| |
| |
Foreword | |
| |
| |
Preface | |
| |
| |
Acknowledgments | |
| |
| |
| |
Why Enzymes as Drug Targets? | |
| |
| |
| |
Enzymes Are Essentials for Life | |
| |
| |
| |
Enzyme Structure and Catalysis | |
| |
| |
| |
Permutations of Enzyme Structure During Catalysis | |
| |
| |
| |
Other Reasons for Studying Enzymes | |
| |
| |
| |
Summary | |
| |
| |
References | |
| |
| |
| |
Enzyme Reaction Mechanisms | |
| |
| |
| |
Initial Binding of Substrate | |
| |
| |
| |
Noncovalent Forces in Reversible Ligand Binding to Enzymes | |
| |
| |
| |
Electrostatic Forces | |
| |
| |
| |
Hydrogen Bonds | |
| |
| |
| |
Hydrophobic Forces | |
| |
| |
| |
van der Waals Forces | |
| |
| |
| |
Transformations of the Bond Substrate | |
| |
| |
| |
Strategies for Transition State Stabilization | |
| |
| |
| |
Enzyme Active Sites Are Most Complementary to the Transition State Structure | |
| |
| |
| |
Steady State Analysis of Enzyme Kinetics | |
| |
| |
| |
Factors Affecting the Steady State Kinetic Constants | |
| |
| |
| |
Graphical Determination ofk cat and K M | |
| |
| |
| |
Reactions Involving Multiple Substates | |
| |
| |
| |
Bisubstrate Reaction Mechanisms | |
| |
| |
| |
Summary | |
| |
| |
References | |
| |
| |
| |
Reversible Modes of Inhibitor Interactions with Enzymes | |
| |
| |
| |
Enzyme-Inhibitor Binding Equilibria | |
| |
| |
| |
Competitive Inhibition | |
| |
| |
| |
Noncompetitive Inhibition | |
| |
| |
| |
Mutual Exclusively Studies | |
| |
| |
| |
Uncompetitive Inhibition | |
| |
| |
| |
Inhibition Modality in Bisubstrate Reactions | |
| |
| |
| |
Value of Knowing Inhibitor Modality | |
| |
| |
| |
Quantitative Comparisons of Inhibitor Affinity | |
| |
| |
| |
RelatingK i to Binding Energy | |
| |
| |
| |
Defining Target Selectivity byK i Values | |
| |
| |
| |
Potential Advantages and Disadvantages of Different Inhibition Modalities In Vivo | |
| |
| |
| |
Knowing Inhibition Modality Is Important for Structure-Based Lead Organization | |
| |
| |
| |
Summary | |
| |
| |
References | |
| |
| |
| |
Assay Considerations for Compound Library Screening | |
| |
| |
| |
Defining Inhibition Signal Robustness, and Hit Criteria | |
| |
| |
| |
Measuring Initial Velocity | |
| |
| |
| |
End-Point and Kinetic Readouts | |
| |
| |
| |
Effects of Enzyme Concentration | |
| |
| |
| |
Balanced Assay Conditions | |
| |
| |
| |
Balancing Conditions for Multisubstrate Reactions | |
| |
| |
| |
Order of Reagent Addition | |
| |
| |
| |
Use of Natural Substrates and Enzymes | |
| |
| |
| |
Coupled Enzyme Assays | |
| |
| |
| |
Hit Validation and Progression | |
| |
| |
| |
Summary | |
| |
| |
References | |
| |
| |
| |
Lead Optimization and Structure-Activity Relationships for Reversible Inhibitors | |
| |
| |
| |
Concentration-Response Plots and IC 50 Determination | |
| |
| |
| |
The Hill Coefficient | |
| |
| |
| |
Graphing and Reporting Concentration-Response Data | |
| |
| |
| |
Testing for Reversibility | |
| |
| |
| |
Determining Reversible Inhibition Modality and Dissociation Constant | |
| |
| |
| |
Comparing Relative Affinity | |
| |
| |
| |
Compound Selectivity | |
| |
| |
| |
Associating Cellular Effects with Target Enzyme Inhibition | |
| |
| |
| |
Cellular Phenotype Should Be Consistent with Genetic Knockout or Knockdown of the Target Enzyme | |
| |
| |
| |
Cellular Activity Should Require a Certain Affinity for the target Enzyme | |
| |
| |
| |
Buildup of Substrate and/or Diminution of Product for the Target Enzyme Should Be Observed in Cells | |
| |
| |
| |
Cellular Phenotype Should Be Reversed by Cell-Permeable Product or Downstream Metabolites of the Target Enzyme Activity | |
| |
| |
| |
Mutation of the Target Enzyme Should Lead to Resistance or Hypersensitivity to Inhibitors | |
| |
| |
| |
Summary | |
| |
| |
References | |
| |
| |
| |
Slow Binding Inhibitors | |
| |
| |
| |
Determiningk obs : The Rate Constant for Onset of Inhibition | |
| |
| |
| |
Mechanisms of Slow Binding Inhibition | |
| |
| |
| |
Determination of Mechanism and Assessment of True Affinity | |
| |
| |
| |
Potential Clinical Advantages of Slow Off-rate Inhibitors | |
| |
| |
| |
Determining Inhibition Modality for Slow Binding Inhibitors | |
| |
| |
| |
SAR for Slow Binding Inhibitors | |
| |
| |
| |
Some Examples of Pharmacologically Interes | |