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Publication Abstract from PubMed

Modification of proteins with polyubiquitin chains is a key regulatory mechanism to control cellular behavior and altera-tions in the ubiquitin system are linked to many diseases. Linear (M1-linked) polyubiquitin chains play pivotal roles in several cellular signalling pathways mediating immune and inflammatory responses and apoptotic cell death. These chains are formed by the linear ubiquitin chain assembly complex (LUBAC), a multi-protein E3 ligase that consists of 3 subunits, HOIP, HOIL-1L and SHARPIN. Herein, we describe the discovery of inhibitors targeting the active site cysteine of the catalytic subunit HOIP using fragment-based covalent ligand screening. We report the synthesis of a diverse library of electrophilic fragments and demonstrate an integrated use of protein LC-MS, biochemical ubiquitination assays, chemi-cal synthesis and protein crystallography to enable the first structure-based development of covalent inhibitors for an RBR E3 ligase. Furthermore, using cell-based assays and chemoproteomics we demonstrate that these compounds effec-tively penetrate mammalian cells to label and inhibit HOIP and NF-kappaB activation, making them suitable hits for the devel-opment of selective probes to study LUBAC biology. Our results illustrate the power of fragment-based covalent ligand screening to discover lead compounds for challenging targets, which holds promise to be a general approach for the de-velopment of cell-permeable inhibitors of thioester-forming E3 ubiquitin ligases.

Fragment-based covalent ligand screening enables rapid discovery of inhibitors for the RBR E3 ubiquitin ligase HOIP.,Johansson H, Tsai YI, Fantom K, Chung CW, Kumper S, Martino L, Thomas DA, Eberl HC, Muelbaier M, House D, Rittinger K J Am Chem Soc. 2019 Jan 18. doi: 10.1021/jacs.8b13193. PMID:30657686^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.