4. Protein Aggregation Diseases

Question 1

What are the two types of protein aggregates?

What is the difference between them?

Answer 1

1. Non-amyloid aggregates
   
   • Non cross-ß structure
   • Causes sickle-cell anemia
2. Amyloid aggregates
   
   • Cross-ß structure
   • Causes Alzheimer’s, Parkinson’s, cataractsm, etc…

Question 2

sickle-cell anemia

Answer 2

• Disease characterized by low number of RBCs
• RBCs have a lifespan of 10-20 days (compared to a regular lifespan of 120 days)
• A single mutation in hemoglobin causes sickle-cell disease
  
  • Causes non-amyloid aggregates
• ​Africa has the highest prevalence of sickle-cell disease in the world
• In the United States, African-Americans are more likely to have sickle-cell disease

Question 3

What is the mutation that causes sickle-cell anemia?

Answer 3

The most common mutation that causes sickle cell anemia is a single nucleotide change from GAG to GTG at amino acid position 6 in the β-chain of Hb.​

Mutation from GAG to GTG replaces Glu by Val at amino acid position 6 in the β-chain of Hb. Mutated β-chain is called as sickle(s) chain and mutated Hb as HbS.

Question 4

What is the phenotype and genotype of an individual who inherits both sickle-cell disease genes?

Answer 4

A person will be born with sickle cell disease only if two genes are inherited—one from the mother and one from the father.

All hemoglobin molecules in patients consist of 2 alpha and 2 s (sickle or mutated beta) chains.

Question 5

What is the genotype of an individual who is a carrier of the sickle cell trait?

Answer 5

A person who inherits just one gene is healthy and said to be a “carrier” of the disease or sickle cell trait. A carrier has an increased chance of having a child with sickle cell disease if he or she has a child with another carrier.

In these persons, half of their hemoglobin molecules consist of 2 alpha and 2 beta chains, and half consist of 2 alpha and 2 s chains.

Question 6

Describe the mutation that causes sickle-cell anemia?

Answer 6

The mutation is from Glu to Val at 6th amino acid in the ß-chain, which is a charged residue to hydrophobic residue mutation.

Mutation makes Hb more hydrophobic at the site of mutation. Individual sickle Hb (Hb S) molecules stick to one another forming long rod-like fibers, twined as a helical bundle.

Question 7

What structure do the aggregates look like in sickle-cell anemia? What does this ultimately cause the tertiary structure of Hb to look like?

Answer 7

• Aggregates are of α-helical structure.
• The content of α-helical structure is similar between sickle and adult hemoglobins.
• The interface between four polypeptide chains is more open.

Question 8

What disease is the 5th leading cause of death?

Answer 8

• 5.3 million Americans have AD, 5th leading cause of death in those ≥65 years old. Following AD diagnosis, survival is typically 4-6 yrs.

Alzheimer’s disease is the most common cause of dementia in older people.

Question 9

What is the typical age that Alzheimer’s disease presents?

Answer 9

• The prevalence of AD increases exponentially with age.
• Therefore, most cases present in people older than 65 years but about 5% of cases occur younger than age 65 and can be as young as 40 years.

Question 10

What are the four risk factors of AD?

Answer 10

1. Age
   
   • In general, the autosomal dominant form of Alzheimer’s disease begins between age 40-60 years, and ‘late onset’ familial or sporadic Alzheimer’s disease begins between 70-85 years.
2. Genetic Factors
   
   • Alterations on chromosome 1, 14, or 21
3. Head injury
4. Gender
   
   • Alzheimer’s disease is more common in women.

Note: Prevalence is independent of the race.

Question 11

What are the cognitive clinical presentations of AD?

Answer 11

• Memory loss (poor recall and losing items)
• Language impairment
• Impaired executive function (unable to perform tasks or movements when asked)
• Disorientation (Impaired perception of time and unable to recognize familiar people)

Question 12

What are the noncognitive clinical presentations of AD?

Answer 12

• Depression, hallucinations, delusions
• Behavioral disturbances (physical and verbal aggression, motor hyperactivity, uncooperativeness, wandering etc)

Question 13

What are the functional clinical presentations of AD?

Answer 13

• Inability to care for self (dressing, bathing, toileting, and eating)

Question 14

What is considered to be the fundamental cause of AD?

Provide evidence.

Answer 14

• Postulated that amyloid beta (Aβ) deposits are the fundamental cause of the disease.
• Support for this postulate comes from the location of the gene for the amyloid beta precursor protein (APP) on chromosome 21
• People with trisomy 21 (Down Syndrome) who thus have an extra gene copy almost universally exhibit AD by 40 years of age.

Question 15

What is the pathophysiology that characterizes AD?

Answer 15

Alzheimer’s disease is characterized by loss of neurons and synapses in the cerebral cortex and certain subcortical regions.

Question 16

In AD, misfolding occurs in what two proteins?

Answer 16

1. Amyloid Plaques*-deposits of the beta-amyloid protein generated from Alzheimer amyloid precursor protein (APP) which accumulates in spaces between nerve cells. This interferes with the neuron to communicate with one another resulting in abnormal brain function and apoptosis.
2. Neurofibrillary Tangles*-accumulate inside of nerve cells. They are deposits of tau proteins that are involved in microtubule formation.

Question 17

Describe amyloid precursor proteins in a patient with AD.

Answer 17

Amyloid precursor protein (APP) is cleaved by β-secretase and γ-secretase to form the neurotoxic Aβ42 fragment that aggregates into the amyloid plaque that is a hallmark feature of AD. The gene for APP is located on chromosome 21. The exact function of APP is not known. The Aβ fragment has no structure in its native state.

Question 18

Describe the role of Tau in a healthy neuron.

Answer 18

• Tau is a major microtubule associated protein (MAP) of a mature neuron.
• Its primary role is to stimulate tubulin assembly into microtubules and to stabilize neuronal cytoskeleton by interacting with microtubules.
• Tau binds to axonal microtubules and result in the formation of microtubule bundles.

Question 19

Describe Tau in a patient with AD.

Answer 19

• Phosphorylation level of tau isolated from autopsied AD brain is 3-4 fold higher than normal human brains.
• Disruption in the balance of protein kinases and phosphatases contributes to abnormal phosphorylation of tau observed in AD.
• Results in the destabilization of microtubules and self-aggregation of tau into neurofibrillary tangles (NFTs).
• Break down of microtubule network leads to a compromised axonal transport and ultimately loss of synapses and degeneration.

Question 20

Describe the kinetics of amyloid formation.

Answer 20

1. Native protein, if structured, partially unfolds leading to the exposure of hydrophobic residues.
2. More than one unfolded molecule with exposed hydrophobic patches stick with one another to form circular aggregates known as oligomers.
3. Oligomers stack to form long protofibrils.
4. Protofibrils twine around one another to form long amyloid fibrils.
5. Amyloid fibrils precipitate resulting in large amyloid plaques.

Question 21

Are oligomers or protofibrils toxic?

Answer 21

Oligomers.

Question 22

Describe the difference between a normal ß-hairpin structure and a cross-ß helical structure.

Answer 22

• Cross-ß helical structure.
• Long ß-sheet is between two different molecules.
• ß-strands from the same molecule perpendicular to the long axis, but no H-bonds.
• H-bonds between ß-strands from different molecules parallel to the long axis.

Question 23

Where do the side chains of the ß-amyloid structure protrude?

Answer 23

• Dry interface between the two sheets; Wet interface on the outside surfaces.
• Complementarity of side chains protruding into the dry space.

Question 24

What are the two processes of amyloid growth involved in AD?

Answer 24

1. Initial formation of small nuclei (nucleation)
2. Fibril growth starting from nuclei (propagation)

Question 25

What is the difference between amyloid formation of pre-formed seeds and those without seeds?

Answer 25

* Kinetics: Two steps * Lag phase: Nucleation * Propagation * Pre-formed seeds: * No lag phase * No need for nucleation

Question 26

In Alzheimer's disease, when do amyloid aggregates start accumulating in a brain?

Answer 26

Amyloid aggregates start accumulating in human brain at least 20 years before the onset of Alzheimer’s disease symptoms.

Question 27

What is the order of pathophysiological characteristics that occur before the onset of AD symptoms?

Answer 27

Question 28

What protein becomes misfolded in Parkinson's disease?

Answer 28

* A natively unstructured protein, a-synuclein, becomes misfolded. * Forms amyloid-like filaments * Dopaminergic neurons develop characteristic filamentous inclusions called Lewy Bodies * Problem: accumulation of protein aggregates rich in β-pleated sheet conformation conferring toxic properties

Question 29

What characterizes Parkinson's Disease?

Answer 29

* Progressive neurodegenerative disorder characterized by tremor at rest, rigidity, bradykinesia, stooped posture, loss of postural reflexes, and shuffling gait. * Autonomic dysfunction and cognitive deficits may occur with disease progression.

Question 30

What is the incidence of Parkinson's disease (i.e., how many Americans, demographs, gender)?

Answer 30

* As many as one million Americans live with Parkinson's disease. * Approximately 60,000 Americans are diagnosed with Parkinson's disease each year. * **Incidence of Parkinson’s increases with age**, but an estimated four percent of people with PD are diagnosed before the age of 50. * Men are one and a half times more likely to develop Parkinson's than women.

Question 31

What can trigger or cause Parkinson's disease as a secondary condition?

Answer 31

1. Drug-induced parkinsonism - most commonly antipsychotic and dopamine-blocking medications 2. Vascular disease 3. Metabolic conditions, such as hypoparathyroidism 4. Structural lesions including tumors, infarctions, and hydrocephalus 5. Toxins or pesticides ## Footnote *About 25% of individuals have PD secondary to these triggers.*

Question 32

Huntington's Disease/Chorea

Answer 32

Huntington's Disease is a neurological disorder characterized by jerky involuntary movements affecting especially the shoulders, hips, and face. It is an inherited **autosomal dominant** progressive neurodegenerative disease characterized by: * Chroeiform movements * Psychiatric problems * Dementia

Question 33

If you are a child of HD affected parents, what is the likelihood of developing HD?

Answer 33

50%

Question 34

Describe the wild-type Huntington peptide.

Answer 34

* Necessary for normal embryonic development * Helps with cell regulatory processes * Gene is located on Chromosome 4 * Huntington gene is widely distributed throughout the brain and many tissues throughout the body * Wild Type Huntington contains 6-34 Cytosine-adenine-guanine (CAG) trinucleotide repeats in the HTT gene near the amino terminus. * CAG trinucleotide sequence codes for the amino acid glutamine

Question 35

What is the genetic determinant of HD?

Answer 35

* Number of CAG repeats important in diagnosis, disease onset and severity. * Toxicity of the Huntington gene occurs with expanded trinucleotide repeats * More than 35 repeats: expanded, unstable, disease-causing * 36-39 repeats: possibly disease causing * More than 40 repeats: disease causing

Question 36

Why are three theories as to why multiple CAG sequences are so detrimental in HD?

Answer 36

* Normal tertiary protein conformation destabilized by expanded glutamine → formation of insoluble beta-pleated sheets. * Expanded polyglutamate tract → increased transglutamate-mediated cross linking with other polyglutamine proteins * Disruption of several intracellular pathways.

Question 37

Where do protein aggregates occur in HD?

Answer 37

Protein aggregates appear in the cytoplasm and nucleus of affected cells.

Question 38

What area of the brain does HD affect? What is this brain responsible for?

Answer 38

HD mainly attacks the cells in the basal ganglia in the brain which controls: * Movement * Emotion * Cognitive ability

Question 39

In the picture below, what brain correlates to the toxicity of HD?

Answer 39

Neither; toxicity of Huntington does not correlate with degree of cell/ neuronal atrophy.

Question 40

Rhes

Answer 40

* May cause Huntington's Disease * Small guanine nucleotide binding protein selectively located in the striatum * Possibly related to cell dysfunction * Exact mechanism still unknown

Question 41

amyotrophic lateral sclerosis (ALS)

Answer 41

* Neurodegenerative disease affecting nerve cells in the brain and spinal cord * Progressive weakness of voluntary muscles, resulting in paralysis * Leads to death due to respiratory muscle failure * Form of motor neuron disease (MND)

Question 42

What occurs in both the initial and advanced stages of ALS?

Answer 42

* Initial stages: * Muscle weakness in hands, arms, legs, muscles of speech, swallowing or breathing * Twitching and cramping of muscles * Advanced stages: * Impaired speech * Difficulty breathing and swallowing

Question 43

What are the four causes of ALS?

Answer 43

Question 44

superoxide dismutase 1 (SOD1)

Answer 44

* **Protects cells from superoxide toxicity** by converting to harmless water. * **SOD1 is a homodimer in which each subunit binds one copper atom and one zinc atom. Cu carries out the chemical change. Zn binding promotes dimerization and stabilizes the native form.** * Mutations in SOD1 cause familial ALS.

Question 45

What causes SOD1 to fold?

Answer 45

* ALS-causing SOD1 mutations promote production of Copper-deficient misfolded species. * The aggregation is from a well-folded, structured protein, unlike the case of Alzheimer’s where the aggregation is from a natively unfolded protein.

Question 46

How do amyloid aggregates cause toxicity? That is, what cell machinery is affected by misfolded proteins?

Answer 46

1. Coaggregation of essential cytoplasmic components 2. Poisoning of the proteasome thereby inhibiting timely degradation of many cellular proteins 3. Saturation of cytoplasmic chaperones that catalyze essential protein folding and refolding 4. Damaging mitochondria by aggregation onto the cytoplasmic surface and/or transport into the mitochondrial intermembrane space

Question 47

prion disease

Answer 47

* Transmissible spongiform encephalopathies (TSEs) * Fatal, degenerative neurological disease * Replicate by binding to normal prion protein (PrPC) which converts to disease-causing prion protein (PrPSC) * **PrP^C – rich in α-helices** * **PrP^SC – rich in ß-sheets** * Aggregation of PrP^SC leads to disease

Question 48

What are human prion disease

Answer 48

* Creutzfeldt-Jakob Disease (CJD) * Variant Creutzfeldt-Jakob Disease (vCJD) * Gerstmann-Straussler-Scheinker Syndrome * Fatal Familial Insomnia * Kuru

Question 49

Creutzfeldt-Jakob Disease (CJD)

Answer 49

* Rapidly degenerative brain disorder * Leads to a rapid decrease in * Mental function * Movement

Question 50

What are the three types of CJD?

Answer 50

1. Sporadic = spontaneous * Idiopathic * **85% of cases** 2. **Familial** = inherited genetic mutation * Defect in the prion protein gene * **5 to 15%** of cases 3. Acquired = exposure to infectious prion * **Contaminated meat** = human variant of BSE * US-EU Beef Hormone Dispute * Transplant of contaminated tissues * Contaminated instruments during surgery

Question 51

What is the prevalence and statistics of CJD?

Answer 51

* Rare * 1 out of 1 million people worldwide * In the US: 250 – 300 new cases yearly * Age * Sporadic * Onset = 55 to 70 years old * Median age of death = 68 years * Familial * Onset = 45 to 60 years old * Death: months – 2 years

Question 52

What are the risk factors for CJD?

Answer 52

* Age * Genetic: **chromosome 20** * Patients who received contaminated * Corneal grafts * Dura mater grafts * Sterotactic electroencephalography electrodes * Neurosurgical instruments * Human growth hormone * Human pituitary gonadotropin

Question 53

What are the early signs of CJD?

Answer 53

* Concentration * Memory * Judgment difficulties

Question 54

What are the symptoms of later-stage CJD?

Answer 54

* Myoclonic jerks or seizures * Abnormal reflexes * Muscle stiffness * Strong startle response * Weakness and loss of muscle tissue

Question 55

What are the complications of CJD?

Answer 55

* Infection * Loss of ability to * Interact with others * Function * Care for oneself * Heart failure * Respiratory failure * Death

Question 56

What misfolded protein causes cataracts?

Answer 56

* The lens is made of crystallin proteins * Must be well-ordered for vision clarity * Any disruption in the arrangement of the crystallins causes cataract. * Crystallins also act as chaperones. Defective crystallins result in more protein aggregates.

Question 57

What are the two types of crystallins that are relevant to cataracts?

Answer 57

**Human γ D-crystallin and Human γ S-crystallin are monomeric two-domain major proteins of the human eye lens and significant components of mature-onset cataracts.** Mature-onset cataracts are thought to be an aggregated state of modified, damaged, or partially unfolded states of lens crystallins.

Question 58

What are the five different types of cataracts?

Answer 58

Question 59

What are the three signs of cataracts?

Answer 59

* Blurred vision * Starts as a small blurry area that continually grows * Yellowing/browning of vision * Unable to identify blues, purples, and greens * Poor night vision

Question 60

What are the facts and figures of cataracts in the U.S.?

Answer 60

* Currently, one out of every 3 Americans over 65 years old have cataracts. * By the year 2030, approximately 70 million Americans will be over 65 years. * Estimated 21 million Americans will have cataracts in 2030.

Question 61

How should you prevent cataracts?

Answer 61

* Wear sunglasses * Eat a healthy diet * Stop smoking * Decrease alcohol consumption * Control diabetes