Protein folding and misfolding
UNFOLDED PROTEINS
NATIVE FOLDED
PROTEINS
AMYLOID PROTEIN
1
ll living organisms use proteins for structure, energy production, information
processing, etc. Finished proteins have complicated three-dimensional
structures that are necessary for their diverse activities. However, proteins
are first produced as a linear string of amino acids, and they must then fold
into the right conformations in the crowded environment of a cell (~300mg/
ml). Diseases can ensue when proteins have problems reaching their
functional conformations: sickle cell anemia, cystic fibrosis, Alzheimer’s etc.
3
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Levinthal paradox
Assume each amino acid backbone can be in 3
conformational states, for 101 residues, there
are 3100 = 5 x 1047 conformations.
If the protein can sample a new conformation at
a rate of 1013 s-1, it will take 1027 years to try
them all. Longer than the age of the universe!
Therefore, proteins must fold in “pre-arranged
pathways” and in a cooperative manner.
Levinthal C. Extrait du Journal de Chimie
Physique 1968; 65:44 Zwanzig et al., PNAS 1992;
89:20-22
Protein folding

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Native state corresponds to structure that is most stable under
physiological conditions
Assumption that native like interactions are on average more
stable than non-native ones—search mechanism is able to find
native state.
Mechanism is efficient in that only a small fraction of the
conformations need to be sampled during folding
6
Folding Funnel
Methods. 2004,34(1):4-14.
A schematic energy landscape for protein
folding. The surface shown here is derived from
a computer simulation of the folding of a highly
simplified model of a small protein. Such a
surface serves to “funnel” the multitude of
denatured conformations to the unique native
structure. The critical region on a simple surface,
such as this one is the saddle point
corresponding to the transition state, the barrier
that all molecules must cross to be able to fold to
the native state. An ensemble of structures
corresponding to the experimental transition
state for the folding of a small protein is
indicated in the figure; this ensemble was
calculated by using computer simulations
constrained by experimental data from
mutational studies of acylphosphatase. The
yellow spheres represent the three “key
residues” in the structure; when these residues
have formed their native-like contacts, the
overall topology of the native fold is established.
The structure of the native state is shown at the
bottom of the surface, while at the top are
indicated schematically some contributors to the
distribution of unfolded states that represent the
starting point for folding. Also indicated on the
surface are highly simplified average trajectories
for the folding of individual molecules.
7
Protein misfolding and aggregation
IDP
Transient dimer
protofibrils
Amyloid fibril
Example: α-synuclein (Parkinson’s),
IAPP (Diabetes), Aβ(Alzhermer’s),
polyQ (Huntington’s)
8
Aggregation of α-synuclein is related to
Parkinson’s Disease
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Parkinson’s disease (PD) is the second most
prevalent neurodegenerative disease in the
world.
Aggregated α-synuclein (aSyn) is found in the
Lewy body or Lewy neurites of brain cells
which is the hallmark of PD.
Lewy body
Goedert M, Nat Rev Neurosci.2001,2(7):492-501.
aSyn forms amyloid fibrils in vivo and in vitro.
Conway KA, Biochemistry.2000,;39(10):2552-63.
9
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Alzheimer afflicts 10 percent of those over 65 years old and
perhaps half of those over 85. Every year Alzheimer’s not only
kills 100,000 Americans, but also costs society $82.7 billion to
care for its victims.In 1991, several different re- search groups
found that individuals with specific mutations in their amyloid
precursor protein developed Alzheimer’s disease as early as age
40. The body processes amyloid precursor protein into a soluble
peptide (small protein) known as Aβ; under certain
circumstances, Aβ then aggregates into long filaments that
cannot be cleared by the body’s usual scavenger mechanisms.
These aggregates then form the β-amyloid, which make up the
neuritic plaque inAlzheimer patientspeptide (small protein) known
as Aβ; under certain circumstances, Aβ then aggregates into
long filaments that cannot be cleared by the body’s usual
scavenger mechanisms. These aggregates then form the βamyloid, which make up the neuritic plaque inAlzheimer patients.
TEM shows that all variants
make fibrils
T53A
mouse
M100L,G103N,Y107A
N87S
G121D, S122N
T53A,N87S
human
Scale bar: 200 nm
11
Protein folding in the cell

Cellular environment is very complex—concentration is 350mg/
ml.

To avoid aggregation auxilliary proteins exist to assist the
proteins to fold efficiently and without aggregation
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Folding catalysts to enhance slow steps in folding and molecular
chaperones to avoid protein misfolding

Sophisticated mechanisms of quality control to check whether
proteins are correctly folded and to destroy proteins that are not
good.
12
Regulation of protein folding in the ER
Many newly synthesized proteins are
translocated into the ER, where they fold
into their three-dimensional structures
with the help of a series of molecular
chaperones and folding catalysts (not
shown). Correctly folded proteins are
then transported to the Golgi complex
and then delivered to the extracellular
environment. However, incorrectly folded
proteins are detected by a quality-control
mechanism and sent along another
pathway (the unfolded protein response)
in which they are ubiquitinated and then
degraded in the cytoplasm by
proteasomes.
Nature. 2003,426:884-890.
13
Folding in the cell

Proteins are synthesized on ribosomes in cellular DNA. Folding occurs in the
cytoplasm after release from the ribosome and in other folding specific
compartments such as ER. Incompletely folded proteins expose to solvent
some regions that are buried in the native state and prone to inappropriate
interactions with other molecules in the crowded cellular environment. Range of
strategies to prevent such behavior

Molecular chaperones either interact with protein on ribosome or guide later
stages of folding—chaperones work by reducing the probability of competing
reactions such as aggregation

After folding in the ER secretion to the golgi apparatus. In the ER proteins must
undergo quality control check—if they do not pass they are targeted for
degradation

Incorrect folding—cystic fibrosis and cancer—cannot exercise function. Other
proteins with high propensity to misfold form aggregates such as Alzheimers,
type II diabetes and BSE or vCJD.
14
Protein misfolding

Misfolding is defined as reaching a state that has a
significant proportion of nonnative interactions between
residues
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Proteins important in biological processes—failure to fold
correctly—gives rise to disease

Misfolded proteins do not exercise proper function or
form deposits in the cells.

Not only “loss of function” but “toxic gain of function” as
aggregates induce cell damage or cell death
15
Protein folding and disease

Mistakes in folding will give rise to malfunctioning of biological
processes and to disease.

Example cystic fibrosis—protein does not fold due to familial
mutation and results in reduced level or reduction of protein.

Amyloid diseases—failure to fold results in aggregation and
deposits of protein in one or more tissues.
16
Aggregation diseases
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Give rise to deposition of proteins in the form of amyloid fibrils
and plaques
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Deposits in brain, organs such a liver and spleen

Fibrillar structures have similar morphology
17
Amyloid diseases
Semin Cell Dev Biol. 2004,15(1):3-16.
18
Amyloid fibrils

Amyloid fibrils appear in a class of diseases called amyloid
diseases such as Alzheimer’s, Parkinson’s, Huntington’s
disease.

Point of contention is whether the most important toxic species is
the amyloid fibril or a smaller aggregated form of the same
protein or peptide.

Amyloid fibril formation is not a property that is restricted to
proteins associated with disease.

Amyloid fibril appears to be a stable structural state of a
polypeptide competing thermodynamically and kinetically with
globular monomeric states and unfolded monomeric states.
19
Mechanism of
protein aggregation
Figure 2 A schematic representation of some of the
many conformational states that can be adopted by
polypeptide chains and of the means by which they
can be interconverted. The transition from β-structured
aggregates to amyloid fibrils can occur by addition of
either monomers or protofibrils (depending on protein)
to preformed β-aggregates. All of these different
conformational states and their interconversions are
carefully regulated in the biological environment, much
as enzymes regulate all the chemistry in cells, by
using machinery such as molecular chaperones,
degradatory systems, and quality control processes.
Many of the various states of proteins are utilized
functionally by biology, including unfolded proteins and
amyloid fibrils, but conformational diseases will occur
when such regulatory systems fail, just as metabolic
diseases occur when the regulation of chemical
processes becomes impaired.
20
Overview of possible fates of newly
synthesized chain
J Mol Med. 2003;81(11):
678-99.
21
Fate of polypeptide chain

Equilibrium 1 is affected by mutations,
misprocessing, aberrant interactions with metals,
changes in environmental conditions. This
increases the population of partially folded or
unfolded species—species are more aggregation
prone than native state.

Equilibrium 2 affected by mutations so as to
kinetically favor aggregate nucleation
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Reaction 3 is irreversible and mature fibrils are the
final stable product if aggregation process.
22
Common features in self assembly
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First phase involves the formation of oligomeric species as a
result of non specific interactions.
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Smaller oligomers are soluble and earliest species observable
by AFM are bead like structures.
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Prefibrillar aggregates then transform into protofibrils.
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Self assemble into mature fibrils.
23
J Mol Med. 2003;81(11):678-99.
Fig. 4. Some amyloid-related peptides/proteins form early aggregates of globular appearance
that further organise into beaded chains, globular annular 'doughnut' shaped assemblies
eventually giving mature protofilaments and fibrils. Pre-fibrilar aggregates may interact with
reconstituted phospholipid membranes and with cell membranes where they form aspecific
channels (pores) disrupting cellular homeostasis. The latter possible mechanism of toxicity is
similar to that displayed by antimicrobial peptides, pore-forming eukaryotic proteins and bacterial
toxins and newly synthesised cyclic peptide antibiotics (see text). The electron micrographs of
the globular and beaded chains of Aβ peptides are taken from Harper et al. [200]. The electron
micrographs of the rings of the α-synuclein A53T (upper row) and A30P (middle row) mutants
and of the Alzheimer precursor protein artic mutant (lower row) are from [201]
24
Characterization of amyloid
structures
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Amyloid deposits all show characteristic optical behavior, green
birefringence on binding to certain dye molecules such as Congo
red because of the regularly spaced β-sheets
β-structure is seen by CD and FT-IR

EM—long and unbranched; fibrils are usually 10 nm in diameter
and consist of between 2 to 6 protofibrils twisted around each
other.

X-ray fibre diffraction organized core has a cross-β structure
where sheets are assembled from b-strands that run
perpendicular to fibrillar axis
25
Biochimie. 2004;86(9-10):589-600
26
Schematic representation of the mechanism of
formation of amyloid fibrils and the various
types of antibodies that can be produced. The
first step in the process of formation of an
amyloid fibril (black box) is the formation of an
amyloidogenic intermediate via the partial
unfolding of the native state (arrow 1) or via
partial folding of otherwise naturally unfolded
species (arrow 2). The second step is the selfassociation of the amyloidogenic intermediates,
which eventually leads to the formation of
amyloid fibrils (red box). The amyloidogenic
intermediates have a high tendency to
aggregate and become stabilised by the
formation of intermolecular β-sheets. Small
oligomers are formed initially that act as nuclei
to direct the further growth of aggregates
(*Here the nucleus is for simplicity shown as a
dimer). This growth leads to the formation of
higher order oligomers referred to as prefibrillar aggregates (PA, sometimes also
referred to as amorphous aggregates, micelles
or protofibrils). These aggregates convert into
protofilaments (P) directly or indirectly, and
finally into mature fibrils (F). Such fibrils usually
consist of two to six protofilaments that are
often twisted around each other to form a ropelike structure as shown in Box 1c. (**According
to this scheme, antibodies 3 and 5 also bind to
PA species but they are not represented for
clarity).
Biochimie. 2004;86(9-10):589-600
27
Fig. 3. A molecular model of an amyloid fibril. This model is derived
from cryo-EM analysis of fibrils grown from an SH3 domain. The
fibril consists of four “protofilaments” that twist around one another
to form a hollow tube with a diameter of approximately 6 nm. The
model illustrates one way in which regions of the polypeptide chain
involved in β-sheet formation could be assembled within the fibrils.
Semin Cell Dev Biol. 2004,15(1):3-16.
28
Q Rev Biophys. 2006;39(1):1-55
Fig. 15. Structural model for the protofilament in Aβ1–40
fibrils prepared with gentle agitation (Petkova et al. 2006),
exhibiting the morphologies and NMR signals shown in
Figs. 4, 8, 10a, 12–14. Residues 1–8 are
conformationally disordered and are therefore omitted.
The long axis of the fibril is perpendicular to the page in
panels (a)–(c). The long axis is vertical and parallel to the
page in panel (d). (a) All-atom representation of a pair of
peptide molecules. Residues 10–22 and 30–40 have βstrand conformations, forming two separate in-register,
parallel β-sheets. The protofilament is a four-layered βsheet structure with C2 symmetry about its long axis.
Blue double-headed arrows indicate side-chain–sidechain and side-chain–backbone contacts established by
2D 13C–13C NMR measurements as in Fig. 12. Purple
double-headed arrows indicate contacts established by
measurements of 15N–13C dipole–dipole couplings, as in
Fig. 16. (b) Average structure resulting from 10
independent molecular dynamics/energy minimization
runs on a cluster of 12 peptide molecules, with
interatomic distance and backbone torsion angle
restraints dictated by solid-state NMR data. Hydrophobic,
polar, negatively charged, and positively charged sidechains are colored green, purple, red, and blue
respectively. The four-layered β-sheet structure is
stabilized primarily by hydrophobic interactions in the
core of the protofilament. Polar and charged side-chains
are on the exterior, with the exception of oppositely
charged K28 and D23 side-chains, which form salt
bridges. (c, d) Cartoon representations, with residues 12–
21 in red and residues 30–40 in blue. A left-handed twist
of 0·833°/Å is imposed, although direct experimental
constraints on the twist in agitated Aβ1–40 protofilaments
29
are not yet available.
Figure 5 (a) Insulin structure showing the
three native disulfide bonds. A chain,
green; B chain, blue; disulfide bonds, gold.
(b) Topology diagram of insulin color
coded as in a. (c) Possible topology for the
amyloid protofilament. Orientations of the
termini and disulfide bonds within the
curved structure are arbitrary. The C
terminus of chain B (dashed) is not
required for amyloid fibril formation (see
ref. 43). (d) β-strand model of a
protofilament. Each chain is shown in two
segments, a straight and a curved β-strand
(PDB accession no. 1umu, residues 93–
100). Each insulin molecule would occupy
two layers, connected by the interchain
disulfide bonds. (e) A possible β-strand
model docked into the EM density of the
compact fibril (transparent gray surface).
The four protofilaments are colored
separately.
PNAS. 2002;99(14):9196-201
30
Figure 6 Models for protofilament packing.
(a) A twisted pair of rectangular
protofilaments in which an interactive
surface is colored purple. The protofilament
twist accompanies the filament twist. (b) A
supercoiled pair of protofilaments in which
the regions involved in packing interactions
rotate around each protofilament. In the
correlated twist model a, interacting regions
would be fixed relative to the cross-β
structure, and other regions could
accommodate large loops and/or folded
domains that would not interfere with
protofilament packing. Similar models can
be constructed with more than two
protofilaments, in which the cross section
rotates as a rigid unit in the helical
structure. Keeping the cross section fixed
means that all packing contacts can be
preserved in the helical fibril. This is the
case for the four-protofilament model in Fig.
5 e.
PNAS. 2002;99(14):9196-201
31
Figure 1 Recent three-dimensional structural
models of fibrillar aggregates from different
sources. (a) The protofilament of Aβ viewed down
the long axis of the fibril. The segments 12–24
(red) and 30–40 (blue) are shown. (b) The fibril
from the C-terminal domain 218–289 of the fungal
prion protein HET-s The ribbon diagram shows
the four β-strands (orange) (residues 226–234,
237–245, 262–270, and 273–282) and the long
loop between β2 and β3 from one molecule.
Flanking molecules along the fibril axis (gray) are
shown. (c) Atomic structure of the microcrystals
assembled from the GNNQQNY peptide. Each βstrand is a peptide molecule. (d) The
protofilament from amylin. Green, yellow, and
pink β-strands indicate residues 12–17, 22–27,
and 31–37, respectively. The unstructured Nterminal tail is shown on the right of the panel
along with the disulfide bridge between Cys2 and
Cys7. (e) The fibril from the NM region of Sup35p.
The colored ribbons indicate residues 25–38
(red), 39–90 (blue), and 91–106 (green). The
unstructured regions 1–20 (red dashed lines) and
158–250 (black dashed lines) are shown.
Annu Rev Biochem. 2006;75:333-66.
32

In native state polypeptide chain is buried. Conditions that favor
aggregation from folded proteins are those that stimulate at least partial
unfolding for example low pH or elevated temperature.

Mutations that destabilize the native state are commonly involved in
familial forms of amyloid disease.

Fragmentation of proteins

Formation characterized by a lag phase followed by a period of rapid
growth
33
Determinants of protein aggregation

Thought at first that only certain proteins could form fibrils.

Ability to form amyloid fibrils is generic feature of polypeptide chain.

Fibrils formed by many proteins that are not associated with disease.

Amyloid fibrils are organized and adopted by unfolded chain when it
behaves like a polymer.
34
Arch Biochem Biophys. 2008;469(1):100-17
Representative structures
of proteins involved in
disease-related amyloid
fibril formation
Fig. 6. Representative structures of proteins
involved in disease-related amyloid fibril
formation. The polypeptides are coloured
according to the aggregation tendency of
their amino acid sequences predicted using
the algorithm TANGO. Sequences shown in
blue are predicted to have no β-aggregation
propensity, while polypeptide stretches
coloured in yellow, orange and red indicate
an increasing propensity to aggregate.
Notably, the peptide structures were
obtained in the presence of fluoroalcohols
(calcitonin and Aβ1–42) or SDS micelles
(amylin), and these sequences might be
substantially less ordered in the absence of
these additives. Note also that for insulin,
amylin and calcitonin, the pro-peptides as
well as the mature sequences have been
implicated as potentially amyloidogenic.
35

Although the fundamental cross-β structure is common to all amyloid
fibrils, the packing of β-sheets into protofilaments must vary to some
extent according to the constraints of the constituent polypeptide.

Although different amyloid protofilaments appear similar in size, the
protein sequence can influence the sheet packing into protofilaments
36
Amyloid structure
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NO complete structure has been determined
Core structure is stabilized by hydrogen bonds involving the main
chain.
Explains why fibrils formed from different amino acid sequences
are similar in appearance.
Side chains are incorporated in manner most favorable for given
sequence
Proportion of polypeptide chain that is incorporated in the core
structure varies substantially—sometimes very few residues
Very different from globular proteins where highly specific
packing of side chains may override main chain preferences
37
Questions about fibrils

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Are cross β structures comprised of parallel β-sheets, antiparallel
β-sheets or mixed β-sheets?
Are β-sheets structurally ordered or disordered?
Which segments of the protein sequence participate in the βsheets?
Do amyloid fibrils really contain multiple layers of β-sheets and
how are these layers arranged relative to each other?
Which aspects of amyloid structure are universal and which are
sequence dependent?
What are intermolecular and intramolecular interactions that
stabilize amyloid structures apart from the backbone H-bonds?
38
Generic nature of amyloid formation

Ability to form amyloid structures appears generic
but propensity can vary dramatically between
sequences

Mutation of single aa in 100 residue protein changes
rate at which aggregation occurs

Change in rate can be correlated with changes in
properties such as charge, secondary structure
propensity and hydrophobicity (Chiti)
39
Importance of hydrophobicity, net charge,
and secondary structure propensity

Hydrophobic interactions—important driving forces in aggregation:
experiments show that residues promoting aggregation are not spread
all over the sequence—instead clustered within narrow range of
sequence. Different from protein folding where distant residues may
nucleate folding
40
Charge in protein aggregation

Electrostatic effects important in modulating aggregation

Net charge is a major determinant of aggregation
Unfolded protein aggregates more rapidly when numbers of positively
and negatively charged aa are equal and resulting net charge is zero

41
Importance of propensity to form
secondary structure in aggregation


Sequences with high propensity to form β-structures are highly
amyloidogenic
Hecht and co-workers designed combinatorial library of sequences that
shared identical patterns of regularly alternating polar and non-polar
residues ideal for β-sheet formation. Alternating periodicity is
interrupted by lysine residues at positions originally occupied by nonpolar aa-- amyloid formation was interrupted and sequences folded into
monomeric β-sheets.
42
Amino acid sequences have evolved to take into
account the influence of hydrophobicity, charge
and β-sheet propensity

Has an evolutionary pressure existed to select against protein
sequences with a high propensity to aggregate?

Folding is a strategy to escape aggregation
Hydrophobics are buried
Amide and carbonyl groups engaged in H-bonding
Charges are exposed—promote repulsive action

Folded proteins are susceptible to aggregation
Transiently adopt unfolded or partially folded conformation

Unfolded proteins: lower hydrophobic content and higher net
charge contributes to maintain aggregation propensity low.
43
Prevention of amyloid like aggregation as a driving force of
protein evolution (Montsellier and Chiti EMBO reports)

Folding is a primary strategy to prevent aggregation

Negative design to control the assembly of folded proteins:
Richardsdon and Richardson have shwon that proteins have
evolved structural adaptations to protect the peripheral b strands—
thay rae protected by structural strategies such as covering them
with a loop or a helic, formation of continuous sheet to create a beta
barrel, distrotionsof b structure…

Conservation of gly and pro residues —proline have structural
constraints that make it difficult to adapt the into a beta-structure.
Gly have a high level of conformatioanl flexibility and high entropic
cost to being incorporated into secondary structure
44

Use of gatekeeper residues to control aggregation: it was found
that positions flanking aggregating stretches are enriched with
residues such as proline, lysine, arginine, glutamate and aspartate
—pro Is b-breaker and other 4 are at the bottom of the aggregation
propensity scales-90% of 26,000 aggregating sequences found by
TANGO have at least 1 of the 5 residues at the 1st position on either
side of the segment.
45
Strategies for therapeutic intervention

Distinct steps in the aggregation process where intervention might be
able to prevent or reverse the formation of aggregates

Destabilization of native state by genetic mutation—stabilizing native
states of disease associated variants; use small molecule analogs:
Example thyroxine a natural ligand of transthyretin blocks that rate at
which transthyretin aggregates.

Aggregation due to fragments: peptide fragments result from natural
processing or incomplete degradation of full length protein and cannot
fold without the rest of the polypeptide chain. Solution: block the
enzymes that generate the fragments. Alzheimers comes from 40 or 42
residue peptide that is derived from the amyloid precursor protein (APP)
–secretase enzymes process APP and secretase inhibitors are under
development to treat Alzheimers.
46
Therapeutic approaches

Enhance clearance of aggregation prone species. Active
immunization with Aβ peptides results in clearance of amyloid
deposits in transgenic mice or a ligand that binds to serum
amyloid P (SAP) a protein that associates with amyloid deposits
and blocks natural clearance mechanisms reduces SAP levels

Alternative is to intervene in the aggregation process directly by
small peptides or peptide analogs designed to bind to fibrils—
problem is that inhibition of amyloid fibril formation might occur at
the expense of the small aggregate precursors. This may be
problem as small aggregates may be the primary pathogenic
agents in neurodegenerative diseases.

Because of the generic nature of the amyloid formation perhaps
one drug can block a number of diseases
47
Strategies for therapeutic intervention
Therapeutic intervention in
amyloid diseases. The
conversion of normal soluble
peptides and proteins into
insoluble aggregates that are
deposited in a variety of tissues is
shown (5). Highlighted are stages
in the aggregation process where
therapeutic intervention may be
able to prevent or reverse
aggregation. Therapeutic
strategies include (A) stabilizing
the native state; (B) inhibiting
enzymes that process proteins
into peptides with a propensity to
aggregate; (C) altering protein
synthesis; (D) stimulating
clearance of misfolded proteins,
for example, by boosting their
proteasomal degradation; (E)
inhibiting fibril assembly; (F)
preventing accumulation of fibril
precursors.
Science (2004):304.1259-1262
48
Folding vs. aggregation: kinetic
partitioning



Amyloid fibrils are formed in a nucleation dependent manner—
protein monomer is converted into a fibrillar structure via
transient aggregation
Residues key to aggregation are thought to be different from
those that drive the folding of polypeptide chain—although the
major driving forces, hydrogen bond formation and burial of
hydrophobic surface area are the same for both processes
Although a large part of the peptide is involved in fibril structure,
some aa sequences are more prone to aggregation than others
—this may be similar to the fact that only a few residues define
the folding nucleus in protein folding.
49
Folding vs. aggregation
Ability of proteins to fold rapidly to their globular native structure
allows them to escape aberrant side reactions that would give
access to the aggregation funnel and lead to thermodynamic
ground state of intermolecular assembly, the amyloid fibril
Evolution has shaped the folding and aggregation funnels to allow
trapping of the native functional state which is thermodynamically
a metastable structure in the context of the entire protein
landscape.
Proteins are dynamic and have transient partial unfolding events.
Native proteins are marginally stable compared to denatured
proteins and partially folded states can be formed from the folded
structure by local and sub-global unfolding events. However,
cooperativity of folding and cellular rescue machinery help to
avoid populations of partially folded forms.
50
Folding vs aggregation

Changes in aa sequence, alterations in the folding conditions,
breakdown of the cellular control system allows the shift towards
aggregation funnel, where polypeptide chain folds into the
thermodynamically stable fibril conformation.

Question: at which points do folding and aggregation landscapes meet?
Does the separation between the fates occur at the unfolded state or
are partially folded forms a common entity?

Questions about the nature and frequency of different unfolding events,
structural properties of different ensembles, barrier heights between
them are still to be defined.
51
References
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Brockwell DJ, Smith DA, Radford SE.
Protein folding mechanisms: new methods and emerging ideas.
Curr Opin Struct Biol. 2000 Feb;10(1):16-25.
Dobson CM.
The structural basis of protein folding and its links with human
disease.
Philos Trans R Soc Lond B Biol Sci. 2001 Feb 28;356(1406):133-45.
Petkova AT, Ishii Y, Balbach JJ, Antzutkin ON, Leapman RD, Delaglio
F, Tycko R.
A structural model for Alzheimer's beta-amyloid fibrils based on
experimental constraints from solid state NMR.
Proc Natl Acad Sci U S A. 2002 Dec 24;99(26):16742-7.
Chiti F, Stefani M, Taddei N, Ramponi G, Dobson CM.
Rationalization of the effects of mutations on peptide and protein
aggregation rates.
Nature. 2003 Aug 14;424(6950):805-8.
Stefani M, Dobson CM.
Protein aggregation and aggregate toxicity: new insights into
protein folding, misfolding diseases and biological evolution.
J Mol Med. 2003 Nov;81(11):678-99.
Dobson CM.
Protein folding and misfolding.
Nature. 2003 Dec 18;426(6968):884-90.
Dobson CM.
Principles of protein folding, misfolding and aggregation.
Semin Cell Dev Biol. 2004 Feb;15(1):3-16.
Dobson CM.
Protein chemistry. In the footsteps of alchemists.
Science. 2004 May 28;304(5675):1259-62.
Dobson CM.
Experimental investigation of protein folding and misfolding.
Methods. 2004 Sep;34(1):4-14.
Dumoulin M, Dobson CM.
Probing the origins, diagnosis and treatment of amyloid diseases
using antibodies.
Biochimie. 2004 Sep-Oct;86(9-10):589-600.






Pawar AP, Dubay KF, Zurdo J, Chiti F, Vendruscolo M, Dobson CM.
Prediction of "aggregation-prone" and "aggregationsusceptible" regions in proteins associated with
neurodegenerative diseases.
J Mol Biol. 2005 Jul 8;350(2):379-92.
Jahn TR, Radford SE.
The Yin and Yang of protein folding.
FEBS J. 2005 Dec;272(23):5962-70.
Chiti F, Dobson CM.
Protein misfolding, functional amyloid, and human disease.
Annu Rev Biochem. 2006;75:333-66.
Bemporad F, Calloni G, Campioni S, Plakoutsi G, Taddei N, Chiti F.
Sequence and structural determinants of amyloid fibril
formation.
Acc Chem Res. 2006 Sep;39(9):620-7
Tycko R.
Molecular structure of amyloid fibrils: insights from solid-state
NMR.
Q Rev Biophys. 2006 Feb;39(1):1-55.
Monsellier E, Chiti F.
Prevention of amyloid-like aggregation as a driving force
of protein evolution.
EMBO Rep. 2007 Aug;8(8):737-42.

Jahn TR, Radford SE.
Folding versus aggregation: polypeptide conformations on
competing pathways.
Arch Biochem Biophys. 2008 Jan 1;469(1):100-17.

Vilar M, Chou HT, Lührs T, Maji SK, Riek-Loher D, Verel R,
Manning G, Stahlberg H, Riek R.
The fold of alpha-synuclein fibrils.
PNAS. 2008 Jun 24;105(25):8637-42.
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Protein Misfolding, Aggregation and Conformational Diseases: Part
A: Protein Aggregation and Conformational Diseases
Chapter 2. The generic nature of protein folding and misfolding
Edited by Anthony Fink and Vladimir Uversky
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