Protein Structures: Data Representation

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• Protein Structures Data Representation
• Primary Structure character string.
• Secondary Structure
• Tertiary Structure
• Quaternary Structure

Identifying sub-structures in a large protein
based on sequence. 3-Dimensional
Representation Protein Database Bank (PDB) This
is a complicated file format structure that
support numerous programs, and contains
information regarding the primary structure
(sequence), 3-D structures (x, y, z coordinates),
size and linking of specific atoms in structures,
etc.
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Secondary Structure Prediction 1) Hydropathy
Plot 2) Alpha Helix 3) Beta Sheet
A Hydropathy plot identifies domains within a
protein that are soluble (region of charged
amino acids) or insoluble (region of uncharged
amino acids).
An alpha helix is a group of amino acids within a
proteins that arrange themselves in a helical
structure.
A beta sheet is a group of amino acids within a
protein that arrange themselves in a stable
aligned (parallel) configuration.
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Secondary Structure Prediction Hydropathy
Plot Commonly used to identify alpha helices that
span a membrane (i.e. anchor protein to cell
wall).
1) Choose a moving window that travels along
the protein sequence a) calculates the overall
solubility of the amino acids in the
window. b) moves in amino acid c) repeat
calculation d) continue this though the entire
protein sequence.
Transmembrane domains are 20 amino acids, but any
size window can be used.
ELRLRYCAPAGFALLKCNDADYDGFKTNCSNVSVVHCTNLMNTTVTTGLL
LNGSYSENRT
1) Calculate average using amino acids-specific
constants.
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Secondary Structure Prediction Hydropathy Plot
ELRLRYCAPAGFALLKCNDADYDGFKTNCSNVSVVHCTNLMNTTVTTGLL
LNGSYSENRT
X (-3.5)(3.8)(-4.5)(3.8)(-4.5)(-1.3)(2.5)
(1.8)(-1.6)(1.8)(-0.4)(2.8)(1.8)(3.8)(3.8)
(-3.9)(2.5)(-3.5)(-3.5)(1.8)
WINDOW SIZE 20
Solubility Constants (Kyte Doolittle) A
Alanine 1.8 R Arginine -4.5 N Asparagine
-3.5 D Aspartic acid -3.5 C Cysteine 2.5 Z
Glutamine -3.5 E Glutamic acid -3.5 G Glycine
-0.4 H Histidine -3.2 I Isoleucine 4.5 L
Leucine 3.8 K Lysine -3.9 M Methionine 1.9 F
Phenylalanine 2.8 P Proline -1.6 S Serine
-0.8 T Threonine -0.7 W Tryptophan -0.9 Y
Tyrosine -1.3 V Valine 4.2
X 30.05 / 20 X 1.503
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Protein Folding Computationally Modeling
Biochemistry
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• OBJECTIVE
• Utilize the sequence information, along with
  temperature-dependent biomolecular interaction
  constants, to computationally predict a
  proteins tertiary structure.
• CHALLENGES
• It is NOT known how proteins fold in nature.
• More detailed or mathematically-intensive methods
  cant be completed in a reasonable time (given
  current computer capabilities).
• There are essentially no experimental methods to
  verify or validate that a predicted protein is
  correct or how correct.

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Monte Carlo simulation of a folding event. Each
frame displays the average position of a 48-mer
chain during a 104 iteration time window. The
color of each bead represents the variance of the
position of the bead during this time interval,
with yellow/green indicating large fluctuations
and blue indicating small fluctuations. The
entire folding event takes 8 x 105 iterations.
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• Evolution of Protein Folding Methods
• 1) Lattice Methods 3D lattice of residue or
  atomic positions.
• 2) Off-Lattice Methods Not reliant on
  predetermined 3D positions. Can include solvent
  effects.
• 3) All Atoms Methods/Modeling EXTREMELY
  computationally intensive.

• Tactics
• Initially calculate secondary structures minimums
  (fold sheets and helices), then calculate minima
  for remaining sequence.
• Emulate Protein synthesis process, starting from
  amino-terminus.
• Utilize existing NMR and X-ray crystal structures
  that match sequence under investigation.

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Protein Self-Assembly Good AND Bad
Quaternary Structure the interaction of multiple
proteins to form larger functional
structures. Many proteins bind to themselves to
form homodimers and homopolymers. Many proteins
bind to other proteins to form heterodimers and
heteropolymers.
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Many diseases involve self-aggregating proteins
(especially neurodegenerative diseases). Mad Cow
Disease (Prion Proteins) Alzheimers Disease
(beta-Amyloid Peptide) Huntingtons Disease Why
neuro-diseases? 1) Because the blood flow
(nutrients) to the brain is highly regulated, and
proteins that aggregate tend to collect and are
NEUROTOXIC. Note that these proteins ALSO
aggregate in peripheral tissues, but are
cleared and do not appear to be sufficiently
toxic. 2) Brain cells (neurons) do NOT
regenerate in a manner equivalent to peripheral
tissues (particularly in older people). 3) Loss
of neuronal cells leads to altered cognitive
capabilities, which is not the case in peripheral
tissues (e.g. slight muscle atrophy).
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Neurodegenerative Protein Diseases Beta Sheet
Structures!!! Beta-sheet structures are
sometimes called amyloid structures. Hence the
term Amyloidopathy NOTE The molecular forces
that assemble beta-sheet structures ALSO cause
them to self-assemble!
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2 key concepts regarding age-related
diseases. 1) Increased human health longevity
invents diseases. Before the modern age, nature
had rarely seen a 60 year old human. Imagine the
age-related diseases of the future when the
average human life span is gt120 years. 2)
Evolutionary pressures did not select for humans
to live much longer than 35-40 years. So
inherited mutations that lead to age-related
diseases were not selected out of the human
population. This fact has NOT changed in modern
times.
Alzheimers Disease 40-90 (sporadic at 60,
familial at 40), increases with age Men more
common under the age of 80 yrs Women more common
over the age of 80 yrs (J Neurol Neurosurg
Psychiatry 199966177 in BMJ 1999 Feb
27318(7183)614)
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Alzheimers Disease
Amyloid Precursor Protein
Beta Amyloid Protein
42 amino acids long
Self Aggregation
Neuronal cell nuclei (blue circles)
Senile Plaque
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Beta-Amyloid Aggregated in Water
500 nm
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Huntingtons Disease Incidence 2-8 persons per
100,000 worldwide with focal population
clusters Cause Known excess of trinucleotide
(CAG) repeats (encode glutamine) CAG
repeats 6-34 Normal Gene 36-120 HD Mutation
(majority 40-50 CAG repeats, 33-40 yr
onset) Number of repeats inversely related to
age of onset. Juvenile onset is rare and
involves CAG repeats gt60.
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Huntingtin Gene
10-30 CAG codons
Normal
Abnormal
gt 40 CAG codons
Huntingtin Protein
Abnormal
Normal
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Figure 1. Specific localization of huntingtin
aggregates in HD-repeat mutant mouse
brain.Low-magnification micrographs are shown of
brain sections from HD-repeat mutant (a) and
wild-type (b) mice at 27 months of age. Only the
striatum (Str) in the HD-repeat mutant mouse
brain was immunoreactive with EM48. Ctx, cortex.
High-magnification light micrograph (c) and
electron microscopy (d) show EM48-immunoreactive
aggregates in the neuronal nucleus (arrows). n,
Nucleus. Immunofluorescent double labelling shows
that striatal neurons containing intranuclear
EM48-reactive aggregates are labelled by
antibodies to calbindin-D (stars in e), but not
by antibodies to nitric oxide synthase (NOS f)
or parvalbumin (PARV g). Scale bars, 10  m
(a-c,f- g) and 0.5  m (d).
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Prion Protein Diseases
Creutzfeldt-Jakob Disease CJD humans
variant Creutzfeldt-Jakob Disease vCJD humans acquired from cattle with BSE
Bovine Spongiform Encephalopathy BSE "mad cow disease"
Kuru   infectious in humans who practiced cannibalism in Papua New Guinea
Gerstmann-Sträussler-Scheinker disease GSS inherited disease of humans
Fatal Familial Insomnia FFI inherited disease of humans
Scrapie   infectious disease of sheep and goats
other animal TSEs   cats, mink, elk, mule deer
1) Inter-species effect due to similarity between
prion protein sequences. 2) The role of the
normal prion protein in nature is not
understood. 3) The disease involves a
mis-folding of the prion protein to a beta-sheet
structure, which then self-aggregates.
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The illustration below compares a normal prion
protein (PrpC) to a disease-causing form (PrpSc).
The two structures exhibit two different, classic
protein motifs, called "alpha helices," and "beta
sheets." Alpha helices, seen here in the normal
prion (left), consist of linked amino-acid
building blocks that spiral around like a coiled
spring. Beta sheets form when amino acid chains
line up in a flat plane within the protein, as in
the disease-causing protein shown here.
Transmissible Spongiform Encephalopathy
Disease Form (self aggregating)
Normal Form