Enzymes

Chapter 1: A Brief History of Enzymology

• • Key Learning Points
• • 1.1 Enzymes in Antiquity
• • 1.2 Early Enzymology
• • 1.3 The Development of Mechanistic Enzymology
• • 1.4 Studies of Enzyme Structure
• • 1.5 Enzymology Today
• • 1.6 Summary
• • References and Further Reading

Chapter 2: Chemical Bonds and Reactions in Biochemistry

• • Key Learning Points
• • 2.1 Atomic and Molecular Orbitals
• • 2.2 Thermodynamics of Chemical Reactions
• • 2.3 Acid–base Chemistry
• • 2.4 Noncovalent Interactions in Reversible Binding
• • 2.5 Rates of Chemical Reactions
• • 2.6 Summary
• • References and Further Reading

Chapter 3: Structural Components of Enzymes

• • Key Learning Points
• • 3.1 The Amino Acids
• • 3.2 The Peptide Bond
• • 3.3 Amino Acid Sequence or Primary Structure
• • 3.4 Secondary Structure
• • 3.5 Tertiary Structure
• • 3.6 Subunits and Quaternary Structure
• • 3.7 Cofactors in Enzymes
• • 3.8 Conformational Dynamics and Enzyme Function
• • 3.9 Methods of Protein Structure Determination
• • 3.10 Summary
• • References and Further Reading

Chapter 4: Protein–Ligand Binding Equilibria

• • Key Learnings Points
• • 4.1 The Equilibrium Dissociation Constant, K d
• • 4.2 The Kinetic Approach to Equilibrium
• • 4.3 Binding Measurements at Equilibrium
• • 4.4 Graphic Analysis of Equilibrium Ligand-Binding Data
• • 4.5 Equilibrium Binding with Ligand Depletion (Tight Binding Interactions)
• • 4.6 Competition Among Ligands for a Common Binding Site
• • 4.7 Protein Dynamics in Receptor–Ligand Binding
• • 4.8 Orthosteric and Allosteric Ligand Binding Sites
• • 4.9 Experimental Methods for Measuring Ligand Binding
• • 4.10 Summary
• • References and Further Reading

Chapter 5: Steady-State Kinetics of Single-Substrate Enzyme Reactions

• • Key Learning Points
• • 5.1 The Time Course of Enzymatic Reactions
• • 5.2 Effects of Substrate Concentration on Velocity
• • 5.3 The Rapid Equilibrium Model of Enzyme Kinetics
• • 5.4 The Steady-State Model of Enzyme Kinetics
• • 5.5 The Significance of k cat and K m
• • 5.6 Experimental Measurement of k cat and K m
• • 5.7 Other Linear Transformations of Enzyme Kinetic Data
• • 5.8 Measurements at Low Substrate Concentrations
• • 5.9 Deviations From Hyperbolic Kinetics
• • 5.10 Summary
• • References and Further Reading

Chapter 6: Chemical Mechanisms in Enzyme Catalysis

• • Key Learning Points
• • 6.1 Substrate–Active Site Complementarity
• • 6.2 Rate Enhancement Through Transition State Stabilization
• • 6.3 Chemical Mechanisms for Transition State Stabilization
• • 6.4 The Serine Proteases: An Illustrative Example
• • 6.5 Enzymatic Reaction Nomenclature
• • 6.6 Summary
• • References and Further Reading

Chapter 7: Experimental Measures of Steady-State Enzyme Activity

• • Key Learning Points
• • 7.1 Initial Velocity Measurements
• • 7.2 Detection Methods
• • 7.3 Separation Methods in Enzyme Assays
• • 7.4 Factors Affecting the Velocity of Enzymatic Reactions
• • 7.5 Reporting Enzyme Activity Data
• • 7.6 Enzyme Stability
• • 7.7 Summary
• • References and Further Reading

Chapter 8: Transient-State Kinetics

• • Key Learning Points
• • 8.1 Timescale of Pre-Steady-State Turnover
• • 8.2 Instrumentation for Transient Kinetic Measurements
• • 8.3 Estimating Initial Conditions for Transient Kinetic Measurements
• • 8.4 Examples of Some Common Transient Kinetic Reaction Mechanisms
• • 8.5 Examples of Transient Kinetic Studies from the Literature
• • Deformylase
• • 8.6 Summary
• • References and Further Reading

Chapter 9: Enzyme Regulation

• • Key Learning Points
• • 9.1 Active and Inactive Conformational States
• • 9.2 Post-Translational Modifications
• • 9.3 Enzyme Regulation Through Protein–Protein Interactions
• • 9.4 Small-Molecule Allosteric Ligands
• • 9.5 Quantitative Measurements of Enzyme Activation and Inhibition
• • 9.6 Regulation of Protein Kinases
• • 9.7 Summary
• • References and Further Reading

Chapter 10: Reversible Inhibitors

• • Key Learning Points
• • 10.1 Equilibrium Treatment of Reversible Inhibition
• • 10.2 Thermodynamic Modes of Reversible Inhibition
• • 10.3 Effects of Inhibitors on Steady-State Parameters
• • 10.4 Concentration-Response Plots of Enzyme Inhibition
• • 10.5 Effects of Substrate Concentration on Inhibitor Concentration–Response Curves
• • 10.6 Mutually Exclusive Binding of Two Inhibitors
• • 10.7 Structure–Activity Relationships and Inhibitor Design
• • 10.8 Summary
• • References and Further Reading

Chapter 11: Tight-Binding Inhibitors

• • Key Learning Points
• • 11.1 Identifying Tight-Binding Inhibition
• • 11.2 Distinguishing Inhibitor Type for Tight-Binding Inhibitors
• • 11.3 Determining K I for Tight-binding Inhibitors
• • 11.4 Use of Tight-Binding Inhibitors to Determine Active Enzyme Concentration
• • 11.5 Summary
• • References and Further Reading

Chapter 12: Time-Dependent Inhibition

• • Key Learning Points
• • 12.1 Progress Curves for Slow-Binding Inhibitors
• • 12.2 Distinguishing Between Slow-Binding Schemes
• • 12.3 Distinguishing Between Modes of Inhibitor Interaction with Enzyme
• • 12.4 Determining Reversibility
• • 12.5 Examples of Slow-Binding Enzyme Inhibitors
• • 12.6 Summary
• • References and Further Reading

Chapter 13: Enzyme Reactions with Multiple Substrates

• • Key Learning Points
• • 13.1 Reaction Nomenclature
• • 13.2 Bi–Bi Reaction Mechanisms
• • 13.3 Distinguishing Between Random and Compulsory-Ordered Mechanisms by Inhibition Pattern
• • 13.4 Isotope Exchange Studies for Distinguishing Reaction Mechanisms
• • 13.5 Using the King–Altman Method to Determine Velocity Equations
• • 13.6 Cleland’s Net Rate Constant method for Determining v max and v max @k m
• • 13.7 Summary
• • References and Further Reading

Chapter 14: Enzyme–Macromolecule Interactions

• • Key Learning Points
• • 14.1 Mutlitprotein Enzyme Complexes
• • 14.2 Enzyme Reactions on Macromolecular Substrates
• • 14.3 Summary
• • References and Further Reading

Chapter 15: Cooperativity in Enzyme Catalysis

• • Key Learning Points
• • 15.1 Historic Examples of Cooperativity and Allostery in Proteins
• • 15.2 Models of Allosteric Behavior
• • 15.3 Effects of Cooperativity on Velocity Curves
• • 15.4 Sigmoidal Kinetics for Nonallosteric Enzymes
• • 15.5 Summary
• • References and Further Reading

Chapter 16: Evolution of Enzymes

• • Key Learning Points
• • 16.1 Early Earth Conditions
• • 16.2 Natural Selection
• • 16.3 Genetic Alterations
• • 16.4 Enzyme Families and Superfamilies
• • 16.5 Enzyme Promiscuity as a Springboard of Evolution
• • 16.6 Protein Dynamics and Conformational Selection in Evolution of Neofunctionality
• • 16.7 Ancestral Enzyme Reconstruction
• • 16.8 Contemporary Enzyme Evolution
• • 16.9 Summary
• • References and Further Reading

Chapter 17: Enzymes in Human Health

• • Key Learning Points
• • 17.1 Enzymes as Therapeutic Agents
• • 17.2 Enzyme Inhibitors as Therapeutic Agents
• • 17.3 Enzyme Essentiality in Disease
• • 17.4 Enzyme-Mediated Target Protein Degradation
• • 17.5 The Role of Enzymology in Drug Discovery and Development
• • 17.6 Summary
• • References and Further Reading