664.13 - First-in-class Deubiquitylase Inhibitors Reveal New Enzyme Conformations

Francesca Chandler (University of Leeds), Miriam Walden (University of Leeds), Poli Adi Narayana Reddy (The Wistar Institute), Joel Cassel (The Wistar Institute), Lillie Bell (University of Leeds), Lisa Campbell (University of Leeds), Martina Foglizzo (University of Leeds), Frank Sicheri (Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital), Joseph Salvino (The Wistar Institute), Roger Greenberg (Basser Center for BRCA, Perelman School of Medicine, University of Pennsylvania), Elton Zeqiraj (University of Leeds)

Cells maintain protein homeostasis by adding a small protein, ubiquitin, to regulate a variety of cellular processes, dictating protein activity, localisation or degradation. The addition of ubiquitin, known as ubiquitylation, is a reversible process making it a versatile post-translational modification aptly suited for cell signalling. Removal of ubiquitin is catalysed by deubiquitylating enzymes, commonly referred to as DUBs. BRCC36 isopeptidase complex (BRISC) is a multi-protein DUB complex which hydrolyses lysine-63-linked ubiquitin chains on Type I interferon receptors (IFNAR1/2), thus regulating interferon-dependent signalling. Therefore, BRISC-mediated inflammatory signalling amplification is a promising target for autoimmune disease drug development. We performed a high-throughput screen to identify small molecules which inhibit BRISC enzymatic activity. Employing an integrative structural biology approach (cryo-electron microscopy, native mass spectrometry, hydrogen-deuterium exchange mass spectrometry), complemented with biochemical assays, we have uncovered new enzyme conformations, revealing a remarkable mode of action for BRISC inhibitors. Exploring these key mechanisms will expand current knowledge of inflammatory signalling pathways and establish the use of DUB inhibitors as therapeutics to combat autoimmune disease and hyperactive cytokine signalling.

F.C. is supported by a Wellcome Trust four-year PhD studentship (219997/Z/19/Z). E.Z. is supported by a Sir Henry Dale Fellowship from the Wellcome Trust amp;amp; Royal Society (200523/Z/16/Z).