The Ubiquitin Proteasome System

Characterization of RING-Between-RING E3 Ubiquitin Transfer Mechanisms.- Single Turnover RING/U-Box E3-Mediated Lysine Discharge Assays.- Methods for NAD-Dependent Ubiquitination Catalyzed by Legionella pneumophila Effector Proteins.- Using In Vitro Ubiquitylation Assays to Estimate the Affinities of Ubiquitin-Conjugating Enzymes for their Ubiquitin Ligase Partners.- Competition Assay for Measuring Deubiquitinating Enzyme Substrate Affinity.- Enzymatic Assembly of Ubiquitin Chains.- Ubiquitin-Activated Interaction Traps (UBAITs): Tools for Capturing Protein-Protein Interactions.- Generating Intracellular Modulators of E3 Ligases and Deubiquitinases from Phage-Displayed Ubiquitin Variant Libraries.- Integrated Proteogenomic Approach for Identifying Degradation Motifs in Eukaryotic Cells.- A Method to Monitor Protein Turnover by Flow Cytometry and to Screen for Factors that Control Degradation by Fluorescence-Activated Cell Sorting.- E. coli Based Selection and Expression Systems for Discovery, Characterization, and Purification of Ubiquitylated Proteins.- Strategies to Trap Enzyme-Substrate Complexes that Mimic Michaelis Intermediates during E3-Mediated Ubiquitin-Like Protein Ligation.- Small-Angle X-Ray Scattering for the Study of Proteins in the Ubiquitin Pathway.- Methods for Preparing Cryo-EM Grids of Large Macromolecular Complexes.- Recombinant Expression, Unnatural Amino-Acid Incorporation, and Site-Specific Labeling of 26S Proteasomal Subcomplexes.- Native Gel Approaches in Studying Proteasome Assembly and Chaperones.- Measuring the Overall Rate of Protein Breakdown in Cells and the Contributions of the Ubiquitin-Proteasome and Autophagy-Lysosomal Pathways.- Methods to Rapidly Prepare Mammalian 26S Proteasomes for Biochemical Analysis.- Measurement of the Multiple Activities of 26S Proteasomes.- Exploring the Regulation of Proteasome Function by Subunit