtina
Zdroj: Chembiochem : a European journal of chemical biology [Chembiochem] 2024 Mar 01; Vol. 25 (5), pp. e202300797. Date of Electronic Publication: 2024 Jan 31.
DOI: 10.1002/cbic.202300797
Abstrakt: ADP-ribosylation is a post-translational modification catalyzed by the enzyme family of polyadenosine diphosphate (ADP)-ribose) polymerases (PARPs). This enzymatic process involves the transfer of single or multiple ADP-ribose molecules onto proteins, utilizing nicotinamide adenine dinucleotide (NAD + ) as a substrate. It, thus, plays a pivotal role in regulating various biological processes. Unveiling PARP-selective protein targets is crucial for a better understanding of their biological functions. Nonetheless, this task proves challenging due to overlapping targets shared among PARP family members. Therefore, we applied the "bump-and-hole" strategy to modify the nicotinamide binding site of PARP1 by introducing a hydrophobic pocket ("hole"). This PARP1-mutant binds an orthogonal NAD + (Et-DTB-NAD + ) containing an ethyl group ("bump") at the nicotinamide moiety. Furthermore, we added a desthiobiotin (DTB) tag directly to the adenosine moiety, enabling affinity enrichment of ADP-ribosylated proteins. Employing this approach, we successfully identified protein targets modified by PARP1 in cell lysate. This strategy expands the arsenal of chemically modified NAD + analogs available for studying ADP-ribosylation, providing a powerful tool to study these critical post-translational modifications.
(© 2024 The Authors. ChemBioChem published by Wiley-VCH GmbH.)
Databáze: MEDLINE
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