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Simulations of Peptide Aggregation. Joan-Emma Shea Department of Chemistry and Biochemistry The University of California, Santa Barbara. Protein and Peptide Aggregation. PROTEIN AGGREGATION AND DISEASE. Parkinson. Alzheimer. Prion (“Mad Cow”). Huntington. - PowerPoint PPT Presentation
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Simulations of Peptide Aggregation
Joan-Emma Shea
Department of Chemistry and BiochemistryThe University of California, Santa Barbara
Protein and Peptide Aggregation
PROTEIN AGGREGATION AND DISEASE
Alzheimer
Huntington
Parkinson
Prion(“Mad Cow”)
Proteins not associated with a specific disease can also aggregate to form amyloid fibrils
APPLICATION TO BIOMATERIALS
Nano-tube and nano-sphere fabrication using aromatic di-peptides. [Gazit et al., Nano Lett, 4 (2004) 581]
Use of peptides to form nanoscale-ordered monolayers, COSB 2004, 14: 480
Time scalesSide-chain rotations
Loop closure
Helix formation Folding of-hairpins
Protein folding
Protein aggregation
All-atom: MolecularDynamics (MD)With EXPLICIT Solvent
MULTISCALE APPROACH
Off-lattice minimalist:Langevin dynamics
All-atom: MolecularDynamics (MD) withIMPLICITsolvent
COARSE GRAINING
1. Dimerization Mechanisms of 4 tetrapeptides KXXE
2. Design of Inhibitors of aggregation of the AlzheimerAmyloid-beta (A) peptide.
Fibrilsof KFFE
KXXE peptides
Tjernberg et al. , JBC 277 (2002) 43243
KAAE
KVVE KFFE
KLLE
Fibrils!
PEPTIDE MODEL
CHARMM19 Force Field and Generalized Born Implicit Solvent
1010
54)(
RrRrr
RerU pc
R
ep=1 kCal/mol, =1Å,
R=17Å
Confining Sphere:
NVTNVTT1
TN
T2
))((,1min mnij EEe
Simulation protocol: Replica Exchange MD6 replicas for monomer: total simulation time: 40 ns12 replica for dimers: total simulation time: 400 ns
Heterogeneous dimers
RMSD (Å)
E (
kC
al/m
ol)
Association temperature Ta
KFFE > KVVE > KLLE > KAAE
Thermodynamicstability of dimers:
Aa
La
Va
Fa TTTT
What determines transition temperature Ta?
Ta=
SU / translconfig SS
Propensity for structureSalt bridgesHydrophobic contactsAromatic interactions
monomererchain UU int
T=325K
T=240K T=275K
Free energy as a function of interaction energy and radius of gyration
T=285K
U (kCal/mol)
Rg
(Å
)
U
UU
T=325K
T=240K T=275K
Free energy as a function of interaction energy and radius of gyration
T=285K
U (kCal/mol)
Rg
(Å
)
U
UU
Interaction energy: KFFE > KLLE ~ KVVE > KAAE
Most favorable
Least favorable
Entropy loss due to dimerization
Monomers, Rg>5A
-strand
RandomCoil
Helix
Entropy loss due to dimerization
Monomers, Rg>5A
-strand
RandomCoil
Helix
Entropy loss due to dimerization
Monomers, Rg>5A Dimers, Rg<5A
-strand
RandomCoil
Helix
Entropy loss due to dimerization
Monomers, Rg>5A Dimers, Rg<5A
-strand
RandomCoil
Helix
-strand basin more populated for monomerof KVVE than KLLE: SL > SV
Association temperature
SUTa /
Energetic effect
Entropic effect
FVAL SSSS
ALVF UUUU ~
Aa
La
Va
Fa TTTT
KFFE KLLE
Kinetic accessibility of dimers
E (Kcal/mol) E (Kcal/mol)
Rg
(Å
)
KFFE KLLE
“DOWNHILL” DIMERIZATION FOR KFFE
FREE ENERGY BARRIERFOR KLLE
Different mechanisms of dimerization
PHE-PHE come together first (stabilized by vdw interactions), followed by the overall collapse of the structure with formation of peptide backbones contacts .
Rg
ove
r C
Rg over PHE atoms Rg over LEU atoms
LEU side chain formation and overall collapse with formation ofpeptide backbone interactions occur simultaneously.
KFFE KLLE
CONCLUSIONS
Aa
La
Va
Fa TTTT for dimerization of the
KXXE peptides1.
2. Strong sequence dependence of free energy landscapes for dimerization, with KFFE experiencing a barrierless transition.
3. KFFE dimers are the most thermodynamically stable and kinetically accessible.
4. Dimer trends match experimental trends observed for fibrils.
Aβ40: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVV
Aβ42: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVVIA
LARGER SYSTEMS: OLIGOMERIZATION OF THE ALZHEIMER AMYLOID BETA (A) PEPTIDES
AMYLOID BETA (A) PEPTIDES AGGREGATE TO FORM TOXIC OLIGOMERS AND FIBRILS
Aβ40: DAEFRHDSGYEVHHQKLVFFAEDV25GSNKGAIIGL35MVGGVV
FRAGMENT 25-35 OF THE (A) PEPTIDE APPEARS TO BETOXIC IN MONOMERIC, SMALL OLIGOMERIC AND FIBRILLAR FORMS
NO STRUCTURE OF THE MONOMERIC PEPTIDE AVAILABLE IN AQUEOUS SOLVENT
PEPTIDE ADOPTS A HELICAL STRUCTURE IN APOLARORGANIC SOLVENT (SUCH AS HEXAFLUOROISOPROPANOL HFIP)
EFFECTS OF SOLVENT ON FREE ENERGY LANDSCAPE OF THE MONOMER
REPLICA EXCHANGE SIMULATION IN:
1) HFIP/WATER CO-SOLVENT 2) PURE WATER
GROMOS96 FORCE FIELD, EXPLICIT SOLVENT
40 REPLICAS, 16 NS EACH, TOTAL SIMULATION TIME OF 640 NS
FREE ENERGY SURFACE IN HFIP/WATER COSOLVENT:
HELIX STABILIZATION
T=300 K
45 %
8%
N
N
N
N
C
CC
C
HFIP DISPLACES WATER NEAR THE PEPTIDE, AND FORMS A “COAT” AROUND THE PEPTIDE
FREE ENERGY SURFACE IN PURE WATER
FORMATION OF COLLAPSED-COILS and -HAIRPINS
T=300K
N
N
N
N
N C
C
C
C
C
Possible importance of turn in toxicity of monomer
REPLICA EXCHANGE MD ON DIMERS IN WATER:HETEROGENEOUS DIMERS
TWO TYPES OF ORDERED DIMERS
Experimentally: two different protofilament of diameters:1.41 +/- 0.48 nm3.58 +/- 1.53 nm
PEPTIDE INHIBITORS OF ALZHEIMER A AGGREGATION
Alzheimer’s disease (AD) is a neurodegenerative disease of the central nervous system.
ALZHEIMER DISEASE IS CHARACTERIZED BY THEPRESENCE OF NEUROFIBRILLAR TANGLES AND
AMYLOID PLAQUES IN THE BRAIN
Aβ40: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVV
Aβ42: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVVIA
AMYLOID PLAQUES CONSIST OF AMYLOID BETA (A) PEPTIDES GENERATED FROM THE PROTEOLYTIC CLEAVAGE OF THE APP TRANSMEMBRANE PROTEIN
Amyloid Plaques
Aβ40: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVV
Aβ42: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVVIA
In healthy individuals, the A peptides are broken down and eliminated. In AD, these peptides self-assemble into amyloid fibrils
Fibril
Both small soluble oligomers and fibrils appear to be toxic to cells.
Native Protein
Misfolded Protein
Soluble Oligomer
Protofibrils
Fibril
N-methylated A(16-20)m peptides can:1) prevent the aggregation of full length A peptide 2) disassemble existing fibrils and possibly small oligomers.
Aβ40: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVV
N-METHYLATED PEPTIDE INHIBITORS
16K(me)LV(me)FF
Fibrils ofA(1-40)peptides
After IncubationwithA(16-20)mpeptides
Meredith and co-workers, J. Pep. Res. (2002) 60, 37-55
Fragment A(16-22) KLVFFAE aggregates to form fibrils
Tycko et al. Biochemistry, 39 (45), 13748 -13759, 2000
Antiparallel arrangements from solid state NMR
MODEL SYSTEM
Aβ40: DAEFRHDSGYEVHHQ16KLVFFA22EDVGSNKGAIIGLMVGGVV
A(16-22) KLVFFAE PROTOFIBRIL
INITIAL STRUCTURE:
Two parallel bilayers
Peptides in layer antiparallel
lys16 and glu22 point to solvent
leu17, phe19, ala21 point inside core
A(16-22) KLVFFAE PROTOFIBRIL
INITIAL STRUCTURE:
Two parallel bilayers Peptides in layer antiparallel
lys16 and glu22 point to solvent
leu17, phe19, ala21 point inside core
GROMOS96 FORCE FIELDEXPLICIT SPC WATER(23000 atoms)
REACTION FIELD/ PME TWO 20 NS SIMULATIONS
REPRESENTATIVE A(16-22) PROTOFIBRIL
(LAST 7 NS OF SIMULATIONS)
Distance between bilayers: 0.93 nm (Tycko: 0.99nm)Distance between peptides: 0.44-0.52 nm (Tycko: 0.47 nm)
Structure of N-methylated A(16-20)m Inhibitor Peptide
Replica Exchange Molecular Dynamics, 30 replicas,20 ns per replica
A(16-20)m more rigid than A(16-22), with -strand content:This pre-organization may allow A(16-20)m to successfullycompete with free A(16-22) for binding to fibril.
Interaction of A(16-20)m Inhibitor Peptide with protofibril
INITIAL STRUCTURE
A(16-20)m with N-methyl groups pointing away fibril
A(16-20)m with N-methyl groups pointing toward fibril
Interaction of A(16-20)m Inhibitor Peptide with protofibril
AFTER 50 ns
A(16-20)m with N-methyl groups pointing toward fibril
Intercalates betweenlayers
A(16-20)m with N-methyl groups pointing away fibril
Forms hydrogen bonds with fibril(antiparallel)
POSSIBLE MECHANISM OF FIBRIL DISRUPTION
Inhibitor drifts from edge of fibril to side and inserts in fibril (between strands 6 and 7) with Lys pointing to solvent and hydrophobic residues inserted in fibril.
POSSIBLE MECHANISM OF INHIBITION OF FIBRIL GROWTH
Inhibitor forms hydrogen bond with fibril, with antiparallelalignment, possibly preventing additional A(16-22) peptidesfrom binding.
DESTABILIZATION OF FIBRIL WHEN INHIBITOR INSERTEDIN FIBRIL
Inhibitor inserted in fibril affects the “twist” of the fibriland the distance between strands.
CONCLUSIONS AND FUTURE DIRECTIONS
1. Results suggest possible mechanisms of fibril inhibition and disassembly
2. Extend the simulations to consider other orientations of the inhibitor peptides
3. Study and design new inhibitors
ACKNOWLEDGEMENTS
Dr. P. Soto
Other group members: M. Friedel, M. Griffin, A. Jewett, W. B. Lee and E. Zhuang
Funding: NSF Career, David and Lucile Packard Foundation, A. P. Sloan Foundation, Army Research Office.
Collaborator: Prof. Stephen Meredith, University of Chicago
Dr. G. WeiDr. A. Baumketner
University of California Santa Barbara