1. Peptide Aggregation and Pore Formation in a Lipid Bilayer; a Combined CG and AA MD Study Lea Thøgersen, University of Aarhus Pushing the Boundaries of Biomolecular Simulation June 11, 2008

2. Outline Introduction Results Conclusions  Coarse-Grained Molecular Dynamics  Alamethicin  Modelling Setup  NAMD CG vs MARTINI CG  Microsecond Action  Structure  Reverse Coarse-Graining  Water Pore Formation  Structural Changes

3. Coarse Grained MD – Why? Reduction in degrees of freedom Fast frequenzy movements removed Smoother potential surface Longer time steps can be taken Microsecond simulations possible All-atom MD: time step of 1-2 fs, time frame sampled ~ 100 ns Coarse-grained MD: time step of 20-50 fs, time frame sampled ~ 1 μs Coarse-graining

4. Coarse Grained Molecular Dynamics Shelley, Shelley, Reeder, Bandyopadhyay, Klein; A Coarse Grain Model for Phospholipid Simulations J. Phys. Chem. B (2001) 105 4464 Shih, Arkhipov, Freddolino, Schulten; Coarse Grained Protein-Lipid Model with Application to Lipoprotein Particles J. Phys. Chem. B (2006) 110 3674 NAMD CG Marrink, de Vries, Mark; Coarse Grained Model for Semiquantitative Lipid Simulations J. Phys. Chem. B (2004) 108 750 MARTINI CG Marrink, Risselada, Yefimov, Tieleman, de Vries; The MARTINI Force Field: Coarse Grained Model for Biomolecular Simulations J. Phys. Chem. B (2007) 111 7812 Monticelli, Kandasamy, Periole, Larson, Tieleman, Marrink; The MARTINI Coarse-Grained Force Field: Extension to Proteins J. Chem. Theory and Comput. (2008) 4 819

5. 20 amino acid antimicrobiel peptide. Part of the immune system (for fungi). Forms channels in membranes which allow water and ions to go through. Destroys membrane potential. Alamethicin Motivation: Potentially a good and simple membrane channel model Insight into this family of proteins could be valuable in the development of antibiotics Gln7 Glu18 Gln19

6. Widely Accepted Channel Model Tieleman, Hess, Sansom; Analysis and Evaluation of Channel Models: Simulations of Alamethicin. Biophys. J. (2002) 83 2392 Spaar, Münster, Salditt; Conformation of Peptides in Lipid Membranes Studied by X-Ray Grazing Incidence Scattering. Biophys. J. (2004) 87 396

7. Modelling Setup 25 peptides 330 DMPC lipids 26452 water ~117000 atoms ~ 11700 beads PBC 120 Å × 124 Å × 90 Å To study alamethicin interaction with membrane and each other

8. Apolar Polar Nonpolar Charged hydrogenbond donor acceptor both none hydrogenbond donor acceptor both none NAMD CG versus MARTINI CG NAMD CGMARTINI CG 1 4 4 1 2 1 2 4 2 degree of polarity 1-5 (MARTINI) (NAMD) AA GLYALAAIB NAMD MARTINI Nda N0 C5C4 CCCNda

9. 32 ns 7 ns Alamethicin Behavior Alamethicin Hydrophilic sidechains Lipid Polar headgroup Non-polar tails C-term N-term 120 ns

10. Peptide Aggregation I 0-1 μs

11. Peptide Aggregation II 0 μs1 μs4 μs MARTINI NAMD

12. Structure of the Clusters 0 0.5 1 Gln7

13. Helix Tilt Alamethicin, DMPC lipid peptid:lipid - 1:15 15 N-Aib8 alamethicin Exp θ = 10˚ MD Vosegaard, Bertelsen, Pedersen, Thøgersen, Schiøtt, Tajkhorshid, Skrydstrup, Nielsen; Resolution Enhancement in Solid-State NMR of Oriented Membrane Proteins by Anisotropic Differential Linebroadening JACS (2008) 130 5028 θ Aib8

14. Reverse Coarse-Graining CG t = 0μs CG t = 1μs AA - rev CG t = 1μs AA after SA AA t = 0μs

15. CG water vs AA water 900-1000 ns δ-δ- δ+δ+ δ+δ+ Class: Polar 5.0Å2.8Å TIP3P model 35-40 ns

16. Structural Changes 18 of 25 peptides remain α-helical

17. Conclusions Alamethicin monomers readily aggregate and form clusters that grow in size over time. Large diversity in form of clusters and structure of peptides. Reverse CG required to  obtain detailed water interaction  validate results obtained on the long time scale

18. Acknowledgements Emad Tajkhorshid & the NAMD people Theoretical and Computational Biophysics Group University of Illinois at Urbana-Champaign, USA Birgit Schiøtt & the Biomodelling Group Department of Chemistry University of Aarhus, Denmark Niels Christian Nielsen & Thomas Vosegaard Laboratory for Biomolecular NMR spectroscopy Department of Chemistry University of Aarhus, Denmark Funding: &