Transcript Slide 1

Protein Toxicity in Parkinson
And Alzheimer’s Disease
Chuck Sanders, Dept. of Biochemistry
Rm 5110C MRBIII, [email protected]
Readings on Role of α-Synuclein in Parkinson Disease
100 years of Lewy pathology.
Goedert M, Spillantini MG, Del Tredici K, Braak H.
Nat Rev Neurol. 2013 Jan;9(1):13-24.
α-Synuclein: membrane interactions and toxicity in Parkinson's disease.
Auluck PK, Caraveo G, Lindquist S.
Annu Rev Cell Dev Biol. 2010;26:211-33.
Pathological roles of α-synuclein in neurological disorders.
Vekrellis K, Xilouri M, Emmanouilidou E, Rideout HJ, Stefanis L.
Lancet Neurol. 2011 Nov;10(11):1015-25
α-Synuclein oligomers and clinical implications for Parkinson disease.
Kalia LV, Kalia SK, McLean PJ, Lozano AM, Lang AE.
Ann Neurol. 2012 Aug 28. doi: 10.1002/ana.23746.
PD
AD
From: May 2010 US National Vital Statistics Report
Protein Deposition in Neurodegenerative Diseases: A Histological View
A. Abbott Nature Outlook 2011
NEJM 349, 583 (2003)
note that list is
incomplete and growing
Key Questions About Protein Aggregation in Alzheimer’s and Parkinson Diseases
What triggers protein aggregation?
In inherited form of disease?
In sporadic (idopathic) form of disease?
What form of the protein is toxic?
Disease promoted by toxic “gain of function”, loss of native function, or both?
What organ, tissue, and cells are affected… as a function of time?
Where in (or out) of cell does protein aggregate, deposit and exert its toxicity?
What is the mechanism of toxicity and is it direct or pathway-based?
Does protein deposition spread from cell to cell and tissue to tissue? If so, how?
Before we explore these issues, we need to review the biophysics and biochemistry
of protein folding and misfolding.
Thermodynamic Stability of Proteins
folded
unfolded
C
Keq,fold
Keq,fold=
[folded]
[unfolded]
o
G

N
Standard Free Energies of Folding for Single Domain Proteins
Protein
Gofold (kcal/mol)
lambda repressor
alpha spectrin SH3 domain
arc repressor
cytochrome C (with heme and Fe(II)
CD2
procarboxypeptidase
U1A spliceosomal protein
Hpr
-3.0
-2.9
-6.3
-15
-8.2
-4.1
-9.3
-4.6
Most folding proteins are only moderately stable:
The folded state is usually favored over the unfolded state by only -3 to -10 kcal/mol.
Many Proteins are Natively Unfolded (Natively Disordered)
The unfolded state is energetically favored, unless some the protein comes in contact
with something that stabilizes the folded state (e.g., a cognate ligand or a membrane).
Polypeptides <50 Residues Usually Do Not Fold In Solution
Unless They Bind to Another Protein or to A Membrane
Protein Folding Pathways: Kinetics Revisited
o
G
Rate depends on the pathway between states and associated G*.
Favorable Go doesn’t mean the process will take place at a significant rate
“Kinetically trapped”
Proteins fold because it is both thermodynamically and kinetically
favorable for them to do so.
2 states
1 barrier in each direction
G*forward G*reverse
Kinetics
G
o
G* : free energy of activation
rate is proportional to rate constant, k,
which is proportional to:
G*
e RT
There is an inverse logrithmic relationship
between rate and activation energy.
Mutations can perturb the heights of the rate
barriers and/or the energies of the free energy
states. Consider a mutation that lowers the
energy of a folding intermediate:
Examples of Potentially
Deadly Mutations
Barrier to folding becomes too
high.
Normal barrier to misfolding is
reduced.
unfolded
G
o
Mutation that lowers
energy of folding
intermediate (only).
folded Folding intermediate is
stabilized.
Folded form is destabilized.
Disrupt interactions with the
protein folding machinery.
o
G
Note that this mutation not only makes the intermediate
much more stable (longer-lived), but that the energy
barriers from this state to both initial and final states
are now larger than for wild type.
Protein Aggregation
When protein sticks to protein to form very large oligomers
Both folded and unfolded proteins can sometimes aggregate.
Aggregates can be disordered (amorphous) or can be ordered.
Amyloid fibrils are a common type of ordered aggregate.
The rate of formation of ordered aggregates is usually limited by
the rate of a requisite nucleation step.
The Formation of Ordered Aggregates Is Rate-Limited By A Nucleation Event
This could involve the rare collision of minor conformations of the protein or could depend on
the appearance of post-translationally modified form of a protein that can serve as an
aggregation seed. The rareness of the seeding species and events can make it very hard to
pin down the exact nature of the nucleation event, especially in living tissue.
Figure: Aguzzi and Polymenidou, Cell, 2004
Selkoe, Nature, 2003
cross-β fibril formation on the
road to mature amyloid; initial
self-association might be
staged from native or
denatured states in some
instances.
Protein Folding in the Cell
The main locations of protein folding in the cell
are the cytosol and the lumen of endoplasmic
reticulum.
ER: Membrane proteins, secreted proteins, and
organellar proteins.
Protein folding in the cell…
3-D structure is determined by sequence.
Folding pathways differ from test tube:
Folding is often co-translational.
Chaperone proteins play roles.
Other folding accessory proteins.
Folding is monitored by quality control.
Folding-defective protein is degraded.
There are two major degradation protein pathways in mammalian cells.
The ubiquitination/proteasome pathway.
The autophagy/lysosomal pathway(s).
Members of Alois Alzheimer’s research group
at the Royal Psychiatric Clinic of the University Alois
of Munich, Germany in 1910
Alzheimer
Goedert, M. et al
Nat Rev Neurol 9, 13-24, 2013
Fritz Jakob
Heinrich Lewey
http://coloradodementia.org
Marwan Sabbagh
Extracellular Amyloid Plaques Are One of the Hallmarks of AD;
Central Component: The Amyloid-β Polypeptide
A Second Hallmark of Alzheimer’s Disease Are Intracellular Deposits Called
“Neurofibrillary Tangles”.
The Central Component of NFTs is the Hyperphosphorylated Tau Protiein
Intracellular Lewey Bodies Are the Hallmark Parkinson Disease
Central Component: α-Synuclein
The superstructure of amyloid-β in
(i) amyloid fibrils in amyloid plaques
(ii) hyperphosphorylated tau in neurofibrillar tangles,
(iii) α-synuclein in Lewey bodies
is thought to be organized around
“amyloid” cross-beta fibrils formed
by each of these three proteins.
The Amyloidogenic Pathway and Alzheimer’s Disease
cytosol
Amyloid Precursor
Protein
secretase
cleavage
Amyloid-Beta
cytosol
C99
Oligomers
secretase
cleavage
Neuron
Cell Death
The Amyloid Precursor Proteins and
The Amyloid-β Polypeptides
Long forms Aβ42 and Aβ43 thought to be more toxic than short forms Aβ38 and Aβ40
Retaining half of the TM segment of the APP, the Aβ polypeptides (especially) the
long forms are only marginally water-soluble. They have a high propensity to
aggregate and may also retain significant affinity for membrane surfaces.
The function of APP and of its fragments (extracellular domain: β-APP,
Aβ, and APP intracellular domain:AICD) are not well understood. Lots of bits
and pieces of evidence in support of various roles. However, APP-knockout mice
are a little slow and a little stupid, but live. Lethality occurs only when two
homologs of APP– the APP-like proteins (ALP1 and ALP2) are also knocked out.
This implies that AD is not strongly related to the loss of native function of APP or
of its derived fragments.
Schematic of the protein sequence of human α-synuclein.
Bosco D A et al. Cold Spring Harb Perspect Biol
2011;3:a007500
©2011 by Cold Spring Harbor Laboratory Press
Natively disordered in solution.
Binds to membrane surface via N-terminal
domain, which is thought to form helices.
C-terminal domain remains disordered, but
engages in complexes with other proteins.
Functions of α-Synuclein
α-synuclein stabilizes synaptic vesicles (and other vesicles such as the ER-to-Golgi
transport vesicles) to prevent premature membrane fusion and loss of vesicle
contents. However, knockout of α-synclein is not fatal and does not lead to
neurodegeneration. Parkinson’s does not seem to be primarily loss of function in nature.
Annual Reviews