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The Development of a Fluorescence‐Based Competitive Assay Enabled the Discovery of Dimeric Cyclic Peptide Modulators of Ubiquitin Chains
Author(s) -
Vamisetti Ganga B.,
Meledin Roman,
Nawatha Mickal,
Suga Hiroaki,
Brik Ashraf
Publication year - 2021
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.202013392
Subject(s) - ubiquitin , peptide , chemistry , conjugate , cyclic peptide , combinatorial chemistry , drug discovery , high throughput screening , peptide library , biochemistry , computational biology , peptide sequence , biology , gene , mathematical analysis , mathematics
Development of modulators targeting specific interactions of ubiquitin‐based conjugates with their partners is a formidable task since it requires a suitable screening assay and homogeneous ubiquitin conjugates. We developed a novel high‐throughput strategy for screening ligands for Lys48‐linked tetraubiquitin chain in a relatively simple, fast, and affordable manner. This approach combined with a state‐of‐the‐art toolbox of chemical protein synthesis and a specially optimized Cys deprotection protocol enabled us to design highly potent, Lys48‐linked tetraubiquitin chain selective “next generation” dimeric peptide modulators. The dimeric peptide exhibited cancer cell permeability and induced cell death with higher efficiency compared to its monocyclic analogue. These features make our dimeric peptide a promising candidate for further studies using in vivo models. Our assay can be adopted for other various ubiquitin chains in their free or anchored forms as well as conjugates for Ub‐like modifiers.