1. INTERACTIONS IN PROTEINS AND THEIR ROLE IN STRUCTURE FORMATION

2. Levels of protein structure organization

4. Dominant forces in protein folding Electrostatic forces Hydrogen bonding and van der Waals interactions Intrinsic properties Hydrophobic forces Conformational entropy (opposes folding)

5. Can we say that there are „dominant” forces in protein folding? Hardly. Proteins are only marginally stable (5 – 20 k B T/molecule). For comparison: water-water H-bond has about 5 kcal/mol (9 k B T/molecule) Consequently, even the tiniest force cannot be ignored. However, different types of interactions play different role Hydrophobic interaction: compactness Local interactions: chain stiffness Hydrogen bonds: architecture

6. Local and nonlocal interactions

7. Long-range vs. short-range interactions n<=3: long range interactions n>3: short-range interactions Long-range: electrostatic (charge-charge, charge-dipole, and dipole-dipole) interactions Short-range: van der Waals repulsion and attraction, hydrophobic interactions

8. Electrostatic interactions

9. Lots of like-charges (e.g., side-chain ionization by pH decrease/increase) destabilize protein structure Increase of ionic strength destabilizes protein structure 5 – 10 kcal/mol / counter-ion (salt-bridge) pair A protein contains only a small number of salt bridges, mainly located on the surface (nevertheless, they can be essential).

10. Example of a surface salt bridge: salt bridge triad between Asp8, Asp12 and Arg110 on the surface of barnase

11. Replacement of charged residues with hydrophobic residues can increase the stability by 3-4 kcal/mol. Example: ARC repressor Wild type: salt triad between R31, E36, and R40 Mutant: hydrophobic packing between M31, Y36, and L40

13. Potentials of mean force

14. Maksimiak et al., J.Phys.Chem. B, 107, 13496-13504 (2003)

15. Masunov & Lazaridis, J.Am.Chem.Soc., 125, 1722-1730 (2003)

17. Hydrogen-bonding and van der Waals forces

18. A w +B w A n +B n (AB) w (AB) n  G 1 =-2.40 kcal/mol  G 3 =+3.10 kcal/mol  G 4 =+0.62 kcal/mol  G 3 =+3.10 kcal/mol Free energies of N-methylamide dimerization in water (w) and CCl 4 (n) solution and transfer between these solvents

19. Local interactions are largely determined by Ramachandran map

20. Conformations of a terminally-blocked amino-acid residue C 7 eq C 7 ax E Zimmerman, Pottle, Nemethy, Scheraga, Macromolecules, 10, 1-9 (1977)

21. Energy maps of Ac-Ala-NHMe and Ac-Gly-AHMe obtained with the ECEPP/2 force field

22. Energy curve of Ac-Pro-NHMe obtained with the ECEPP/2 force field  L-Pro  -68 o

23. Energy minima of therminally-blocked alanine with the ECEPP/2 force field

24. Hydrophobic forces

30. Sobolewski et al., J.Phys.Chem., 111, 10765-10744 (2008)

32. Dependence of the PMF and cavity contribution to the PMF of two methane molecules on temperature (Sobolewski et al., PEDS, 22, 547-552 (2009)

33. S. Miyazawa & R.L. Jernigan, R. L. 1985. Estimation of effective interresidue contact energies from protein crystal structures: quasi-chemical approximation. Macromolecules, 18:534-552, 1985.

34. CMFILVWYAGTSQNEDHRKP P K R H D E N Q S T G A Y W V L I F M C Color map of the MJ table

35. Conformational entropy