1. Simulating the first steps of amyloid peptides aggregation
Jessica Nasica
Département de physique - Université de Montréal
GEPROM – 2ième réunion scientifique
May 27th, 2009

2. Overview of the presentation
Medical interest
Problem of amyloid fibril formation
Our simulations & results

3. Protein misfolding and aggregation
Many protein-misfolding diseases include conditions where a protein
forms insoluble aggregates, amyloid fibrils, that deposit toxically
Amyloid diseases
e.g. :
Alzheimer’s
Parkinson’s
Huntington
type II diabetes
Dobson et al. (2003), Nature 426

4. Amyloid fibril formation
Amyloid fibrils insoluble fibrous aggregates with a highly organized
Macrostructure made of β-sheets.
Mechanism of formation  3 steps:
Alignment of the molecules to form β-sheets  fastest stage  involves H-bonds
Formation of the cross-β structure  slower than step 1  involves Van-der-Waals forces  interdigitation of residues side chains  “steric zipper” structure
Fibril formation  involves non-covalent bonds
Amyloid fibril formation is a nucleated-growth process stabilized by the
protein concentration and by the formation of steric zippers
Nelson et al. (2005), Nature 435

5. Amyloid fibril polymorphism
For the Aβ40 sequence, they construct a full molecular model
showing 2 distinct possible morphologies for the fibril structure.
Paravastu et al. (2008), PNAS &
Petkova et al. (2006), Biochemistry 45

6. Amyloid oligomers
intermediate states during the formation of amyloid fibrils
thought to be more toxic than the fibrils themselves
Different types of soluble amyloid oligomers share a common structure  suggests they share a common toxicity mechanism
Lashuel et al. (2002), Nature 418
Kayed et al. (2003), Science 300

7. Theories on intermediate oligomer states
Oligomers as protofibrils
Oligomers as pores  may form annular pore-like structures to go through the cell membranes : β-barrel
Esposito et al. (2006), PNAS 103
Amyloid oligomer pore structure observed experimentally (left) & numerically (right)
Lashuel et al. (2008), Nature 418
Irbäck et al. (2007), Proteins 71

8. The “short peptides” approach
Nelson et al. (2006), Current opinion in structural biology 16
The gain-of-interaction model for amyloid structure suggests that
only a small portion of a native protein is responsible for amyloid fibrils.
Under conformational changes
this small portion is exposed & binds to an identical portion on another molecule
builds up a fibril or
Cross-β spine (no domain swapping) Cross-β spine with domain swapping
For the budding yeast Sup35p fibril-forming protein
GNNQQNY
GNNQQNY
The essential element involved in the fibril structure is the small portion

9. Experiments done on GNNQQNY
Atomic structure of cross-β spine constructed from X-ray diffraction analysis
Fomation of a double β-sheet with parallel β-stands
Side chains form a self-complementing steric zipper
Interdigitation of the side-chains would stabilize the sheets
Nelson et al. (2005), Nature 435

10. Our short-term goals
To understand the aggregation process of short peptides
Kinetics of aggregation
Final structures (morphologies accessible)
To study different sizes of systems
Trimer GNNQQNY
Pentamer GNNQQNY
20-mer GNNQQNY
50-mer GNNQQNY

11. Our simulation methods
Replica exchange MD
launch n molecular dynamical simulations in parallel, at n different temperatures
at regular intervals, try an exchange of configurations between two adjacent temperatures using a Metropolis accept-reject criterion
REMD accelerates sampling (in some cases) for the cost of losing dynamical information.
REMD still provides thermodynamical information
Final structures of simulations tested with an all-atom potential (GROMACS)

12. Our simulations : small aggregates
We observe a strong tendency to form planar β-sheets
GNNQQNY trimer
GNNQQNY pentamer

13. Our simulations : bigger aggregates
Formation of twisted pair-of-sheet “protofibril-like” structures
Formation of β-barrel-like structures
GNNQQNY 20-mer – 300 ns simulation
Similar to the atomic structure described by Nelson et al. (2005)

14. GNNQQNY 20-mer results
We observe a nucleated-growth aggregation process
&
a clear loss of entropy as the energy of the system drops rapidly

15. GNNQQNY 20-mer results
β-sheets favor a parallel orientation of the β-strands
Consistent with the atomic description given by Nelson et al. (2005)
Statistics over 300 ns

16. Conclusions
We have described the first steps of aggregation of GNNQQNY. The results are consistent with experimental results on that sequence.
Polymorphism exists and we already see a clear separation between 2 different semi-organized structures.
The obtained β-barrel-like structures might not be on the fibril formation pathway. It seems it is a separate possible morphology.
For small aggregates (trimers, pentamers), the study of 2 other sequences (SSTSAA,SNQNNF) shows similar features compatible with the GNNQQNY results.

17. Future work
Lots of statistics obtained from our 20-mer simulations  a lot of the analysis is still being done to understand
The formation process
The triggering of the nucleated-growth process
Simulation of the GNNQQNY 50-mer
Simulation on other short sequences (SSTSAA, SNQNNF,…)

18. Acknowledgments Collaborators MONTREAL Normand Mousseau PARIS
Philippe Derreumaux
MILAN
Giorgio Colombo
Massimiliano Meli (Ph.D.)
Funding
GEPROM
CRSNG
FESP
Resources
Réseau québécois de calcul de haute performance
Special thanks to:
Rozita Laghaei (post-doc), Lilianne
Dupuis (Ph.D.) and Jean-François St-
Pierre (Ph.D.)