A Hitch-Hiker’s Guide to Molecular Thermodynamics What really makes proteins fold and ligands bind

1. A Hitch-Hiker’s Guide to Molecular ThermodynamicsWhat really makes proteins fold and ligands bind Alan Cooper Chemistry Department Joseph Black Building, Glasgow University Glasgow G12 8QQ, Scotland Amsterdam: November 2002

2. + “Concepts and tools for medicinal chemists” What makes this protein fold, and what controls its stability ?

3. + “Concepts and tools for medicinal chemists” What makes this protein fold, and what controls its stability ? What are the thermodynamic forces responsible for ligand binding ? Can we use them to design better ligands ?

4. “ Concepts and tools for medicinal chemists” Thermodynamic homeostasis, compensation; hydrogen-bonded lattices… ...the role of water in biomolecular interactions Microcalorimetry: analytical uses for biomolecular interactions and stability

5. A bluffer’s guide to Thermodynamic Equilibrium… There is a natural tendency for all things (even atoms & molecules) to roll downhill - to fall to lower energy. H wants to be negative This is opposed (at the molecular level) by the equally natural tendency for thermal/Brownian motion (otherwise known as “entropy”) to make things go the other way… …and this effect gets bigger as the temperature increases. T.S wants to be positive

6. Thermodynamic Equilibrium, expressed in terms of the Gibbs Free Energy change, reflects just the balance between these opposing tendencies… G = H - TS Equilibrium is reached when these two forces just balance (G = 0). The standard free energy change, G, is just another way of expressing the equilibrium constant, or affinity (K) for any process, on a logarithmic scale… G = -RTlnK

7. Both enthalpy and entropy are integral functions of heat capacity... ….from which DG = DH - T.DS So DCp is the key - if we can understand heat capacity effects, then we can understand everything else.

8. Calorimetric techniques... • Differential scanning calorimetry (DSC) • Isothermal titration calorimetry (ITC) • Pressure perturbation calorimetry (PPC)

9. So, what is the role of water? So DCp is the key - if we can understand heat capacity effects, then we can understand everything else. And DCp is largely determined by the interactions between water and the macromolecule(s). In figure b many more waters are free than in a. And free waters are happy waters!

10. DG=DH-TDS DG=-RTln(K) ΔG must negative for a reaction to take place. ΔG = 1.38 kCal/Mole means a factor 10 difference in an equilibrium. Example: A <==> B [A] = [B] G=17.2 for [A] and for [B], so we have a 50/50 equilibrium (it is impossible to know that G=17.2, we can only know that ΔG is 0; but lets pretend…) If we make G=18.6 for [A] (again, this is nonsence because we cannot know G, only ΔG) (so, G is 1.38 bigger for [A] which means better for [B]) then [B] becomes 10 times bigger than [A].

11. DG=DH-TDS Good for ΔH: 1) Contacts in protein (H-bonds, Van der Waals interactions, salt bridges, aromatic stacking, etc). 2) H-bonds between water molecules Bad for ΔH: 1) H-bonds between water and part of protein that gets buried.

12. DG=DH-TDS Good for ΔS: Entropy of water. Bad for ΔS: Entropy of protein.