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Thermodynamic and Kinetic Origins of Alzheimer's and Related Diseases: a Chemical Engineer's Perspective Carol K. Hall Department of Chemical & Biomolecular Engineering North Carolina State University Protein Folding: The ABCs.

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Thermodynamic and Kinetic Origins of Alzheimer's and Related Diseases: a Chemical Engineer's PerspectiveCarol K. HallDepartment of Chemical & Biomolecular Engineering North Carolina State University

protein folding the abcs

Protein Folding: The ABCs

A. A protein is a chain of amino acid residues arranged in a unique sequence.

c physiological proteins exist in the folded or native state the state with the lowest free energy
C. Physiological proteins exist in the folded or “native” state, the state with the lowest free energy.

Villin headpiece protein



(moderate T or low denaturant)

D. Proteins unfold into a “random coil” if temperature raised or denaturant (urea, GuHCl) added.

E. Of all the forces thought to govern protein folding, hydrophobicity and hydrogen bonding are considered most important.


(high T or high denaturant)






Prion disease (e.g. Mad Cow)

Amyloid Lateral Sclerosis

( Lou Gehrig’s)

Huntington’s Disease




alpha synuclein

prion protein



Amyloidoses:Diseases characterized by the abnormal aggregation of proteins into ordered structures, called “fibrils” or “amyloid.”
alzheimer s disease
Alzheimer’s Disease
  • 100 years ago --Dr. Alois Alzheimer described abnormal clumps in brain of deceased dementia patient, Auguste D.
  • Clinical symptoms: severe dementia, loss of memory & motor skills----> death
  • Late onset disease : 5-10% of 65-74 year olds,

50% of 85+ year olds

  • 4.5 million Americans
  • Costs $100 billion/year
  • US Research Budget $650 million/year.
structure of amyloid fibrils



Structure of Amyloid Fibrils

Fibrils are ordered aggregates of peptides characterized by cross-beta structure

AFM on fibrils of A-ß protein

-sheets in a


issues in amyloid disease research
Issues in Amyloid Disease Research
  • Identity of toxic species--- early oligomers or fibrils?
  • Kinetics of fibril nucleation and growth
  • Structure of fibrils
  • Interactions with inhibitors


To develop a computational tool that :

allows investigation (particularly visualization) of spontaneous fibril formation.

reveals the basic physical principles underlying fibril formation


Six Blind Men and Elephant

polyalanine a model system for studying protein fibrillization

Polyalanine– A Model System for Studying Protein Fibrillization

Speculation - fibril formation is natural consequence of peptide geometry, hydrogen-bonding capability and hydrophobic interactions under slightly-denatured, concentrated conditions.

Polyalanine peptides form fibrils in vitro at high concentrations (C > 1.5 mM) and high temperature (T > 40oC) (Blondelle et al., Biochem. 1997).

Peptide Sequence: KA14K


beta-sheets in a fibril

molecular dynamics simulations of protein folding

Molecular Dynamics Simulations of Protein Folding

Packages: Amber, CHARMm, ENCAD, ECEPP, Discover, UNRES, etc.

Force fields: describe interactions between all atoms on protein and in solvent at atomic resolution

Desired Output: “folding” trajectory of a protein

Limitation: very difficult to simulate folding of a single protein even with the fastest computers

Implications : sacrifice details to study protein aggregation

discontinuous molecular dynamics

Discontinuous Molecular Dynamics

Traditional MD:

Forces based on Lennard Jones (LJ) potential.

Follow particle trajectories by numerically integrating Newton’s 2nd law every picosecond.

Discontinuous MD:

Forces field based on square-well potential.

Follow particle trajectories by analytically integrating Newton’s 2nd law

Particles move linearly between collisons, capture or bounce

prime pr otein i ntermediate resolution m od e l





PRIME (Protein Intermediate Resolution Model):
  • United atom: NH, CaH, CO, R

R= CH3 for alanine

  • Steric Interactions:

hard spheres with realistic diameters

  • Pseudo-bonds maintain:

ideal backbone bond angles


residue L-isomerization

  • Covalent bond and pseudo-bond lengths set to ideal experimental values

Smith and Hall. PROTEINS (2001) 44 344

Nguyen et al. Protein Sci (2004) 13 2909-2924

model forces hydrogen bonding
Model Forces: Hydrogen Bonding

Hydrogen bonds between backbone amine and carbonyl groups are modeled with a directional square-well attraction of strength eH-bonding.






Square-well attraction



Define reduced temperature as: T*=kBT/εH-bonding


Model Forces: Hydrophobic Interactions

  • Solvent effects captured implicitly .
  • Hydrophobic side chains cluster together to avoid water
  • Hydrophobic interaction modeled as square-well attraction between side chains.
  • R= εhydrophobicity/εH-bonding
folding of single ka14k chain
Folding of Single KA14K Chain

Nguyen,Marchut & Hall Biophys. J (2004)


Equilibrium Simulations: 96 Peptides

Use the replica-exchange method to simulate 96-peptide systems at different temperatures and peptide concentrations.

These trends qualitatively agree with experimental data (Blondelle 1997)

Nguyen & Hall Biophys. J. (2004)


Fibril Structure: Intra-sheet Distance

Intra-sheet distance: 4.92 ± 0.01A, comparable to experimental values of 4.76A (Shinchuk et al., Proteins, 2005)


Fibril Structure: Inter-sheet Distance

Inter-sheet distance: 7.52 ± 0.23A, comparable to experimental values of 5.4A (Shinchuck et al., Proteins, 2005)


Fibril Structure: Peptide Orientation

Most peptides are in-register, same as experimental results for the A-ß(10-35) peptide (Benzinger et al., PNAS 1998)


Forming Various Structures versus t* c=5mM, T*=0.14

  • Amorphous aggregates form instantaneously, followed by ß-sheets, and then fibrils after a delay, called the lag time.
  • Appearance of a lag time indicates that this is a nucleated phenomenon.

all aggregates

Nguyen & Hall, J. Biol. Chem (2005)

fibril formation in seeded and unseeded systems at t 0 14 c 2mm

Fibril Formation in Seeded and Unseeded Systems at T*=0.14, c=2mM

Adding a seed eliminates the fibril formation lag time, as found experimentally.

in conclusion technical
In Conclusion---Technical
  • First intermediate resolution simulations of spontaneous “fibril” formation
  • Our results qualitatively agree with experimental data in general, and specifically with those obtained by Blondelle et al. (Biochemistry, 1997) on polyalanines.
  • Next step: Extending PRIME to all 20 amino acids. Which road to take?????
  • Dr. Hung D. Nguyen
  • Dr. Alexander J. Marchut
  • Dr. Anne V. Smith
  • Dr. Hyunbum Jang
  • Dr. Andrew J. Schultz
  • National Institutes of Health
  • National Science Foundation