1. Simulations of the folding and aggregation of peptides, proteins and lipids. BRISBANE School of Molecular and Microbial Sciences (SMMS) Chemistry Building (#68) University of Queensland Brisbane, QLD 4072, Australia Email a.mark@uq.edu.au Phone: +61-7-33469922 FAX: +61-7-33654623 Centre Secr: +61-7-33653975 GRONINGEN Lab. of Biophysical Chemistry University of Groningen Nijenborgh 4 email 9747 AG GRONINGEN The Netherlands tel +31.50.3634457 fax: +31.50.3634800 tel secr: +31.50.3634323 email:a.e.mark@rug.nl secr: mdsecr@fmns.rug.nl http://md.chem.rug.nl Alan E. Mark Herman Berendsen Siewert-Jan Marrink

2. Peptide folding and assembly: Our best example of peptide folding to date is a the beta-hexapeptide shown on the following slides (solvent Methanol). 1.This system is fully reversible. 2.We have simulations of this and other systems to > 200ns at temperatures from 180 -> to 450K. 3.We have replica exchange simulations of a slightly modified system showing 1000’s of individual folding events. 4.As far as we can determine our modified system approaches full convergence in 200-400 ns. 5. Trajectories are available.

3.  - Peptides i)  -amino-acids (additional backbone carbon) ii)Stable 2nd structure. iii)Non-degradable peptide mimetics (e.g. highly selective somatastatin analogue) D. Seebach, B. Jaun + coworkers organic chem ETH-Zurich  -Heptapeptide (M) 31-helix in MeOH at 298 K (left-handed) Daura, X., Bernhard, J., Seebach, D., van Gunsteren, W. F. and Mark, A. E. (1998) J. Mol. Biol. 280, 925-932.

4. unfoldfoldunfoldfold unfold  -Heptapeptide, 340 K

5. Starting structure  -Heptapeptide, 360 K  G folding = -RT ln (folded/unfolded)

6. Predict Probability of Individual Microstates in Solution  G=~6 kJ/mol  G=~8 kJ/mol  G=0 kJ/mol  G=~9 kJ/mol Daura, X., van Gunsteren, W. F. and Mark, A. E. (1999) Proteins: Struct. Funct. Genet. 34, 269-280.

7. Folding Pathways

8. Simulations of peptide folding As part of our program we are looking a range of larger peptides. So far getting reversible folding from random starting structures has proved difficult for systems > 20 a.a. In particular we are investigating a series of related helical peptides (~20 a.a.) with fast folding kinetics AP A 5 (A 3 RA) 3 A YGA Ac-YG(AKA 3 ) 2 AG-NH 2 YGG Ac-YGG(KA 4 ) 3 K-NH 2 So far results are limited but we have seen reversible transitions. An example is given below.

9. AP A 5 (A 3 RA) 3 A Ref: Lednev I. K. et al. J. Am. Chem. Soc. 1999, 121, 8074-8086.  A 21 amino acid, mainly alanine, α-helical peptide (AP).  The folding/unfolding activating barriers based on an nanosecond UV resonance Raman study.  ~8 kcal/mol activation barrier; reciprocal rate constant ~240±60 ns at 37 °C (310 K). MD simulation start from the α-helix structure The GROMOS 45A3 force field was adopted

10. Coil β-Sheetβ-BridgeBendTurnα-helix5-Helix3-Helix Time (ps) Residue Secondary structure The secondary structure as a function of time shows one refolding transition in 100ns.

11. N-ter C-ter 0 ns (starting structure) N-ter C-ter 10 ns N-ter C-ter 30 ns C-ter N-ter 50 ns N-ter C-ter 75 ns N-ter C-ter 70 ns N-ter C-ter 80 ns N-ter C-ter 85 ns N-ter C-ter 100 ns

12. Other peptide systems on which we have simulations showing partial folding or assemble include: 1.Various amyloid forming peptides on surfaces. 2.Betanova (a designed triple stranded peptide) 3.A series of coiled-coils. 4.WW domain peptide (~20 a.a. peptide studied by replica exchange) 5.Several proteins showing recovery from mild denaturing conditions.

13. Spontaneous Aggregation of Lipids and Surfactants I believe this is one area where complexity analysis should be able to perform well as the systems show spontaneous generation of order. We have multiple simulations of: 1.Bilayer formation (course grained and in atomic detail) 2.Vesicle formation (course grained and in atomic detail) 3.Phase transitions (course grained and in atomic detail) 4.Membrane and vesicle fusion. Note: these are highly reproducible collective processes involving 100’s to 1000’s of lipids. A few examples are given below.

14. S.J. Marrink A C eq CB D eq C*C* Spontaneous assembly of phospholipds into a bilayer 0 ns0.2 ns 3 ns 10 ns 20 ns 25 ns

15. Density Evolution Showing the Generation of Order density water head groupslipid tails S.J. Marrink