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This volume of Methods in Enzymology looks at Protein Engineering for Therapeutics. The chapters provide an invaluable resource for academics, researchers and students alike. Wi… Read more
LIMITED OFFER
Immediately download your ebook while waiting for your print delivery. No promo code needed.
This volume of Methods in Enzymology looks at Protein Engineering for Therapeutics. The chapters provide an invaluable resource for academics, researchers and students alike. With an international board of authors, this volume is split into sections that cover subjects such as Peptides, and Scaffolds
Volume in series
Preface
Stapled Peptides for Intracellular Drug Targets
1. Introduction
2. All-Hydrocarbon Stapled α-Helical Peptides
3. The Design of Stapled Peptides
4. Stapled Peptide Synthesis
5. Olefin Metathesis
6. N-terminal and Internal Modifications
7. Cleavage of Stapled Peptides from the Solid Support
8. Stapled Peptide Purification
9. Biophysical Characterization
10. Cell Permeability
11. In Vitro Target Interaction and Activity Assays
12. In Vivo Efficacy
13. Strategies for Stapled Peptide Optimization
14. Summary
Acknowledgments
Mapping of Vascular ZIP Codes by Phage Display
1. Introduction
2. Methods
3. Concluding Remarks and Perspectives
Acknowledgments
Engineering Cyclic Peptide Toxins
1. Introduction
2. Peptide Design
3. Peptide Synthesis
4. Structural Analysis
5. Stability Assays
6. Special Considerations for Cyclic Peptides
Acknowledgments
Peptide Discovery Using Bacterial Display and Flow Cytometry
1. Introduction
2. Protocols
3. Conclusions
Acknowledgments
Designed Ankyrin Repeat Proteins (DARPins)
1. Introduction
2. Applications of DARPins
3. Protocols for DARPins in Biomedical Applications
Acknowledgment
Target-Binding Proteins Based on the 10th Human Fibronectin Type III Domain (10Fn3)
1. Introduction
2. Library Design
3. Choice of Selection Platform
4. Phage Display, mRNA Display, and Yeast-Surface Display of 10Fn3-Based Libraries
5. Production
6. Conclusion
Acknowledgments
Anticalins
1. Introduction
2. Cloning and Expression of Lipocalins and Anticalins in E. coli
3. Construction of a Genetic Anticalin Library
4. Preparation and Selection of a Phage Display Library for Anticalins
5. Preparation and Selection of a Bacterial Surface Display Library for Anticalins
6. Colony Screening for Anticalins with Specific Target-Binding Activity
7. Screening for Anticalins with Specific Target-Binding Activity Using Microtiter Plate Expression in E. coli
8. Measuring Target Affinity of Anticalins in an ELISA
9. Measuring Target Affinity of Anticalins via Surface Plasmon Resonance
10. Application of Anticalins in Biochemical Research and Drug Development
T Cell Receptor Engineering
1. Introduction
2. Stability and Affinity Engineering of T Cell Receptors by Yeast Surface Display
3. Affinity Engineering and Selection of T Cell Receptors by T Cell Display
4. Expression, Purification, and Applications of Soluble scTv Proteins
5. Recipes for Media and Buffers
Acknowledgments
Engineering Knottins as Novel Binding Agents
1. Introduction
2. Knottins as Scaffolds for Engineering Molecular Recognition
3. Engineering Knottins by Yeast Surface Display
4. Knottin Library Construction
5. Screening Yeast-Displayed Knottin Libraries
6. Knottin production by chemical synthesis or recombinant expression
7. Cell binding assays
8. Summary
Acknowledgments
Practical Theoretic Guidance for the Design of Tumor-Targeting Agents
1. Introduction
2. What Molecular Size Is Best for Tumor Uptake?
3. Will Targeting Increase Nanoparticle Accumulation in a Tumor?
4. How Does Affinity Affect Biodistribution?
5. What Dose Is Necessary in Order to Overcome the “Binding Site Barrier”?
6. Conclusions
Reengineering Biopharmaceuticals for Targeted Delivery Across the Blood–Brain Barrier
1. Introduction
2. Blood–Brain Barrier Receptor-Mediated Transport and Molecular Trojan Horses
3. Reengineering Recombinant Proteins for Targeted Brain Delivery
4. Genetic Engineering of Expression Plasmid DNA Encoding IgG Fusion Proteins
5. Pharmacokinetics and Brain Uptake of IgG Fusion Proteins
6. CNS Pharmacological Effects of IgG Fusion Proteins
7. Immune Response Against IgG Fusion Proteins
8. Summary
Engineering and Identifying Supercharged Proteins for Macromolecule Delivery into Mammalian Cells
1. Introduction
2. Methods
3. Conclusion