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This book aims to advance the understanding of deubiquitinases (DUBs) and DUB-like enzymes. Chapters detail methods used to identify, classify,and biochemically characterize DUBs along with approaches that enable both the determination and alteration of DUB biological function.. Written in the format of the highly successful Methods in Molecular Biology series, each chapter includes an introduction to the topic, lists necessary materials and reagents, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols.
Authoritative and cutting-edge, Deubiquitinases: Methods and Protocols aims to be a useful and practical guide to new researchers and experts looking to expand their knowledge.
List of contents
Characterization of Deubiquitinase Catalytic State Using a Structure-Based Approach.- Synthetic Diubiquitin Fluorogenic Substrates to Study DUBs.- Fluorometric Characterization of DUB Activity: From Single Enzyme Reactions to Live Intact Cell.- Using Activity-Based Proteomics for the Quantification of Deubiquitinases in Animal Tissue.- Design and Synthesis of Ubiquitin-Based Chemical tools with Unnatural Amino Acids for Selective Detection of Deubiquitinases.- High Throughput Assay for Characterizing Rpn11 Deubiquitinase Activity.- Activity-Based Protein Profiling (ABPP) for Cellular Deubiquitinase (DUB) and Inhibitor Profiling at Deep and High-Throughput Levels.- High-Throughput MALDI-TOF Mass-Spectrometry-based Deubiquitylating Enzyme Assay for Drug Discovery.- Bioinformatical Approaches to The Discovery and Classification of Novel Deubiquitinases.- Application of Archaea in Deubiquitinase-Like Enzyme Discovery and Activity Assay.- Characterization of the Ubiquitin and ISG15 Deconjugase Activity of SARS-CoV-1 and 2 Papain-like Protease.- Next-generation Phage Display to Identify Peptide Ligands of Deubiquitinases.- Identification of Deubiquitinase Substrates in Xenopus Egg Extract.- Identification of Deubiquitinase Substrates in Saccharomyces cerevisiae by Systematic Overexpression.- Real-time Deubiquitination Assays using a Free-Ubiquitin Sensor.- A Method to Monitor Activity of SARS-CoV-2 Nsp3 from Cells.- Use of Deubiquitinase Fusion Proteins to Characterize Endocytic Trafficking in Yeast.
