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vi CHAPTER 1; GENERAL INTRODUCTION 1 Dissertation Organization 1 Refolding of Recombinant Proteins 1 Analytical Techniques for Monitoring Protein Refolding 2 Capillary Zone Electrophoresis-Laser Induced Tryptophan Fluorescence Detection for Monitoring Refolding of a Large Multimeric Protein 3 Capillary Isoelectric Focusing-Electrospray Ionization Mass Spectrometry for Monitoring Disulfide Bond Induced Protein Refolding 8 CHAPTER 2; MONITORING THE REFOLDING PATHWAY FOR A LARGE MULTIMERIC PROTEIN USING CAPILLARY ZONE ELECTROPHORESIS 13 Abstract 13 Introduction 14 Experimental 17 Capillary Zone Electrophoresis-Laser Induced Fluorescence Detection 17 Protein Denaturation and Refolding 19 Slab Gel Electrophoresis 20 Tryptophan Fluorescence Spectrophotometry 20 Chemicals 21 Results and Discussion 21 Monitoring Tailspike Refolding Using Capillary Zone ElectrophoresisLaser Induced Fluorescence Detection 21 Monitoring Tailspike Refolding Using Tryptophan Fluorescence Spectrophotometry 26 Monitoring Tailspike Refolding Using Slab Gel Electrophoresis 27 Temperature Dependence of Productive vs. Aggregation Pathways 27 Acknowledgments 29 References 29 CHAPTER 3: INVESTIGATING TEMPERATURE EFFECTS ON REFOLDING AND AGGREGATION OF A LARGE MULTIMERIC PROTEIN USING CAPILLARY ZONE ELECTROPHORESIS 38 Abstract 38 Introduction 39 Experimental 42 CZE-LIFD 42 Protein Denaturation and Refolding 43 Slab Gel Electrophoresis 44

Monitoring protein refolding using diverse separation and spectroscopy techniques