Heterologous Gene Expression in E.Coli: Methods and Protocols

Part I: High-Throughput Cloning, Expression Screening, and Optimization

1. Recombinant Protein Expression in E. coli: A Historical Perspective

Opher Gileadi

2. N- and C-Terminal Truncations to Enhance Protein Solubility and Crystallization: Predicting Protein Domain Boundaries with Bioinformatics Tools

Christopher D.O. Cooper and Brian D. Marsden

3. Harnessing the Profinity eXactTM System for Expression and Purification of Heterologous Proteins in E. coli

Yoav Peleg, Vadivel Prabahar, Dominika Bednarczyk, and Tamar Unger

4. ESPRIT: A Method for Defining Soluble Expression Constructs in Poorly Understood Gene Sequences

Philippe J. Mas and Darren J. Hart

5. Optimizing Expression and Solubility of Proteins in E. coli Using Modified Media and Induction Parameters

Troy Taylor, John-Paul Denson, and Dominic Esposito

6. Optimization of Membrane Protein Production Using Titratable Strains of E. coli

Rosa Morra, Kate Young, David Casas-Mao, Neil Dixon, and Louise E. Bird

7. Optimizing E. coli-Based Membrane Protein Production Using Lemo21(DE3) or pReX and GFP-Fusions

Grietje Kuipers, Markus Peschke, Nurzian Bernsel Ismail, Anna Hjelm, Susan Schlegel, David Vikstr?m, Joen Luirink, and Jan-Willem de Gier

8. High Yield of Recombinant Protein in Shaken E. coli Cultures with Enzymatic Glucose Release Medium EnPresso B

Part II: Protein Purification and Solubility Enhancement

9. A Generic Protocol for Purifying Disulphide-Bonded Domains and Random Protein Fragments Using Fusion Proteins with SUMO3 and Cleavage by SenP2 Protease

H?seyin Besir

10. A Strategy for Production of Correctly Folded Disulfide-Rich Peptides in the Periplasm of E. coli

Natalie J. Saez, Ben Cristofori-Armstrong, Raveendra Anangi, and Glenn F. King

11. Split GFP Complementation as Reporter of Membrane Protein Expression and Stability in E. coli: A Tool to Engineer Stability in a LAT Transporter

Ekaitz Errasti-Murugarren, Arturo Rodr?guez-Banqueri, and Jos? Luis V?zquez-Ibar

12. Acting on Folding Effectors to Improve Recombinant Protein Yields and Functional Quality

Ario de Marco

13. Protein Folding Using a Vortex Fluidic Device

Joshua Britton, Joshua N. Smith, Colin L. Raston, and Gregory A. Weiss

14. Removal of Affinity Tags with TEV Protease

Sreejith Raran-Kurussi, Scott Cherry, Di Zhang, and David S. Waugh

Part III: