Cover
Protein-Protein Interaction Regulators
Preface
Contents
Chapter 1 - Protein-Protein Interaction Interfaces and their Functional Implications
1.1 Introduction
1.2 Categorizing Protein-Protein Interactions
1.3 Biophysical Principles Governing the Specificity of Protein-Protein Interactions
1.4 Amino Acid Contributions Towards Association and Dissociation Rate Constants
1.5 Structural Changes Associated with Formation of Protein-Protein Complexes
1.6 General Characteristics of PPI Interfaces
1.7 The Role of Interface Water in Binding
1.8 The Core-Rim Model of Protein- binding Interfaces
1.9 Binding Interfaces Viewed as Connected Modules
1.10 Enthalpy-Entropy Compensation in PPIs
1.11 Protein- binding Sites are Imprinted in the Unbound Structure
1.12 Conservation of Interface Residues
1.13 Concluding Remarks
Acknowledgements
References
Chapter 2 - Protein-Protein Interaction Networks in Human Disease
2.1 Introduction
2.2 Methods Commonly Employed in Disease PPI Network Analysis
2.2.1 Yeast- two- hybrid
2.2.2 Protein Microarray
2.2.3 Affinity- purification Coupled to Mass Spectrometry
2.2.4 Proximity Labelling Methods
2.3 Role of PPI Networks in Human Diseases
2.3.1 Cancer
2.3.2 Parkinson's Disease
2.3.3 Cardiomyopathies
2.3.4 Diabetes
2.3.5 Autoimmune Diseases
2.3.6 Microbial Diseases
2.4 Challenges in Working with PPI Networks and Overcoming these Challenges
2.5 Conclusion
Funding
References
Chapter 3 - High Throughput Screening Methods for PPI Inhibitor Discovery
3.1 Introduction
3.2 HTS Methods for PPI Inhibitor Discovery
3.2.1 Overview
3.2.2 Methods to Detect Direct Interactions
3.2.2.1 Fluorescence Polarization.

3.2.2.1.1 Basic Principle.Fluorescence polarization (FP) methodology uses change in molecular rotation as a readout for intermolecular bin...
3.2.2.1.2 General Design and Applications.In the context of high throughput PPI detection, FP assays are typically developed to monitor bi...
3.2.2.2 Methods for Detection of Small Molecule Binders to Proteins
3.2.3 Proximity- based Energy Transfer Methods
3.2.3.1 AlphaScreen
3.2.3.1.1 Basic Principle.Amplified Luminescent Proximity Homogeneous Assay Screen (AlphaScreen) is an energy transfer- based technology f...
3.2.3.1.2 General Design and Applications.Various versions of Alpha Screen have been developed that substitute anthracene and rubrene in a...
3.2.3.2 Resonance Energy Transfer- based Methods
3.2.3.2.1 Basic Principle.Resonance energy transfer (RET) is an optical phenomenon in which the excess energy of an excited molecule, desi...
3.2.3.2.2 General Design and Applications
Fluorescence/Förster Resonance Energy Transfer (FRET). In FRET assays, both the donor (A) and acceptor (B) molecules are fluorop...
Bioluminescence Resonance Energy Transfer (BRET). Bioluminescence resonance energy transfer (BRET) represents another proximity-...
3.2.4 Protein Fragment Complementation Assay (PCA) Methods
3.2.4.1 Bimolecular Fluorescence Complementation (BiFC) and Bimolecular Luminescence Complementation (BiLC)
3.2.4.1.1 Basic Principle.In bimolecular fluorescence complementation (BiFC), a GFP variant is split between residue 157 and 158 into two ...
3.2.4.1.2 General Design and Applications.The BiFC signal can be monitored either through direct measurement of fluorescence intensity or ...
3.2.4.2 Two- hybrid Methods
3.2.4.2.1 Basic Principle.Two- hybrid assays are cell- based assays in which split fragments of a transcription factor or signaling protei.

3.2.4.2.2 General Design and Applications.Many HTS two- hybrid systems used in PPI inhibitor screening currently are modifications of the ...
3.2.5 Phenotype- based Methods: High Content Screening
3.2.5.1 Basic Principle
3.2.5.2 General Design and Applications
3.3 Validation Methods for PPI Inhibitor Discovery
3.3.1 Biosensor Technologies: Surface Plasmon Resonance and Biolayer Interferometry
3.3.2 Affinity Capture Techniques: Co- immunoprecipitation and Pull- down
3.3.3 Ligand- observed NMR and Protein- observed NMR
3.3.4 X- ray Crystallography
3.3.5 Isothermal Titration Calorimetry
3.3.6 Differential Scanning Fluorometry (Protein Thermal Shift Assay)
3.3.7 Cellular Thermal Shift Assay
3.3.8 Microscale Thermophoresis
3.3.9 Proximity Ligation Assay
3.4 Optimization of Identified PPI Inhibitors
3.5 Conclusion
Acknowledgements
References
Chapter 4 - Computational Structural Modeling to Discover PPI Modulators
4.1 Introduction
4.2 Protein Structure Availability and Modeling
4.2.1 Swiss- model
4.2.2 I- TASSER
4.3 Identification of the PPI Interface and Small Molecule Binding Site
4.3.1 Protein-Protein Docking Tools and Software
4.3.1.1 HADDOCK
4.3.1.2 ATTRACT
4.3.1.3 SwarmDock
4.3.1.4 ClusPro
4.3.2 Binding Pocket Identification
4.3.2.1 CASTp
4.3.2.2 PrankWeb
4.3.2.3 ArDock
4.4 Virtual Screening for Small Molecule PPI Modulators
4.4.1 AutoDock Vina
4.4.2 USCF DOCK
4.4.3 GLIDE
4.4.4 FRED
4.4.5 DARC
4.5 Concluding Remarks
Acknowledgements
References
Chapter 5 - Small Molecule Inhibitors of E3 Ubiquitin Ligases
5.1 Introduction
5.2 Inhibitors of SKP2 to Protect p27 from Ubiquitination
5.2.1 Inhibitors of p27Kip1 Degradation Through a Cell- based Screening System
5.2.1.1 1,25- (OH)2 Vitamin D3
5.2.1.2 Silibinin.

5.2.1.3 Epigallocatechin- 3- gallate (EGCG)
5.2.1.4 CpdA
5.2.1.5 SMIP001 and SMIP004
5.2.1.6 Pentagalloylglucose (5gg), Curcumin, Quercetin and Lycopene
5.2.2 Inhibitors of p27Kip1 Degradation Through the Target- based Screening System
5.2.2.1 C1, C2, C16 and C20
5.2.2.2 Compound #25
5.2.3 Screening System for the Inhibitors of Interaction Between Skp2 and p27Kip1
5.2.3.1 NSC689857 and NSC681152
5.2.3.2 Sulfonamidoquinolines
5.2.4 Fragment- based Screening System of Inhibitors of p27Kip1 Ubiquitination
5.2.4.1 Linichlorin A and Gentian Violet
5.3 Modifiers of Other F- box Protein- dependent Interactions
5.3.1 Inhibitors of β-­TrCP-­dependent Ubiquitination
5.3.1.1 Erioflorin
5.3.1.2 GS143
5.3.1.3 Acyclovir Triphosphate
5.3.2 Modifiers of Fbw7- dependent Ubiquitination
5.3.2.1 Oridonin
5.3.2.2 Genistein
5.3.2.3 SCF- I2
5.3.3 Inhibitors of Yeast F- box Protein Met30
5.3.3.1 SMER3
5.4 Conclusions and Perspective
References
Chapter 6 - Hydrogen Bond Surrogate Stabilized Helices as Protein-Protein Interaction Inhibitors
6.1 Introduction
6.2 Attributes of α-­Helices at Protein-Protein Interfaces
6.3 HBS Synthesis
6.3.1 Olefin HBS
6.3.2 Thioether and Disulfide HBS
6.3.3 Olefin α3βHBS
6.4 Conformational Analysis of HBS Helices
6.5 HBS Helices as Ligands for Protein Receptors
6.6 Hypoxia Inducible Factor 1α (HIF-­1α)/p300
6.6.1 Ras/Sos
6.7 Conclusion and Future Directions
References
Chapter 7 - Helix- mimetics as Protein-Protein Interaction Inhibitors
7.1 Importance of Protein-Protein Interactions in Biology
7.1.1 Signaling Pathways
7.1.2 Gene Regulatory Networks
7.2 Aberrant Protein-Protein Interactions in Disease Etiology
7.3 Characteristics of Protein-Protein Interfaces
7.4 Types of Helices in Natural Proteins.

7.5 Strategies for Stabilizing the α-­Helix
7.6 General Classification of Peptidomimetics and Different Types of Helix Mimetics
7.7 Side- chain Cross- linked Helix Mimetics
7.7.1 Lactam Bridges
7.7.2 Disulfide and Related Cross- linkers
7.7.3 Triazole Cross-­linked α-­Helices
7.7.4 Foldamers
7.7.5 Hydrocarbon Stapled Helices
7.7.6 Introduction of Conformation- constraining Amino Acids
7.8 Intracellular Delivery of Helical Peptides
7.8.1 Cell- penetrating Peptides
7.8.2 Endosomal Escape
7.9 Other Intracellular Delivery Methods
7.9.1 Peptide- loaded Nanoparticles
7.9.2 Liposome- mediated Delivery
7.10 Examples of Synthetic Helical Peptides used as Protein-Protein Interaction Inhibitors
7.10.1 p53- MDM2/MDMX Inhibitor
7.10.2 RAS- SOS Inhibitor
7.10.3 cFos-cJun Interaction
7.10.4 HIV1- gp41
7.11 Helical Peptides in Clinical Trials
References
Chapter 8 - Discovery and Development of Mcl- 1 Inhibitors as Anti- cancer Therapeutics: Hit to Clinical Candidate Optimization
8.1 Introduction
8.1.1 A Protein-Protein Interaction Network Regulates Programmed Cell Death
8.2 Discovery of Selective Mcl- 1 BH3 Mimetic Inhibitors
8.1.2 Targeting Bcl- 2 Regulated Apoptosis in Cancer
8.2.1 Discovery of Mcl- 1 Inhibitors from High Throughput Screening
8.2.1.1 Walensky Group (Dana Farber Cancer Institute)
8.2.1.2 Nikolovska- Coleska Group (University of Michigan)
8.2.2 Development of Mcl- 1 Inhibitors Derived from Fragment- based Screening
8.2.2.1 2- Carboxylic Acid Indole- based Inhibitors (Abbvie)
8.2.2.2 Tricyclic Indole- based Inhibitors (Fesik Group - University of Vanderbilt)
8.2.2.3 Thienopyrimidine- based Inhibitors (Servier/Vernalis)
8.2.3 Development of Macrocyclic Mcl- 1 Inhibitors
8.2.3.1 Cyclized Indole 2- Carboxylic Acid Inhibitors (AstraZeneca).

8.2.3.2 Macrocyclic Acylsulfonamide- based Inhibitors (Amgen).