Biophysical Approaches to Drug Design

1. Biophysical Approaches to Drug Design Overall Organization of the Topic • Approaches Useful in Drug Design • Fluorescence Spectroscopy - Principle of Fluorescence - Factors governing fluorescence intensity - Fluorescent groups - Sensitivity of Fluorescence to Environment - Fluorescence Resonance Energy Transfer Biophysical Approaches

2. Biophysical Approaches Useful in Drug Design • Absorption Spectroscopy • Fluorescence Spectroscopy • Infrared Spectroscopy • Raman Spectroscopy • Nuclear Magnetic Resonance Spectroscopy • Electron Spin Resonance Spectroscopy • Circular Dichroism • Centrifugation Techniques • X-ray Crystallography Biophysical Approaches

3. Fluorescence Spectroscopy Principle Biophysical Approaches

4. Electronic States of Molecules Example of Ethylene Biophysical Approaches

5. Obtaining Fluorescence Spectrum Biophysical Approaches

6. Factors Governing Fluorescence Intensity • Internal Conversion (kIC) • Collisional De-excitation (kQ) • Intersystem Crossing (kIS) Biophysical Approaches

7. + + + + _ _ _ _ Tryptophan Tyrosine Phenylalanine Cysteine Adenosine Guanosine Fluorescent Groups • Intrinsic Groups in Proteins and Nucleic Acids - Trp, Tyr, Phe, Cys, Adenine, Guanine, … Biophysical Approaches

8. Fluorescent Groups • External Probes Biophysical Approaches

9. Sensitivity of Fluorescence to Environment • Determination of Equilibrium Dissociation Constants Biophysical Approaches

10. Mutant KDDGO (nM) (kcal/mol) WT 2 -11.9 R47H 59 -9.9 R129H 820 -8.6 K125M 570 -8.4 K114A 10,000 -6.7 Binding Affinities as a Function of Structure • Varying the Receptor • Varying the Ligand Ligand KDDGO (nM) (kcal/mol) DEFGH’ ~1×10-5-19 DEFGH ~1×10-3 -16 DEFG 34 -10.2 DEF 2,000 -7.8 EFGH 6,900 -7.0 Biophysical Approaches

11. Fluorescence Resonance Energy Transfer Principle Biophysical Approaches

12. Fluorescence Resonance Energy Transfer Determining Substrate Specificities Biophysical Approaches