Concepts and Case Studies in Chemical Biology

Retaining the proven didactic concept of the successful "Chemical Biology - Learning through Case Studies", this sequel features 27 new case studies, reflecting the rapid growth in this interdisciplinary topic over the past few years.
Edited by two of the world's leading researchers in the field, this textbook introduces students and researchers to the modern approaches in chemical biology, as well as important results, and the techniques and methods applied. Each chapter presents a different biological problem taken from everyday lab work, elucidated by an international team of renowned scientists.
With its broad coverage, this is a valuable source of information for students, graduate students, and researchers working on the borderline between chemistry, biology, and biochemistry.

Autorentext
Herbert Waldmann obtained his PhD in organic chemistry from the University of Mainz, Germany, in 1985 working with Professor H. Kunz, after which he completed a postdoctoral appointment with Professor G. Whitesides at Harvard University (USA). He was appointed as Professor of Organic Chemistry at the University of Bonn (1991), full Professor of Organic Chemistry at the University of Karlsruhe (1993), and Director at the Max-Planck-Institute of Molecular Physiology Dortmund and Professor of Organic Chemistry at the University of Dortmund, Germany, in 1999. He has received numerous awards, e.g. the Otto-Bayer-Award (2001), the Max Bergmann Medal (2003), and recently the Emil-Fischer-Prize (2012). His research interests lie in bioactive compound development, target identification, and the chemical biology of Ras proteins.

Petra Janning studied chemistry at the University of Munster, Germany, and did her PhD thesis at the Institute for Analytical Sciences in Dortmund, Germany in 1995. After different positions where she worked in the area of analytical chemistry and on the borderline between chemistry and biology she joined the Max-Planck-Institute of Molecular Physiology, Dortmund, in the department of Professor H. Waldmann, where she is currently working. She is responsible for the Bioorganic Practical Course and the analytics in the department. She focused her work on mass spectrometry based methods in proteome research in particular in target protein identification of small molecules.

Inhalt

List of Contributors xvii

Introduction and Preface xxv

Abbreviations xxix

**1 Real-Time and Continuous Sensors of Protein Kinase Activity Utilizing Chelation-Enhanced Fluorescence 1
Laura B. Peterson and Barbara Imperiali

1.1 Introduction 1

1.2 The Biological Problem 1

1.3 The Chemical Approach 3

1.3.1 Chelation-Enhanced Fluorescence 3

1.3.2 -Turn-Focused Kinase Activity Sensors 7

1.3.3 Recognition-Domain-Focused Kinase Activity Sensors 7

1.3.4 Chimeric Kinase Activity Sensors 10

1.4 Chemical Biological Research/Evaluation 12

1.4.1 Kinetic Parameters 12

1.4.2 Assessing Kinase Selectivity 12

1.4.3 Kinase Profiling in Cell Lysates and Tissue Homogenates 14

1.5 Conclusions 14

References 15

**2 FLiK and FLiP: Direct Binding Assays for the Identification of Stabilizers of Inactive Kinase and Phosphatase Conformations 17
Daniel Rauh and Jeffrey R. Simard

2.1 Introduction The Biological Problem 17

2.1.1 Kinase Inhibitors Stabilizing Inactive Enzyme Conformations 17

2.1.2 Monitoring Conformational Changes upon Ligand Binding 19

2.2 The Chemical Approach 20

2.3 Chemical Biological Research/Evaluation 23

2.3.1 Finding the Unexpected 25

2.3.2 Targeting Protein Interfaces iFLiK 26

2.3.3 Screening Akt 27

2.3.4 Targeting Phosphatases FLiP 29

2.3.5 Lessons Learned from High-Throughput Screens 31

2.4 Conclusions 34

References 35

**3 Strategies for Designing Specific Protein Tyrosine Phosphatase Inhibitors and Their Intracellular Activation 37
Birgit Hoeger and Maja Köhn

3.1 Introduction The Biological Problem 37

3.1.1 Chemical Inhibition of Protein Tyrosine Phosphatase Activity 37

3.1.2 PTP1B as Inhibitor Target 40

3.2 The Chemical Approach 41

3.2.1 The Concept of Bivalent Ligands Development of a Specific PTP1B Inhibitor 41

3.2.2 Cell Permeability and Intracellular Activation of a Self-Silenced Inhibitor 43

3.2.3 A Prodrug Strategy to Gain Cell Permeability 44

3.3 Chemical Biological Research/Evaluation 45

3.3.1 An Affinity-Based ELISA Assay to Identify Potent Binders 45

3.3.2 Evaluation of Cell Permeability and Cellular Activity by Monitoring Insulin Receptor Signaling 47

3.4 Conclusions 47

References 48

**4 Design and Application of Chemical Probes for Protein Serine/Threonine Phosphatase Activation 51
Yansong Wang and Maja Köhn

4.1 Introduction 51

4.2 The Biological Problem 52

4.3 The Chemical Approach 54

4.4 Chemical Biological Research/Evaluation 57

4.4.1 Selectivity of PDPs toward PP1 over PP2A and PP2B 57

4.4.2 Studying the Functions of PP1 in Mitosis with PDPs 58

4.4.3 Studying the Functions of PP1 in Ca 2+ Signaling with PDPs 59

4.5 Conclusion 60

References 60

**5 Autophagy: Assays and Small-Molecule Modulators 63
Gemma Triola

5.1 Introduction 63

5.2 The Biological Problem 65

5.2.1 Assays 66

5.2.2 Small-Molecule Modulators of Autophagy 67

5.3 The Chemical Approach 68

5.3.1 Assays 68

5.4 Chemical Biological Evaluation 71

5.5 Conclusion 80

References 80

**6 Elucidation of Protein Function by Chemical Modification 83
Yaowen Wu and Lei Zhao

6.1 Introduction 83

6.2 The Biological Problem 84

6.2.1 Small GTPases 84

6.2.2 Autophagy 85

6.3 The Chemical Approach 88

6.3.1 Expressed Protein Ligation and Click Ligation 88

6.3.2 Site-Specific Modification of Proteins 90

6.3.3 Semisynthesis of Lipidated LC3 Protein 94

6.4 Biological Research/Evaluation 97 6.4.1...