David M. Smith Div. Org. Chem. and Biochem., Institut Ruđer Bošković, Zagreb

1. Specific Interactions Between Sense and Complementary Peptides:What Can Molecular Dynamics Tell Us? David M. Smith Div. Org. Chem. and Biochem., Institut Ruđer Bošković, Zagreb

2. Sense Peptide N → Tyr-Gly-Gly-Phe-Met → C Translation Sense mRNA (+) 5’ → UAU-CCC-GGC-UUC-AUG → 3’ Transcription Complementary DNA 3’ ← ATA-CCC-CCG-AAG-TAC ← 5’ Sense DNA 5’ → TAT-GGG-GGC-TTC-ATG → 3’ Complementary mRNA (-) 3’ ← AUA-CCC-CCG-AAG-UAC ← 5’ C ← Ile-Pro-Ala-Glu-His ← N Complementary Peptide Complementary Peptides: What are they ? ChemBioChem.2002, 3, 136

3. Some Mekler-Idlis Pairs Sense Complementary Sense Complementary Cys Arg Gly Ser Ser Ala Thr Pro Ala Gly Trp Gly Arg Arg Cys Agr Gly Ser Pro Thr Biophyzika.1969, 14, 581

4. Hydrophilic Hydrophobic Hydrophobic Hydrophilic Hydrophilic Hydrophilic Hydrophobic Hydrophobic Hydropathicity and the Molecular Recognition Theory Biochem. Biophys. Res. Commun.1984, 121, 203

5. Sense Peptide N → Tyr-Gly-Gly-Phe-Met → C Sense mRNA (+) 5’ → UAU-CCC-GGC-UUC-AUG → 3’ Complementary DNA 3’ ← ATA-CCC-CCG-AAG-TAC ← 5’ Sense DNA 5’ → TAT-GGG-GGC-TTC-ATG → 3’ Complementary mRNA (-) 3’ → AUA-CCC-CCG-AAG-UAC →5’ N → Ile-Pro-Pro-Lys-Tyr →C Complementary Peptide An Alternative Definition of Complementary Peptides: Pro. Nat. Acad. Sci.1985, 82, 1372

6. Some Root-Bernstein Pairs Sense Complementary Sense Complementary Arg Arg Arg Arg Pro Pro Pro Pro Ala Gly Gly Gly Gly Gly Cys Cys Trp stop Pro Thr J. Theor. Biol.1983, 100, 99

7. The System Ace-Tyr-Gly-Gly-Phe-Met-Nme Ace-Ile-Pro-Pro-Lys-Tyr-Nme Croat. Chem. Acta.1998, 71, 591

8. Bonds Angles Dihedrals Van der Waals Electrostatic Implicit Solvation (Non Elec.) Implicit Solvation (Elec.) Molecular Dynamics: The Force Field

9. Molecular Dynamics in Practise In principle, doing MDs is simply a matter of solving Newton’s equations of motion In practise we must numerically integrate these equations with a finite time step (typically 1-2 fs)

10. Molecular Dynamics in Action 50 ps of MD at 300 K with implicit solvation

11. Analysis: Structural Deviations Root Mean Square Deviation from a reference structure vs time

12. Cluster 1, Pop. = 28% <E> = -57.5 kcal/mol Cluster 2, Pop. = 45% <E> = -58.6 kcal/mol Backbone Overlay Cluster 3, Pop. = 27% <E> = -58.5 kcal/mol Minimum Energy Structure E= -16.7 kcal/mol Analysis: Clustering

13. Analysis: Clustering 28% 45% Implicit solvent, 150 ns 27%

14. Principle Component Analysis Projections onto the eigenvectors of the covariance matrix

15. Principle Component Analysis 28% 45% What about the force field? FF94, Implict solvent, 150 ns 27%

16. Differences in the Force Field 12% 27% FF99, Implict solvent, 150 ns 4% 58%

17. Differences in the Force Field 51% 33% FF03, Implict solvent, 150 ns 16%

18. NMR, experiments in binary bilayered mixed micelles (bicelles) show a well-defined structure: Biophys. J.2004, 86, 1587 What Does Experiment Say NMR experiments in water show an essentially random distribution of conformers

19. Explicit Solvation 50 ps of MD at 300 K with explicit solvation (NVT)

20. Explicit Solvation: Analysis 54 % 27% 19% 40 ns of MD at 300 K with explicit solvation (FF03)

21. A modern solution is to construct several replica simulations with different temperatures and allow them to exchange according to: Replica Exchange Dynamics Single simulations, especially in explicit solvent, are prone to become trapped in potential energy minima Increasing the temperature can facilitate barrier crossings but can lead to irrelevant results

22. Replica Exchange Dynamics 16 replicas simulated for 2.5 ns each, implying 40ns in total. The temperatures range between 275K and 420K such that P≈0.2.

23. Replica Exchange Dynamics 27% 40% 33% 40 ns (16 x 2.5) of MD at 275-420 K with explicit solvation (FF03)

24. Non-Exchanging Replicas Replica exchange MD is an inherently parallel method An alternative approach is to construct several non- interacting replicas (distributed computing) An efficient way to implement this is to first run one simulation at high temperature and to cluster the results The structure closest to the centroid of each cluster can then be used as a starting point for each replica

25. Non-Exchanging Replicas 30% 37% 33% 40 ns (8 x 5) of MD at 300 K with explicit solvation (FF03)

26. Sense and Complementary Peptides 1 ns of MD at 300 K with explicit solvation

27. Sense and Complementary Peptides 19% 30% 16% 23% 13% 40 ns (8 x 5) of MD at 300 K with explicit solvation

28. A Closer Look at the Clusters Cluster 1: Population 30 %

29. A Closer Look at the Clusters Cluster 2: Population 19 %

30. A Closer Look at the Clusters Cluster 3: Population 23 %

31. A Closer Look at the Clusters Cluster 4: Population 13 % Cluster 5: Population 16 %

32. Analysis: Structural Properties Separation of the centres of mass of the two peptides

33. Analysis: Structural Properties Separation of the centres of mass of the complementary residues

34. Conclusions • The force field can have a strong influence on the structural • properties. FF03 can probably be trusted. • Non-interacting replicas constitute a good approximation to • the replica exchange method and a good alternative to a single • long simulation, at least for small peptides. • Met-enkephalin does not have a well-defined native structure • in aqueous solution at 300 K. • Met-enkephalin does exhibit some affinity for its complementary • counterpart but this is apparently not based on the specific • interactions predicted by the Molecular Recognition Theory.

35. Cluster 1, Pop. = 51% <E> = 0.4 kcal/mol Cluster 2, Pop. = 33% <E> = -0.5 kcal/mol Backbone Overlay Cluster 3, Pop. = 16% <E> = 0.4 kcal/mol Minimum Energy Structure E= -16.7 kcal/mol Analysis: Clustering (ff03)

36. Analysis: Clustering (ff03) 51% 33% 16%

37. Principle Component Analysis (ff03) 51% 33% 16%