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Ubiquitin and Protein Degradation, Part A will cover high level purification, bioinformatics analysis and substrate identification of the major proteins involved in protein degradation. The chapters are highly methodological and focus primarily on purification and analysis.
E1 Enzymes E2 Enzymes E3 Enzymes Proteasomes Isopeptidases
Biochemists, biophysicists, cell biologists, molecular biologists, geneticists, developmental biologists
Section I – E1 Enzymes Ch 1 High level expression and purification of recombinant E1 enzyme Ch 2 Expression, purification and characterization of the E1 for human NEDD8, the heterodimeric APPBP1-UBA3 complex Ch 3 A FRET-based assay to study SUMO modification in solution Ch 4 Properties of the ISG15 E1 enzyme UbE1L
Section II – E2 Enzyme Ch 5 Purification and properties of the Ubiquitin Conjugating Enzymes Cdc34 and Ubc13-Mms2 Ch 6 Eexpression, purification, and properties of Ubc4/5 family E2 enzymes Ch 7 In vitro systems for Nedd8 conjugation by Ubc12 Ch 8 Purification and activity assays for Ubc9, the ubiquitin conjugating enzyme for the small ubiquitin-like modifier SUMO Ch 9 High-througput assay to monitor Formation of the E2-Ubiquitin thioester intermediate
Section III – E3 Enzyme Ch 10 Expression and Evaluation of RING Finger proteins Ch 11 Expression and assay of HECT domain ligases Ch 12 High Level Expression and Purification of Recombinant SCF Ubiquitin Ligase Ch 13 In vitro reconstitution of SCF substrate ubiquitination with purified proteins Ch 14 Expression and assay of glycoprotein-specific ubiquitin ligases Ch 15 Affinity purification and assay of APC/C on p13 Suc1 Ch 16 Large-Scale Purification of the Vertebrate Anaphase-Promoting Complex/Cyclosome Ch 17 Purification and assay of the budding yeast Anaphase Promoting Complex Ch 18 Enzymology of the Anaphase-Promoting Complex Ch 19 Identification of Cell Cycle Dependent Phosphorylation Sites on the Anaphase-Promoting Complex/Cyclosome by Mass Spectrometry Ch 20 Purification and assay of Mad2: A Two-state Inhibitor of APC/C Ch 21 Identification, expression and assay of an oxidation-
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- © Academic Press 2005
- 29th August 2005
- Academic Press
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Howard Hughes Medical Institute, Caltech, Pasadena, CA, USA
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