Advanced Chemical Biology : Chemical Dissection and Reprogramming of Biological Systems, 1e

Cover
Title Page
Copyright
Contents
Foreword
Preface
About the Companion Website
Chapter 1 Introduction to Advanced Chemical Biology
1.1 Introduction
1.2 Enabled by Synthetic and Physical Organic Chemistry
1.3 Guided by Biochemistry and Structural Biology
1.4 Enhanced by Engineering and Evolution
1.5 Expanded by Analytical Chemistry and “Omics” Technologies
1.6 Impact on Biological Discovery and Drug Development
1.7 Outlook
References
Chapter 2 DNA Function, Synthesis, and Engineering
2.1 Introduction: A Historical Perspective
2.1.1 The Structure of DNA
2.2 New Nucleobases and Unusual DNA Conformations
2.2.1 G‐Quadruplex DNA Structures
2.2.2 Circular DNA Structures
2.2.3 Aptamers
2.2.4 Other Nucleobases
2.3 The Modern Synthesis of DNA
2.3.1 Solid‐Phase DNA Synthesis
2.3.2 Backbone‐Modified Oligonucleotides
2.3.2.1 Peptide Nucleic Acids (PNAs)
2.3.2.2 Morpholino Nucleic Acids
2.4 DNA Sequencing
2.4.1 Modern Methods to Sequence DNA
2.4.1.1 Sequencing by Synthesis (SBS)
2.4.1.2 Third‐Generation DNA Sequencing
2.5 DNA Engineering
2.5.1 DNA Nanotechnology
2.5.2 DNA‐Templated Nanoparticle Assembly
2.5.3 DNA Nanomachines
2.5.4 DNA Nanotechnology for Biology
2.5.5 DNA‐Based Organelle Mapping Technology
2.5.6 DNA‐Based Technologies for the Detection of Endogenous Nucleic Acids and Proteins
2.5.6.1 Fluorescence In Situ Hybridization (FISH)
2.5.6.2 DNA‐Barcoded Antibodies for Spatial Detection of Proteins
2.5.7 DNA‐Based Super Resolution Imaging
2.5.8 DNA‐Encoded Libraries (DEL)
2.5.9 Digital Data Storage Using DNA
2.6 Tools for Engineering DNA
2.7 Summary and Future Outlook
Acknowledgments
References
Chapter 3 Chemical Approaches to Genome Integrity
3.1 Introduction and Historical Perspective
3.2 Types of DNA Damage
3.2.1 Damage to Nucleobase
3.2.1.1 Oxidation
3.2.1.2 Alkylation
3.2.1.3 Depurination/Depyrimidination
3.2.1.4 Deamination
3.2.1.5 DNA Mismatches
3.2.1.6 DNA Crosslinks
3.2.2 Damage to Sugar
3.2.3 Damage to Phosphate Backbone
3.3 Types of DNA Repair
3.3.1 Direct Repair
3.3.2 Base Excision Repair
3.3.3 Nucleotide Excision Repair
3.3.4 Mismatch Repair
3.3.5 Double‐Strand Break and Interstrand Crosslink Repair
3.4 Identification of Sites of DNA Damage and Modification
3.4.1 Traditional Methods for Damage Detection
3.4.2 Searching for Hotspots of Oxidative Damage – An OG Story
3.4.3 Sequencing for Bulky Adducts – Cisplatin and Pyrimidine Dimers
3.4.4 Sequencing for AP Site and Strand Breaks
3.5 Assays that Allow for Monitoring of the Repair of DNA Damage in Cellular Contexts
3.5.1 Lesion Reporter Assays to Monitor Base Excision Repair
3.5.2 Leveraging Cell‐Based Reporter Assays to Assess Impact of DNA Lesions on Replication and Transcription
3.5.3 Plasmid Reporters Monitoring Several DNA Repair Pathways Simultaneously
3.5.4 Highly Sensitive Fluorescent DNA Repair Probes for Clinical Diagnostics and Imaging in Cells
3.6 Summary and Future Outlook
Acknowledgments
Exam Questions
References
Chapter 4 RNA Function, Synthesis, and Probing
4.1 Introduction
4.2 The Principles of RNA Chemistry
4.2.1 The Impact of a 2′‐Hydroxyl on Nucleic Acid Chemistry
4.2.2 RNA Bases and Base‐Pairing
4.2.3 RNA Secondary Structure
4.2.4 RNA Tertiary Structures and the Ribosome
4.3 Synthesis of RNA
4.3.1 Chemical Synthesis
4.3.2 In Vitro Transcription
4.4 Labeling of RNA
4.4.1 Introducing Modifications Through Chemical Synthesis of RNA
4.4.2 Using Ligation to Introduce Chemical Modifications into RNA
4.4.3 Incorporation of Modified Bases into RNA Using IVT
4.4.4 Approaches to 3′‐End Label RNA
4.4.5 Approaches to 5′‐End Label RNA
4.5 Identification and Engineering of Functional RNAs
4.5.1 Aptamers
4.5.2 Riboswitches
4.5.3 Ribozymes
4.5.4 Genetically Encoded Tags to Label RNA
4.5.5 RNA‐Based Therapeutics
4.6 The Sequencing of RNA
4.6.1 Reverse Transcription of RNA
4.6.2 Long‐Read and Direct RNA Sequencing
4.6.3 Extensions and Alternative Approaches to RNA‐seq
4.7 The Chemical Probing of RNA Structure
4.7.1 In‐Line Probing of RNA Conformation
4.7.2 Reagents for Chemical Probing of RNA Conformation and Base‐Pairing
4.7.3 Reagents for Probing Solvent Accessibility, Tertiary Structure, and Higher Order Interactions
4.8 Summary and Future Outlook
References
Chapter 5 Chemical Approaches to Transcription and RNA Regulation In Vivo
5.1 Introduction/Historical Perspective
5.2 Core Concepts/Landmark Studies
5.2.1 Transcription Regulation in Eukaryotes
5.3 Transcription Regulation by Chemical Targeting of DNA and the Core Transcription Machinery
5.3.1 Cell‐Permeable DNA‐Targeting Small Molecules
5.3.2 Targeting Transcription by Nucleic Acids and Their Analogs
5.3.3 Small‐Molecule Inhibitors of the Transcription Machinery
5.4 Chemical Regulation of Transcription via Targeting of Epigenetic Elements
5.4.1 Transcription Regulation Through Targeting of Histone Modifications
5.4.2 DNA Methylation and Small Molecules Targeting DNA Modifications
5.5 Chemical Approaches to Target Post‐Transcriptional RNA Metabolism
5.5.1 Post‐Transcriptional RNA Metabolism
5.5.2 Regulating RNA Function by Direct RNA Binders
5.5.3 Regulating RNA Function by Targeting RNA‐Binding (Effector) Proteins
5.6 Summary and Future Outlook
References
Chapter 6 Chemical Biology of Genome Engineering
6.1 Introduction to Genome Editing
6.2 Early Genetic Engineering Experiments: Chemical Mutagenesis, Gene Transfer, and Gene Targeting
6.2.1 Chemical Mutagenesis Methods
6.2.2 Gene Transfer
6.2.3 Gene Targeting
6.3 Improving Precision and Programmability with Double‐Stranded DNA Breaks
6.3.1 The Development of Double‐Stranded Break‐Reliant Genome Editing Technologies
6.3.2 Repair of Double‐Stranded DNA Breaks in Mammalian Cells
6.3.3 Meganucleases
6.3.4 Zinc Finger Nucleases (ZFNs)
6.3.5 Transcription Activator–Like Effector Nucleases (TALENs)
6.4 The Golden Age of Genome Engineering: CRISPR‐Based Genome Editing Technologies
6.4.1 Introduction
6.4.2 CRISPR‐Cas9
6.4.3 Programmability Improvements
6.4.4 Efficiency Improvements
6.4.5 Specificity Improvements
6.4.6 Precision Improvements
6.4.7 Epigenome Editing
6.5 Non‐DSB‐Reliant Genome Editing Technologies
6.5.1 Base Editing
6.5.2 Prime Editing
6.6 Gene Editing Methods for Spatial and Temporal Control
6.7 Ethical Implications, Summary, and Future Outlook
References
Chapter 7 Peptide Synthesis and Engineering
7.1 Introduction
7.2 Peptide Synthesis
7.2.1 SPPS Is Optimized for Stepwise Efficiency
7.2.2 Nα‐protecting Groups Ensure Single Coupling of the Incoming Amino Acid
7.2.3 Plastic Resins Are Used During SPPS
7.2.4 Temporary Masking of Reactive Side Chains Is Necessary During SPPS
7.2.5 Peptide Bonds Are Synthesized by a Condensation Reaction Mediated by a Stoichiometric Coupling Agent
7.3 Secondary and Tertiary Structures of Amino Acids
7.3.1 Peptide Backbone Conformations
7.3.2 Biophysical Determinants of Helix Folding and Design of α‐Helix Mimics
7.3.3 β‐Strand and β‐Sheet Mimics
7.3.4 Protein Tertiary Structure Mimics
7.3.4.1 β‐Sheet and β‐Hairpin Mimics
7.3.5 Helical Tertiary Structure Mimics
7.4 Conformationally Defined Peptides as Modulators of Protein Interactions
7.4.1 Peptide Therapeutics
7.5 Summary and Future Outlook
References
Chapter 8 Protein Synthesis and Engineering
8.1 Introduction/Historical Perspective
8.2 Core Concepts/Landmark Studies
8.2.1 Cysteine‐thioester‐Based Ligations: Making the Pieces
8.2.1.1 C‐terminal Pieces: Chemical Synthesis of N‐terminal Cysteine Peptides
8.2.1.2 C‐terminal Pieces: Recombinant Expression of N‐terminal Cysteine Peptides/Proteins
8.2.1.3 N‐terminal Pieces: Chemical Synthesis of Thioester‐Containing Peptides
8.2.1.4 N‐terminal Pieces: Recombinant Expression of Thioester‐Containing Proteins
8.2.1.5 Internal Fragments: Preparation of Cysteine and Thioester‐Containing Peptides/Proteins
8.2.2 Adding More Pieces: Moving Beyond Thioester/Cysteine Ligations
8.2.2.1 Desulfurization
8.2.2.2 Auxiliaries
8.2.2.3 Other Ligation Chemistries
8.3 Putting the Pieces Together: Practical Considerations for Ligation Reactions
8.4 Protein Trans‐splicing
8.5 Examples of Protein Synthesis
8.5.1 Post‐Translational Modifications
8.5.1.1 Cell Signaling
8.5.1.2 Chromatin
8.5.1.3 Amyloid‐Forming Proteins
8.5.2 Chemical and Biophysical Probes
8.5.2.1 Backbone Modifications
8.5.2.2 Segmental Isotopic Labeling
8.5.3 Mirror Image Proteins
8.5.3.1 Racemic Crystallography
8.5.3.2 Mirror Image Display
8.5.4 Protein Ligation in Living Systems
8.5.5 Potential Therapeutic Applications
8.6 Summary and Future Outlook
References
Chapter 9 Directed Evolution for Chemical Biology
9.1 Introduction
9.2 Methodologies
9.2.1 Directed Evolution at the Protein Level
9.2.1.1 Random Mutagenesis
9.2.1.2 Gene Recombination
9.2.1.3 Semi‐Rational Design
9.2.2 Directed Evolution at the Pathway Level
9.2.2.1 Directed Evolution of a Single Enzyme in a Pathway
9.2.2.2 Directed Evolution of an Entire Pathway
9.2.3 Directed Evolution at the Genome Level
9.2.3.1 Adaptive Laboratory Evolution
9.2.3.2 Genome‐Scale Engineering Strategies
9.2.4 Continuous Directed Evolution
9.2.5 Screening or Selection Methods
9.2.5.1 Selection‐Based Techniques
9.2.5.2 Screening‐Based Techniques
9.3 Case Studies
9.3.1 Directed Evolution of a Glyphosate N‐Acetyltransferase
9.3.2 Directed Evolution of a Transaminase for Sitagliptin Manufacture
9.3.3 Directed Evolution of a Cytokine Using DNA Family Shuffling
9.3.4 Efficient Proximity Labeling in Living Cells and Organisms with TurboID
9.3.5 Biocatalytic Cascade Evolution for Manufacturing Islatravir
9.3.6 A Multi‐Functional Genome‐Wide CRISPR System
9.4 Future Perspectives and Conclusion
Acknowledgments
References
Chapter 10 Chemical Biology of Cellular Metabolism
10.1 Introduction/Historical Perspective
10.2 Metabolite Detection and Quantitation
10.2.1 Shotgun Metabolomics
10.2.2 Targeted Metabolomics
10.2.3 Metabolite Flux Analysis
10.2.4 Untargeted Metabolomics
10.2.5 Discovering Structurally Novel Metabolites
10.3 Metabolite Imaging and Sensing
10.3.1 Mass Spectrometry Imaging
10.3.2 Chemical Probes for Metabolite Imaging
10.3.3 Protein and RNA Metabolite Sensors
10.4 Perturbation of Metabolite Levels
10.4.1 Small‐Molecule Inhibitors and Drugs of Metabolism
10.4.2 Enzymatic Perturbation of Metabolism
10.5 The Impact of Chemical Biology in Disease and Drug Discovery
10.6 Summary and Future Outlook
References
Chapter 11 Chemical Biology of Lipids
11.1 Introduction
11.2 Identification of Bulk Lipids
11.2.1 Lipidomics by Mass Spectrometry
11.2.2 Lipid Analysis by Thin‐Layer Chromatography
11.3 Fixing Lipids in Subcellular Space
11.3.1 Protein‐Based Techniques to Localize Lipids
11.3.2 Mass Spectrometry Imaging of Lipids
11.3.3 Lipid Detection Using Modified Lipids as Probes
11.4 Tracing Individual Lipids via In Cellulo Click Chemistry
11.4.1 Alkyne/Azide‐Modified Lipids and Click Chemistry
11.4.2 Bifunctional Lipid Derivatives
11.5 Tools to Elucidate Lipid Signaling
11.5.1 Metabolic Machinery as a Chemical Tool: the Advantage of Chemical Dimerizers
11.5.2 Releasing Bioactive Lipids with Light
11.6 A Comprehensive View of Protein–Lipid Interactions
11.6.1 Trifunctional Lipids
11.6.2 Lipid–Protein Interactome
11.7 Summary and Future Outlook
References
Chapter 12 Protein Posttranslational Modifications
12.1 Introduction
12.2 Functional Impacts of PTMs
12.3 Evolution and PTMs
12.4 Major Classes of PTMs
12.4.1 Phosphorylation
12.4.2 Acetylation
12.4.3 Ubiquitination
12.4.4 Methylation
12.4.5 Glycosylation
12.4.6 Lipidation of Proteins
12.4.7 Oxidation of Proteins
12.4.8 Miscellaneous Modifications
12.5 Writers and Erasers
12.5.1 Protein Kinases and Phosphatases
12.5.2 Acetyltransferases and Deacetylases
12.5.3 Ubiquitin Ligases and Deubiquitinases
12.5.4 Methylation and Demethylases
12.5.5 Glycosyltransferases and Glycosidases
12.5.6 Lipid Transferase and Hydrolases
12.6 Strategies for the Study of PTMs
12.6.1 Mutagenesis
12.6.2 Genetic Codon Expansion
12.6.3 Small‐Molecule Probes and Chemical Complementarity
12.6.4 Chemical Ligation
12.6.5 Protein Microarrays
12.7 Protein PTMs in Diseases
12.7.1 Protein Kinases and Diseases
12.7.2 Lys Acetylation and Cutaneous T Cell Lymphoma
12.7.3 Ubiquitination
12.8 Summary
References
Chapter 13 Chemical Glycobiology
13.1 Introduction
13.2 Total Chemical Synthesis of Structurally Defined Glycans
13.3 Enzymatic and Chemoenzymatic Synthesis of Glycans
13.4 Programmable and Automated Glycan Synthesis
13.5 Synthesis of Glycopeptides and Glycoproteins
13.6 Glycan Microarrays
13.7 Chemical Tagging and Remodeling of Cellular Glycans
13.8 Inhibitors of Glycan‐Processing Enzymes and Glycan Binding Proteins
13.9 Glycan‐Targeted Therapeutics
13.10 Summary and Future Outlook
References
Chapter 14 The Chemical and Enzymatic Modification of Proteins
14.1 Introduction
14.2 General Considerations
14.3 Lysine Modification
14.4 Aspartic Acid, Glutamic Acid, and C‐Terminal Carboxylate Modification
14.5 Tyrosine Modification
14.6 Cysteine Modification
14.7 Methionine Modification
14.8 Tryptophan Modification
14.9 Histidine Modification
14.10 Serine and Threonine Modification
14.11 N‐Terminal Modification
14.12 Enzymatic Approaches to Modifying Proteins
14.12.1 Transpeptidases
14.12.2 Ligases
14.12.3 Activating Enzymes
14.13 Summary and Future Outlook
References
Chapter 15 Genetic Code Expansion
15.1 Introduction
15.2 Genetic Code Expansion Through Directed Evolution of aaRS/tRNA Pairs
15.2.1 The Development of the Mj.TyrRS‐tRNA Based GCE System
15.2.2 The Development of Additional aaRS/tRNA‐Based GCE System
15.2.3 The PylRS‐tRNA Pair as a “one‐stop‐shop” GCE System
15.2.4 Genetic Code Expansion in Multicellular Organisms
15.3 GCE with Genome Recoding Strains and/or Unnatural Codons
15.3.1 GCE with Genome Recoding Strains
15.3.2 GCE with Four‐Base Codons Using Orthogonal Ribosome
15.3.3 Genetic Code Expansion with Unnatural Base Pairs
15.4 GCE‐based Applications
15.4.1 Site‐Specific Posttranslational Modifications (PTMs)
15.4.2 New “Physical” Property Empowered by ncAAs
15.4.3 New Chemical Reactivity Derived from ncAA and Their Unique Applications
15.4.4 Control of Protein Activation
15.5 Therapeutic Conjugates
15.6 Live‐Attenuated Virus and Other Genetically Modified Vaccines
15.7 Summary and Future Outlook
15.7.1 Improving the Efficiency
15.7.2 Expanding the Applications
15.7.3 Exploring the Therapeutic Potential
References
Chapter 16 Bioorthogonal Chemistry
16.1 Introduction and Historical Perspective
16.2 Key Concepts: Bioorthogonality and Bioorthogonal Reactions, Click Chemistry, and the Bioorthogonal Metabolic Reporter Strategy
16.2.1 Bioorthogonality and Bioorthogonal Reactions
16.2.2 Click Chemistry
16.2.3 The Bioorthogonal Metabolic Reporter Strategy
16.3 The Beginnings of Bioorthogonal Chemistry: Oxime and Hydrazone Formation
16.4 The Azide as a Bioorthogonal Handle
16.5 The Staudinger Ligation of Azides and Phosphines
16.6 Cu‐Catalyzed Azide–Alkyne Cycloaddition (CuAAC) of Azides and Terminal Alkynes
16.7 Strain‐Promoted [3+2] Azide–Alkyne Cycloaddition (SPAAC) of Azides and Cyclooctynes
16.8 The Tetrazine Ligation: Rapid Bioorthogonal Inverse Electron‐Demand Diels–Alder Reactions
16.9 Other Bioorthogonal Ligations
16.10 Light‐Activated Bioorthogonal Reactions
16.11 Bioorthogonal Uncaging and Cleavage Reactions
16.12 Mutually Orthogonal Bioorthogonal Reactions
16.13 Fluorogenic Bioorthogonal Reagents
16.14 Applications of Bioorthogonal Chemistry
16.14.1 Bioorthogonal Non‐canonical Amino Acid Tagging (BONCAT)
16.14.2 In Vivo Imaging of Glycans
16.14.3 Therapeutic Applications of Bioorthogonal Chemistry
16.15 Summary and Future Outlook
References
Chapter 17 Cellular Imaging
17.1 Introduction
17.1.1 History
17.1.2 Light and Fluorescence
17.2 Small‐Molecule Fluorophores
17.2.1 Background
17.2.2 Pyrenes and Coumarin Fluorophores
17.2.3 BODIPY Dyes
17.2.4 Fluoresceins and Rhodamines
17.2.5 Phenoxazine and Cyanine Dyes
17.2.6 Use as Biomolecule Labels
17.2.7 Use as Cellular Stains
17.2.8 Fluorescent Indicators
17.2.9 Enzyme Substrates
17.3 Fluorescent Proteins
17.3.1 Background
17.3.2 General Considerations of Fluorescent Proteins
17.3.3 Fluorescent Proteins as Biomolecule Labels and “Stains”
17.3.4 Fluorescent Proteins as Sensors
17.3.5 Fluorescent Proteins as Enzyme Substrates
17.4 Hybrid Small‐Molecule–Protein Systems
17.4.1 Background
17.4.2 Labels
17.4.3 Sensors
17.5 Landmark Study I: Harnessing Photosensitive Fluorophores
17.5.1 Background
17.5.2 Super‐Resolution Microscopy
17.6 Landmark Study II: Ca2+ Imaging
17.6.1 Background
17.6.2 Small‐Molecule Ca2+ Indicators
17.6.3 Genetically Encoded Ca2+ Indicators
17.6.4 Genetically Encoded Indicators for In Vivo Imaging
17.7 Summary and Future Outlook
References
Chapter 18 In Vivo Imaging
18.1 Introduction
18.2 Basic Concepts for Imaging In Vivo
18.3 The Imaging Toolbox: Probes for Imaging Cellular and Molecular Features
18.3.1 “Always On” Probes
18.3.2 “Turn‐On” (Activatable) Probes
18.3.3 Genetically Encoded Probes
18.4 Molecular Imaging Across the Electromagnetic Spectrum
18.4.1 Imaging with X‐rays (CT)
18.4.2 Imaging with Sound (US)
18.4.3 Imaging with Radio Waves (MRI)
18.4.4 Imaging with Radionuclides (PET/SPECT)
18.4.5 Imaging with Optical Light (Fluorescence/Bioluminescence)
18.4.5.1 Targeted Fluorophores and Fluorescent Materials
18.4.5.2 Activatable Probes
18.4.5.3 Genetically Encoded Fluorescent Probes
18.4.5.4 Genetically Encoded Bioluminescent Proteins (Luciferases)
18.4.5.5 Engineered Probes for Sensing Metabolites and Molecular Features
18.5 Multimodality Imaging and Combination Probes
18.6 Emerging Areas in Molecular Imaging
18.7 Summary and Future Outlook
References
Chapter 19 Chemical Biology of Metals
19.1 Introduction
19.2 Metals and the Inorganic Foundations of Life
19.2.1 Metal Complexes are Lewis Acid–Base Complexes
19.2.2 Crystal Field Theory Enables Bonding Analysis from Molecular Shape and d Orbitals
19.2.3 Hard Soft Acid Base Theory Defines Metal–Ligand Preferences
19.3 Non‐Redox Roles for Metals in Biology: Structure and Lewis Acid Catalysis
19.3.1 Metals for Stabilizing Nucleic Acid Structure
19.3.2 Metals as Protein Structural Units: Zinc Finger and EF Hand Motifs
19.3.3 Metals as Lewis Acid Catalysts: Metallohydrolases
19.4 Redox Chemistry: Oxygen Transport and Electron Transfer Proteins
19.4.1 Oxygen Transport Requires Redox‐Active Metal Binding
19.4.2 Marcus Theory and Electron Transfer Proteins
19.5 Redox Chemistry: Metalloenzymes for Redox Catalysis at Oxygen, Nitrogen, and Carbon
19.5.1 Oxygen Evolution in Photosynthesis: Photosystem II
19.5.2 Oxygen Reduction: Respiration with Cytochrome c Oxidase
19.5.3 Oxygen Catalysis: Heme and Non‐Heme Iron‐Dependent Oxidations
19.5.4 Nitrogen Cycle: Nitrogenases and Nitrate/Nitrite Reductases
19.5.5 Bioorganometallic Chemistry: Carbon Cycling and Vitamin B12
19.6 Metals in Medicine: Metallotargets, Metallodrugs, and Metal‐Based Imaging Agents
19.7 Emerging Areas for Metals in Biology: Transition Metal Signaling and Metalloallostery
19.8 Chemical Tools to Study Metal Biology
19.9 Summary and Future Outlook
References
Chapter 20 Redox Chemical Biology
20.1 Introduction
20.2 Activity‐Based Detection of Cysteine Modifications
20.3 Indirect Profiling of Cysteine Oxidation
20.4 Direct Profiling of Cysteine OxiPTMs with Chemoselective Probes
20.4.1 Profiling Protein Sulfenic Acids (−SOH)
20.4.1.1 Sulfenic Acid Probes – A Historical Perspective
20.4.1.2 Chemical Models for the Assessment of Sulfenic Acid Probes
20.4.1.3 Selectivity of Chemical Probes for Sulfenic Acids
20.4.1.4 Quantification of Protein Sulfenic Acids
20.4.1.5 Application of Sulfenic Acid Probes
20.4.2 Profiling Protein Sulfinic Acids (−SO2H)
20.4.3 Profiling Protein Persulfides (−SSH)
20.5 Probes and Biosensors for Reactive Oxygen Species in Cells
20.6 Conclusions and Outlook
References
Chapter 21 Activity‐Based Protein Profiling
21.1 Introduction/Historical Perspective
21.2 Core Concepts/Landmark Studies
21.2.1 Probe Design
21.2.1.1 Reactive Groups
21.2.1.2 Reporter Tags
21.2.1.3 Recognition Group/Linker
21.2.2 Detection Methods
21.2.2.1 Gel‐Based Analysis
21.2.2.2 Fluorescence Polarization
21.2.2.3 Imaging of Proteins in Cells and Organisms
21.2.2.4 Quantitative Proteomics by Mass Spectrometry
21.2.3 Common Applications
21.2.3.1 Profiling Protein Activity and Amino Acid Reactivity in Biological Systems of Interest
21.2.3.2 Competitive ABPP for Ligand Discovery and Optimization
21.2.3.3 Target Identification for Ligands
21.2.3.4 Assignment of Enzyme Function
21.2.3.5 Visualizing Enzyme Localization and Activity in Living Cells and Organisms
21.3 Summary and Future Outlook
References
Chapter 22 Chemical Genetics
22.1 Introduction
22.2 AS‐Protein – Orthogonal Molecular Glues
22.3 AS‐Enzyme – Orthogonal Substrate Pairs
22.3.1 Protein Kinases
22.3.2 Protein Methyltransferases
22.3.3 Protein Lysine Acetyltransferases
22.3.4 PARPs
22.3.5 Glycosyltransferases
22.3.6 PTM Erasers: Lysine Demethylases
22.3.7 Beyond PTM Enzymes
22.4 AS‐Enzyme – Orthogonal Inhibitor Pairs
22.4.1 Protein Kinases
22.4.2 Other Enzymes
22.5 Final Thoughts
22.5.1 Beyond Bump–Hole
References
Chapter 23 Natural Product Discovery
23.1 Introduction and Definitions
23.2 Key Concept: Natural Products are Genetically Encoded
23.3 Key Concept: Structural Differences Between Natural Products and Synthetic Drugs
23.4 Key Concept: Target Specificity and Latent Reactivity
23.5 Key Concept: Natural Product Discovery and Activity‐Guided Fractionation
23.6 Key Concept: Cryptic Biosynthetic Gene Clusters
23.7 Landmark Studies: Penicillin and the Golden Age of Antibiotic Discovery
23.8 Landmark Studies: Activating Silent Biosynthetic Gene Clusters
23.8.1 Manipulation of Culture Conditions
23.8.2 Classical Genetics
23.8.3 Chemical Genetics
23.8.4 Heterologous Expression
23.9 Summary and Outlook
References
Chapter 24 Natural Product Biosynthesis
24.1 Introduction
24.2 Peptide Natural Products
24.2.1 Ribosomally Synthesized and Post‐translationally Modified Peptides (RiPPs)
24.2.2 Non‐ribosomal Peptides
24.3 Polyketide Natural Products
24.3.1 Bacterial Type‐I Polyketides
24.3.2 Bacterial Type‐II Polyketides
24.4 Terpene Natural Products
24.5 Hybrid and Unnatural Natural Products
24.6 Summary and Future Outlook
Acknowledgment
References
Chapter 25 Chemical Microbiology
25.1 Introduction and History
25.2 Cell Envelope Structure and Biosynthesis
25.2.1 Bacterial Cell Structure
25.3 Chemical and Chemoenzymatic Synthesis for Pathway Elucidation
25.3.1 Peptidoglycan (PG) Biosynthesis
25.3.1.1 Reconstructing the Steps in PG Biosynthesis Using Defined Substrates
25.3.1.2 Accessing Lipids I and II
25.3.2 Cell Envelope Components Beyond Peptidoglycan
25.3.2.1 Gram‐Negative Lipopolysaccharides
25.3.2.2 Wall Teichoic Acid Biosynthesis
25.3.2.3 Mycobacterial Galactan
25.4 The Chemical Biology of Antibiotic Action
25.4.1 PG Assembly Is Targeted by Diverse Antibiotics
25.4.2 Penicillin and Other Antibiotics Induce Dominant‐Negative Effects
25.4.3 Identifying Inhibitors of Essential Enzymes Is Not Enough
25.4.4 Identifying Attributes for Compound Uptake in Bacteria
25.5 Chemical Biology Strategies for Imaging PG Assembly and Remodeling
25.5.1 Antibiotic‐Based PG Probes
25.5.1.1 Antibiotics that Bind PG Intermediates
25.5.1.2 Probes from Antibiotics that Act on Enzymes that Generate PG
25.5.2 Substrate Analogue PG Probes
25.6 Labeling Glycan Cell Envelope Components
25.6.1 Diversity and Function of Bacterial Polysaccharides
25.6.2 Probes of Bacterial Glycans
25.6.3 Probes of LPS: AzKdo
25.6.4 Labeling Mycobacterial Glycans
25.6.5 Trehalose Analogs
25.6.6 Imaging Probes
25.6.7 Fluorogenic Probes
25.7 Chemical Probes Applied to the Microbiome
25.7.1 Microbiome: Looking Forward
25.8 Summary and Future Outlook
References
Chapter 26 Chemical Approaches to Analyze Biological Mechanisms and Overcome Resistance to Therapeutics
26.1 Introduction
26.2 Using Chemical Inhibitors as Tools to Probe Cellular Processes
26.3 Using Resistance to Characterize Chemical Inhibitors
26.4 Crash‐Testing Drugs
26.5 RADD – Resistance Analysis During Design
26.6 Designing Inhibitors with Distinct Binding Modes
26.7 Addressing Drug Resistance with Targeted Protein Degradation
26.8 Overcoming Resistance by Using Combinations of Drugs
26.9 Conclusions
References
Chapter 27 Chemical Developmental Biology
27.1 Introduction
27.2 Small‐Molecule Teratogens
27.2.1 Cyclopamine
27.2.2 Thalidomide
27.3 Optochemical and Optogenetic Probes
27.3.1 Optochemical Control of Gene Expression
27.3.2 Optogenetic Control of Cell Signaling
27.4 Lineage Tracing Tools
27.4.1 Chemical Control of Genetic Recombination
27.4.2 DNA Barcoding Strategies
27.5 Summary
References
Chapter 28 Chemical Immunology
28.1 Introduction
28.2 Chemical Dissection of Adaptive Immunity
28.3 Generation and Chemical Engineering of Antibodies
28.4 Antigen Recognition by Immune Cells
28.5 Chemical Innovations for Eliciting and Discovering Antigen‐specific Immune Responses
28.6 Chemical Modulation of Innate Immunity
28.7 Chemical Dissection of Immunity
28.8 Summary and Future Outlook
References
Chapter 29 Chemical Neurobiology
29.1 Introduction
29.2 Actuation
29.2.1 Neuropharmacology Has a Storied History
29.2.2 Molecular Cloning and Structural Biology Have Revolutionized the Field
29.2.3 Caged Ligands and Photopharmacology Allow for Optical Control of Neural Activity
29.2.4 Chemogenetics Enables Cell‐Specific Neuropharmacology in Brains
29.2.5 Tethered Pharmacology Operates on Engineered Receptors or Native Receptors in Genetically Modified Cells
29.2.6 Tethered Photopharmacology Combines Genetic with Optical Control
29.2.7 Synthetic Photoreceptors Can Be Engineered Through Genetic Code Expansion
29.3 Visualization
29.3.1 Chemical Staining and Imaging Methods Have Launched Modern Neuroscience
29.3.2 Calcium Imaging Can Be Used to Monitor Neuronal Activity
29.3.3 Voltage Sensing Provides a Direct Picture of Neuronal Activity
29.3.4 Neurotransmitters Can Be Sensed with Chemogenetic FRET Sensors
29.3.5 Metals and Gases in the Brain Can Be Sensed with Fluorescent Probes
29.3.6 Positron Emission Tomography Requires Fast Chemistry
29.3.7 Proximity Ligation Enables Spatially Resolved Mapping of Neural Networks
29.4 Summary and Outlook
References
Chapter 30 Small‐Molecule Drug Discovery
30.1 Introduction
30.2 Discovery of Chemical Matter
30.2.1 Target‐Based Discovery
30.2.2 HTS‐Compatible Assay Formats
30.2.3 Phenotype‐Based Discovery
30.2.4 HTS: General Considerations
30.2.5 Drug Repurposing and Serendipity
30.2.6 Alternative Small‐Molecule Discovery Approaches
30.3 In Vivo Pharmacology: Invention of Drug Candidates and In Vivo Probes
30.3.1 Drug Absorption, Distribution, Metabolism, and Excretion
30.3.1.1 Drug Absorption and Distribution
30.3.1.2 Physicochemical Properties of Drugs
30.3.1.3 Drug Metabolism and Excretion
30.3.1.4 Pharmacokinetics, Pharmacodynamics, and Biomarkers
30.3.2 Medicinal Chemistry
30.3.3 Drug Toxicity and Human Clinical Trials
30.4 Conclusion
References
Index
EULA