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Rough Games and the Brain:
The Structure and
Function of Proteins
Peggy Brickman
Department of Plant Biology
University of Georgia
1
“Wrestler Chris Benoit Brain’s Forensic Exam
Consistent with Numerous Brain Injuries”
Science Daily (Sept 6, 2007)
• Sports Legacy Institute President, Chris Nowinski, a
former Harvard football player and ex-professional
wrestler, after hearing of Chris Benoit’s death,
phoned Benoit’s father, Michael, with a ghoulish
request: to borrow the remains of his son’s brain.
• “When Nowinski contacted me about conducting
tests on Chris’ brain. I was extremely hesitant given
the circumstances surrounding my son’s death,”
said Michael Benoit.
2
Part I: The Case of Chris Benoit
• Twice recognized by the World Wrestling
Entertainment as the world heavyweight champion,
Canadian Professional Wrestler, Chris Benoit, was
booked to win his third championship the weekend
of his death.
• Instead, that weekend he killed his wife and strangled his seven
year-old son to death, and then hung himself using cords from a
weight machine.
• Medical examiners concluded that the elevated testosterone
levels in Benoit’s body (probably prescribed to remedy
deficiencies resulting after prior steroid abuse) did not
contribute to his violence.
• Nowinski believed that he did know the cause: repetitive head
injuries Benoit and other athletes suffer in contact sports that
result in Chronic Traumatic Encephalophathy (CTE).
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Rough Games
and the Brain…
• Impact: when the head slams into a hard surface, the skull
stops abruptly, while the brain, floating in cerebral fluid,
continues to move and is shaken and sometimes bruised.
• Animation
• Along with other damage, one result is that a nerve cell
protein called ß-Amyloid Precursor Protein (ßAPP) is cut
into pieces called ß-amyloid. Over time, neurofibrillary
tangles containing tau protein fibers accumulate.
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What are the functions of ß-APP, ßamyloid and tau?
• Results in axon growth and signaling to other
neurons. (ß-APP increases the levels of cell structure
protein, actin, for example. Tau is a protein found
with cell structure microtubules.)
• Healing: ßAPP triggers inflammatory response and
acts as anti-coagulant to prevent blood clots.
• May increase the expression of specific genes inside
the cells.
• Why would releasing ß-APP cause harm?
5
Benoit’s Brain
• Neurofibrillary tangles (NFTs, like those seen
below) have been found in patients suffering
cognitive and intellectual dysfunction, including
major depression. Similar NFTs were found in
Chris Benoit’s brain.
• Did wrestling cause his death?
6
Proof needed to demonstrate link
•
•
•
•
The Sports Legacy Institute (SLI), which oversaw and
coordinated the testing, is an independent medical
research organization dedicated to studying the longterm effects of head injuries in sports.
Most CTE occurs in boxers, but also in professional
football players.
Ten percent of retired pro-football players suffer from
depression, the same as the general population.
Before recommending drastic changes or additional
rules for athletes, what type of evidence or
experiments would you need to see linking
concussions to CTE and depression?
7
Concussions and CTE
• Study of more than 2,500 former NFL players by the
Center for the Study of Retired Athletes at UNC
found that cognitive impairment, dementia, and
depression rose proportionately with the number of
concussions they had sustained (Guskiewicz et al,.
2003, 2005, 2007). Those who had sustained 3 or
more concussions were more likely to experience
“significant memory problems” and 5 times more
likely to develop mild cognitive impairment.
• How might the NFL counter this data?
8
Dementia: CTE and Alzheimer’s
Disease
• Early pathologists stained
sections of an Alzheimer’s
brain with iodine and saw
large brown regions.
• Named ß-amyloid mistaken
for amylose
• Debate: Is it carb or lipid or
protein?
• What are the differences?
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CQ#1: Your answer should include the series of
numbers in order from the choices below.
Macromolecules are polymers composed of
monomers. For example polysaccharides like ___
are made of the repeating monomer ___. Fats
like ____ are made of the repeating monomer
____. Proteins are polymers made of repeating
monomers called ____.
1. Triglycerides
2. Amino acids
3. Glucose
4. Nitrogen
5. Nucleic Acids
6. Fatty Acid
7. Steroids
8. Cellulose
9. Phosphates
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II: Amino Acids:
Specific composition/
comparison with
biological molecules.
other
• Tau protein tangles like those in Benoit’s
brain and ß-amyloid:
• 1984: Scientists purified protein from the tangled fibrils
seen in Alzheimer’s brains.
• 1987: cloned the gene which coded for a 695 amino acid
protein (ß-APP) which spanned the phospholipid bilayer.
• ß-amyloid are fragments of the protein that are composed
of 28 amino acids strung together.
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Not all early-onset dementia comes from
brain trauma.
• Most Alzheimer’s occurs in the elderly but about 1/2
million of the 5 million people who develop
dementia or Alzheimer’s each year are under 65.
They didn’t all have brain trauma, but some had a
family history of the disease.
• Maybe these athletes’ dementia was inherited (5% of
Alzheimer’s is caused by inherited dominant
mutations.)
• People with the mutation develop symptoms much
earlier than typical (~age 51).
• Some of these mutations (15%) are changes in the
amino acids in the ß-APP protein.
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Structure of Amino Acids
• All amino acids have an amino and a carboxyl
(acid) attached to a central carbon along
with Hydrogen
H
amino
group
H
O
N
C
H
carboxyl
group
C
OH
R
side-chain
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Amino acids differ in the R group
• 20 Different R groups: Some
• Non-polar (Hydrophobic)
H
O
H
N
C
C
H
OH
CH2
phenylalanine
• Polar (Hydrophilic)
H
O
H
N
• Some even contain sulfur
C
H
C
OH
CH2
glutamine
C
H2N
O
14
15
Amino Acids Form Proteins
• Condensation
reactions create a
covalent bond
(Peptide Bond)
• Forms Polypeptides
• DNA gene mRNA
• Primary structure:
met
gln
thr
phe
H
N
C
N
OH
C
H
O
H
C
H
glu
H
O
H
C
H
N
OH
CH3
CH2OH
alanine
serine
C
H
OH
glycine
H2O
H
O
H
H
O
H
H
C
C
N
C
C
N
C
O
H
H
C
OH
CH3
ala
C
H
H2O
N
O
H
ser
CH2OH
gly
leu
H
lys
tyr
pro
16
Each protein has a different pattern of
amino acids
insulin
gly
ile
val
glu
gln cys cys
ala
ser
val
cys ser leu
tyr
B amyloid
asp ala
glu phe arg
his
ser
gly
tyr
glu
val his his
B amyloid
mutation
asp ala
glu phe arg
his phe ser
gly
tyr
glu
val his his
val
• The R-group of each amino acid is different, and thus
imparts different qualities to the protein.
• Hydrophilic amino acids are attracted to other hydrophilic
substances, and hydrophobic are not.
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III. How Proteins Differ:
Function
Structure
Protection
Metabolic Enzymes
Channels
& pumps
Transport
Gene Expression
and Regulation
Cell Signaling (Hormones)
Movement
18
How Can Proteins Have Many Different
Functions? Scrabble Analogy
• Carbohydrates: Glucose
G1
G1
G1
G1
G1
G1
• Hard to make more
than one word
•
•
•
•
•
•
•
•
Proteins: 20 amino acids:
Glutamine
Isoleucine
E1
Asparagine
I1
Serine
Threonine
N1
Lysine
S1
Arginine
T1
K5
R1
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CQ#2: What do proteins, lipids, and
carbohydrates all have in common?
A: Type of reaction that links them covalently into
large polymers of repeating monomers.
B: Having more than a dozen monomers present in
one polymer.
C: Presence of the element N.
D: Having monomers that can be either hydrophobic
or hydrophilic.
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IV. Proteins Fold into Active Shape
• Primary Structure: sequence of amino acids in
polypeptide. For each protein (ß-APP) the
primary structure is always identical.
ß-amyloid
asp ala
glu phe arg
his
ser
gly
tyr
glu
val his his
ß-amyloid
mutation
asp ala
glu phe arg
his phe ser
gly
tyr
glu
val his his
val
Why would changing one amino acid cause the whole
protein to change shape?
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Secondary Structure
• Hydrogen bonds create shape.
• Due to backbone interactions
• http://www.stolaf.edu/people/giannini/flashanimat
/proteins/hydrophobic%20force.swf
• Some examples of permanent structures:
• a-helix or b-pleated sheet
primary structure
amino acids
b-sheet
ahelix
secondary structure
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Tertiary Structure
• 3D packing of polypeptides. Often
involve H-bonds.
primary structure
amino acids
b-sheet
ahelix
secondary structure
Folding animation
b-sheet
tertiary structure
ahelix
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Quaternary Structure
• Interactions
between two or
more polypeptide
chains.
• Not found in all
proteins.
primary structure
amino acids
b-sheet
ahelix
secondary structure
b-sheet
tertiary structure
ahelix
quanternary structure
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CQ#3: Proteins such as the ß amyloid and
the mutant ß amyloid that results in earlyonset Alzheimer’s differ from one another
because:
A. Peptide bonds linking the amino acids differ from one
protein to another.
B. The two proteins have a different combination of amino
acids along the chain of the polypeptide.
C. Presence of the element N is only found in one protein.
D. The two proteins each contains their own unique types of
amino acids.
E. The number of amino acids in the chains differ.
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Sickle-cell anemia results from a point mutation in the HBB gene. The
mutation results in the replacement of an amino acid that has a
hydrophilic R-group with an amino acid that has a hydrophobic Rgroup on the exterior of the hemoglobin protein. Such a mutation
would most likely result in altered
A. Properties of the molecule as a result of
abnormal interactions between adjacent
hemoglobin molecules.
B. DNA structure as a result of abnormal hydrogen
bonding between nitrogenous bases.
C. Fatty acid structure as a result of changes in ionic
interactions between adjacent fatty acid chains.
D. Protein secondary structure as a result of
abnormal hydrophobic interactions between Rgroups in the backbone of the protein.
IV. ßAPP Proteins and Alzheimer’s
Disease
• ß-amyloid precursor protein (ßAPP)
found in the phospholipid outer cell
membranes of neurons.
Outside the Cell
ßAPP
neurons
Inside the Cell
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CQ#4: Which of the following amino
acids might you NOT expect to find in
the intra-membrane region of the ßAPP protein?
H
A
H
H
N C C
O
OH
CH
CH3 CH3
H
B
H
C
H
H
N C C
H CH
OH
serine
O
N C C
OH
CH3
valine
H
H
alanine
H
O
OH
D
H
H
N C C
O
OH
H CH
O C O-
aspartate 29
ßAPP and Alzheimer’s
• typically polypeptides met
are converted back into
amino acids (by
proteases) by a reaction
called hydrolysis.
• Also used to remove
proteins when they are
no longer needed, and
to send signals.
gln thr phe glu ala ser gly leu lys tyr pro
H
O
H
H
O
H
H
C
C
N
C
C
N
C
O
H
N
C
H
OH
CH3
CH2OH
H2O
H
O
H
N
C
H2O
H
N
OH
O
H
C
H
H
C
H
C
H
N
OH
CH3
CH2OH
alanine
serine
O
H
C
C
H
OH
H
glycine
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Brain Trauma Speeds Hydrolysis
ßamyloid
• ß-amyloid precursor protein (ßAPP)
mutant
more
Outside the Cell
ß-secretase
sticky?
ß-amyloid
ßAPP
Inside the Cell
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http://www.healthscout.com/animation/68/7/main.html
CQ#5: Which amino acid
change in the ß-amyloid protein
fragment would be most likely
to make it stick together?
aqueous environment
outside nerve cell
ß-amyloid
A. Replacing an amino acid with a hydrophilic R-group with
one that is hydrophobic.
B. Replacing an amino acid with a hydrophobic R-group with
one that is even more hydrophobic.
C. Replacing an amino acid with a hydrophobic R-group with
one that is hydrophilic.
D. Replacing an amino acid with a hydrophilic R-group with
one that is even more hydrophilic.
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Shape and stickiness of protein
dictated by amino acids
• Changing amino acid 717 from valine to
phenylalanine can produce ß-amyloid that
sticks much better to other ß-amyloid
fragments and causes plaques to form sooner
in familial Alzheimer’s.
H
O
H
N
C
N
OH
CH3
CH3
valine
O
H
C
H
CH
H
C
H
C
OH
CH2
phenylalanine
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CA#6: Even if they don’t have the mutation, hard-hitting
athletes may want to take some precautions. Perhaps taking a
chemical designed to be more attractive to the basic R-groups
on ß-amyloid fragments than the fragments were to each other
thus preventing their aggregation. Phase III trials of such a drug
(Alzhemed) in 2007 failed to show benefits in 1,000 Alzheimer’s
patients. What kind of bonds in the ß-amyloid protein was
Alzhemed meant to disrupt?
A. Peptide bonds
B. Hydrogen bonds
C. Non-polar covalent bonds D. Polar covalent
bonds
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CQ#7: OK, so that drug didn’t work. What do you think the
NFL should do for football players?
A. Test for Alzheimer’s predisposition mutations as part of
physicals prior to hiring.
B. Impose mandatory rest after head injuries.
C. Fine or fire coaches breaking the rules.
D. Change how the game is played so that concussions are
less likely to occur.
E. It’s just part of the game like shot knees and back pain.
Make sure the former players have great health
insurance that includes psychiatric coverage and nursing
home care.
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NFL and Concussions
• Although continuing to support its policies on
concussions and rejecting any link between
concussions and depression & CTE, NFL has
several initiatives:
– League and players union created a fund to help pay medical expenses
of players suffering from dementia.
– New guidelines include: obligatory neuropsychological testing,
“whistle-blower system” for anonymous reports of any coach’s attempt
to override the wishes of concussed players or medical personnel.
• Handheld EEGs, and functional magnetic resonance imaging also
in testing for early diagnosis.
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CQ#8: You might want to know if you have a mutation
in another gene, ApoE4 that changes the amino
cysteine to arginine, and affects age of onset of
Alzheimer’s. How could changing the sequence result
in a change in function?
A. It could change the amount of the
protein produced.
B. It could change the protein’s 3-D
shape and thus its ability to
interact with other proteins.
C. It could change where the protein
is located in the cell.
D. It could change the number of
amino acids.
Copies of
ApoE 4
mutation
Average age
of onset of
Alzheimer’s
2
68
1
75
0
84
37
Alzheimer’s Prevention
• OK. You’re not a heavy-hitting athlete and don’t have any
family history of Alzheimer’s, just like the 4.8/5 million people
with Alzheimer’s over 65.
• So, there’s a 10% chance you’ll get it by age 65, and a 50%
chance you’ll get it by 85.
• People who eat the so-called 'Mediterranean diet' (fruits,
vegetables, bread, pasta, fish, olive oil and a little red wine,
but low in dairy products and red meat) have a lower risk of
Alzheimer's disease. So do people who exercise regularly, use
(not lose) their minds, and have been taking non-steroidal
anti-inflammatories like aspirin for least two years.
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Interested in Reading More?
• Scientific American:
http://www.scientificamerican.com has a great set of
articles on the latest research, including one in
March 2009 on prions and Alzheimers.
• Articles:
– Medline search
http://www.nlm.nih.gov/medlineplus/alzheimersdisease.h
tml
– Lobo, I. (2008) Epistasis: Gene interaction and the
phenotypic expression of complex diseases like
Alzheimer's. Nature Education 1(1)
– McKee et. al. (2009) Chronic Traumatic Encephalopathy in
Athletes: Progressive Tauopathy After Repetitive Head
Injury. J. Neuropathol Exp Neurol.
39
Heat
pH changes
High salts
41
Image Credits
Slide 3:
Description: Photo of Chris Benoit.
Source: From Wikimedia Commons, by dani nuestro from Bangkok,
http://commons.wikimedia.org/wiki/File:BenoitInTheRing.jpg
Clearance: Creative Commons Attribution-Share Alike 3.0 Unported License(CC BY-SA 3.0).
Slide 4
Description: Concussion illustration.
Source: Adapted from Wikimedia Commons,
http://commons.wikimedia.org/wiki/File:Concussion_Anatomy.png, original by Max Andrews.
Clearance: Used in accordance with the Creative Commons Attribution-Share Alike 3.0 Unported license.
Slide 6
Description: Neurofibrillary tangles in the Hippocampus of an old person with Alzheimer-related pathology.
Source: Wikimedia Commons,
http://commons.wikimedia.org/wiki/File:Neurofibrillary_tangles_in_the_Hippocampus_of_an_old_person_with
_Alzheimer-related_pathology,_Gallyas_silver_stain.JPG, by Patho.
Clearance: Used in accordance with the Creative Commons Attribution-Share Alike 3.0 Unported license.
Slide 9
Description: Illustration comparing healthy brain and Alzheimer’s brain.
Source: Alzheimer’s Association, http://www.alz.org/brain/09.asp
Clearance: Used with permission.
42
Slide 11
Description: Illustration of plaques.
Source: Alzheimer’s Association, http://www.alz.org/brain/11.asp
Clearance: Used with permission
Slide 18
Description: Scrabble analogy for the greater complexity of proteins due to the greater number and type of
monomers.
Source: Peggy Brickman, adapted from “Nutrition” Insel, Turner, and Ross.
Slides 21-23
Description: Illustration of four levels of protein structure.
Source: Wikimedia Commons, http://commons.wikimedia.org/wiki/Image:Proteïen_struktuur.png.
Clearance: Used in accordance with the Creative Commons Attribution-Share Alike 3.0 Unported license.
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