Week 1 ICL Flashcards
1
Q
Can the prediction of a protein’s secondary conformation for it’s primary AA sequence help to understand the proteins capacity to form disease inducing aggregates?
A
Yes
2
Q
Which diseases are associated with protein misfolding?
A
- neurodegenerative diseases
- Alzheimer’s
- spongiform encephalopathies
- Parkinson’s
- dementia with Lewy bodies
- frontotemporal dementia with Parkinsonism
- amyotrophic lateral sclerosis
- Huntington’s
3
Q
What is spongiform encephalophathies?
A
Aggregating prion proteins
4
Q
What is a prion?
A
Self-propagating forms of chromosomally encoded protein
Prions are in the cytoplasms of neurons and they are self-propagating!
They are characterized morphological by spongiform change in trace lilac vacuoles in neurons and glia and clinically by rapid dementia
5
Q
How does a protein become infectious? Like in spongiform encephalophathies
A
- A normal prion protein undergoes conformational alteration to a different conformation (we don’t know how)
- Normal prion exposed to “infectious” prion that has already undergone conformational alteration
- Replication/infection happens through transferring protein misfolding (domino effect) to form insoluble aggregates - once one protein is “infected” all the proteins around it also undergo this bad conformational change
This was studied in mice models with scrapy
6
Q
What are prion diseases due to?
A
The normal alpha-helices of prions get turned into b-sheets that aggregate and are resistant to digestion with protease
B- sheets are normal and okay but not for prions; normal prion proteins are alpha helical and soluble
Disease of propagation of incorrect protein folding of the SECONDARY structure
7
Q
What is Creutzfeldt-Jakob disease?
A
Very rare
Rapid progressive dementia and death - there’s no treatment
It’s called scrapie in mice
Bovine spongiform encephalopathy (BSE)(mad cow disease)
Chronic wasting disease(CWD) in deer
8
Q
What is Kuru?
A
Rare progressive fatal prion disease that resembles Creutzfeldt-Jakob disease and was found in the tribes of Papúa New Guinea because they ate the brains of dead relatives
9
Q
What are the 3 classifications of Creutzfeldt-Jakob disease?
A
- Sporadic
- Inherited/familial
- Transmissible
- Variant
10
Q
What is sporadic Creutzfeldt-Jakob disease?
A
85%
No known source = idiopathic
11
Q
What is inherited/familial Creutzfeldt-Jakob disease?
A
10-15%
Autosomal dominant due to genetic mutation in coding sequence for prion protein
Autosomal dominant = only one genetic allele needs to be effect for it to manifest as the disease
12
Q
What is transmissible Creutzfeldt-Jakob disease?
A
<5%
Iatrogenic = physician caused, transmitted by medical or surgical sources
Injections of human material, corneal transplant, brain electrodes
13
Q
What is variant Creutzfeldt-Jakob disease?
A
<1%
People eating meat from bovine spongiform encephalopathy in the UK
Different clinical course than Creutzfeldt-Jakob disease because it happens at a younger age, longer duration and different histopathology of brain
Current concern is blood transfusion transmission of people that have visited the UK
14
Q
What is the abbreviation for prion? Abnormal prions?
A
PrPC = normal PrP, cellular
PrPSC = abnormal PrP causing prion disease
SC for scrapie because of the animal models used
15
Q
What does PrPSC need to manifest?
A
-/- don’t have either gene for normal prions, both their prion genes have been knocked out by researchers
If you give them PrPSC then they don’t get prion disease!! This means that PrPSC requires expression of the normal prion protein PrPSC +/+ in order to manifest
The weird thing is that these mice without the prion protein -/- seem to be exactly the same as the +/+ mice so we don’t actually know what this prion protein does….
16
Q
What is the general reason for Alzheimer’s, Parkinson’s and Huntington’s?
A
Characterized by clumps of proteinaceous material called amyloid
People are speculating that their spread through the brain in a prion-like behavior aka they’re “infecting” other proteins
The diseases have been successfully spread between lab animals by the inoculation of diseased brain samples into the brains of healthy animals
17
Q
What is the MOST unusual biological aspect of CJD compared to other infectious diseases?
A
It’s caused by an “infectious” protein
18
Q
Prion disease is attributed to an infectious change in the secondary structure of the prion protein, resulting gin irreversible aggregation, neurodegeration and death. Secondary structure of polypeptide chains….
A
Consists primarily of two regularly repeating secondary structures
Alpha helices and b-sheets
19
Q
What are examples of post-translational collagen diseases?
A
Scurvy, ehlers danlos syndrome, osteogenesis imperfecta
20
Q
What are post-translational protein modifications?
A
Covalent post-translational modification by conjugation of “prosthetic groups” that help to augment or inhibit protein function
Ex. Acetylation, glycosylation, methylation, etc.
21
Q
What are the most frequently modified AA in protein?
A
Phosphoserine, phosphotyrosine, and phosphothreonine
Help to modulate different cellular functions
22
Q
What does the hydroxyl group addition to proline and lysine do?
A
Post translational modification that stabilized fibers of collagen
23
Q
How common is collagen?
A
More than 30% of all proteins in the body is some form of collagen (includes bones)
Major extra cellular matrix structural protein
24
Q
What is collagen made of?
A
33% glycine (smallest nonpolar AA) and it’s in every 3rd position!
Elongated triple-helical structure stabilized by inter chain hydrogen bonds from hydroxy-lysine and hydroxy-proline
25
What is collagen important for?
Wound healing!
26
How does your body make collagen protein?
Made in fibroblasts in the RER
It’s then post-translationally modified, forms triple helix and is secreted into the ECM
Then mature collagen fibrils aggregate and become cross-linked to make collagen fibers
Meat is tough if it has a lot of collagen
27
What AA is effected when there are problems with collagen?
Gly-X-Y
Hydroxylated proline and lysine are in the Y position, every 3rd AA spot in the AA chain of collagen
If they’re mutated or missing you have problems
28
What is scurvy? Which post-translational disorder does it lead to?
A consequence of deficient vitamin C needed for hydroxylation of proline and lysine
Vitamin C is a co-factor required for hydroxylation
Without hydroxylation the triple helix can’t hydrogen bond well enough and there’s an instability and loss of strength of collagen
So scurvy leads to a post-translational disorder of collagen formation!
29
What does vitamin C do in collagen formation?
Ascorbate = vitamin C
Vitamin C is a reducing agent to keep Fe+2 from oxidizing to Fe+# which allows hydroxylation of proline and lysine to proceed
Without vitamin C correct collagen post-translational hydroxylation can’t occur which inhibits interchain H-bonding and stable triple helix formation
30
What is scurvy?
A defect in post-translation enzymatic activity
31
What is ehlers-danlos syndrome?
EDS
Mutations to type V collagen result in the classic form of EDS, characterized by skiing fragility and extensibility and joint hyper mobility
32
What are collagenopathies?
Inherited collagen synthesis defects
More than 1000 mutations in 23 genes coding for 13 different collagen types
33
What is osteogenesis imperfecta?
OI
Characterized by bones that break easily
Most patients with severe disease have mutations in the gene for type I collagen
Structurally abnormal chains prevent folding of the protein into a triple helical conformation
34
What are the two causes of ehlers-danlos syndrome?
There are 6 variants but all involve a genetic defect in the synthesis (enzymes) OR structure (mutations in the collagen gene)of collagen
EDS can be caused from mutations of collagen processing enzymes or from mutations in the AA sequence of collagen itself
35
What parts of the body are effected with ehlers-danlos syndrome?
Joints
joint hyper mobility: lose joints, dislocations, pain, hyperextended, early osteoarthritis (like you can bend your fingers totally backwards....)
skin
skin hyper-extensibility: fragile skin that tears or bruises easily, scarring, slow and poor wound healing
36
What kind of inheritance are EDS?
Autosomal dominant!!
So only one allele has to be mutated
Recessive means that enzymes require both alleles to be abnormal because 50% of the enzymes can cover up for the other half but with collagen, if half the collagen is mutated then it’s severely structurally weakened
37
What is the specific mutation in Ehlers-Danlos syndrome?
The 5th 3rd Glycine is turned into an Alanine
Gly-X-Y is supposed to be the structure of collagen so if every 5th codon is alanine instead of glycine, it disrupts the hydroxyl bonding that’s needed between the triple helical bonds in collagen
38
What causes osteogenesis imperfecta?
OI is an inherited disease caused primarily by dominant mutations in genes that encode the a1 or a2 chains of type I collagen required for bone formation
Causes bone fragility = problem is too little bone
Can be mild or lethal in utero depending on mutation
Can be either qualitative defects(abnormal collagen I) or quantitative defects (decreased production of normal collagen I)
39
Where are type I collagen fibers found?
Bones, organ capsules, fascia, cornea, sclera, tendons, meninges, and dermis (the whites of the eyes are blue; this is the phenotype that usually presents)
Constitutes about 30% of the human body by weight and is 90% of the protein within the bones
Type I collagen is what’s defective in osteogenesis imperfecta!!
40
What causes quantitative defects in collagen I in OI?
Frequently result from missense mutations in a glycine-coding codon and cause disease due to a dominant negative effect
Glycine in Gly-X-Y is replaced by AA with bulky side chains which mess with bonding in collagen triple-helix
41
What are the two mechanisms of membrane transport?
1. Diffusion
| 2. Active transport
42
What are the types of diffusion (passive transport)?
1. Simple
2. Osmosis
3. Facilitated (passive transport)
A. Carrier-mediated
B. Ion channel-mediated
Downhill, no energy input used, with the electrochemical gradient, favorable process
43
What are the types of active transport?
1. Primary
2. Secondary
Uphill, energy input used (ATP hydrolysis), against the gradient
44
Is Na+ or K+ higher in the ECM or ICM?
Na higher in the ECM and K higher in the ICM
Na get out of here! You’re kicked out into the ECM!
45
Are transporters specific? Which molecules can be transported by more than one mechanism?
Yes they are highly specific ! They’re usually for one molecule or a group of molecules so a single cell has tons of types of transport mechanisms!
Some molecules are transported by more than one mechanism
- Glucose: facilitated diffusion and secondary active transport
- water: osmosis and aquaporins (Facilitated diffusion)
- Na: ion channels, primary and secondary active transport
46
Can steroids free diffuse through the plasma membrane?
Yes they’re small non-polar structures like cortisol and can diffuse freely and don’t require membrane transporter proteins
47
When does diffusion end?
When equilibrium is reached
48
What kinds of passive transport of saturable? Which aren’t??
Simple diffusion doesn’t involve a protein helper it’s just a passage across the lipid bilayer so it is NOT saturable
Facilitated diffusion utilized specific membrane transport proteins so it IS saturable
49
What are the two ways water leaves the cell?
1. Osmosis (type of diffusion)
2. Facilitated diffusion via aquaporins (transporter proteins)
Water is small and not charged so it can pass through the bilayer lipid core at a slow pace (bulk flow) but it can be efficient if there is a high enough solute concentration difference across the membrane
50
What moves water across membranes?
Osmotic pressure
Osmosis is the movement of water across semipermeable membranes - water moves in the direction to equalize osmotic pressure = solute concentration
51
What are isotonic, hypertonic and hypotonic?
Isotonic = solute concentration inside and outside cell are equal (cell doesn’t change)
Hypotonic = less concentrated outside cell than inside the cell (the cell swells because water goes into the cell to try and dilute the inside)
Hypertonic = more concentrated outside the cell than inside the cell (the cell shrinks because water leaves the cell to try and dilute the outside)
52
How is osmosis relevant in cooking?
Pickles! You put them in high salt water and all the water in the pickle leaves and they wrinkle
If you salt an eggplant all the water comes out of it!
53
A man is admitted and is said to be having an ischemic stroke. Intravenous thrombolytic drug treatment was performed. At 24 hours there was no change in neurological deficit. There was also an increase in intracranial pressure. MRA showed persisting ischemia, severe cerebral edema and signs of hemorrhagic infarction. After a decompressive craniectomy and PT, he is better but still has left handed weakness.
In the first days of hospitalization IV administration of which of the following would be an appropriate treatment for the cerebral edema?
Hyperosmotic saline
If the concentration outside the cell is really high then the water inside the cell will leave to try and equalize the concentration gradient
54
A 41 female has consciousness disturbance and seizures. Her sodium is less than normal. She takes chlorpromazine for schizophrenia for 16 years. After hospitalization her Na levels went back to normal and her brain became less swollen. What’s the best explanation for her condition on admission.
Water intoxication
55
Is facilitated diffusion or simple diffusion faster?
Facilitated diffusion is much faster
Facilitated diffusion always accelerates an even that is already thermodynamically favored aka already going down it’s chemical gradient
56
What happens in carrier mediated transport? What are examples?
Transported molecule binds non-covalently with its specific carrier
Binding sites on carriers with the molecule to be transported are very specific/stereo specific but can sometimes bind chemically related solutes
Occurs via limited number of carrier proteins = process is saturable
Can be bidirectional but in practice this is RARE because it’s always downhill with the chemical gradient so the chances of this gradient switching back and forth doesn’t really happen
Ex. Aquaporins, GLUTS
57
What are aquaporins?
Facilitated diffusion of WATER
NOT an ion channel and they’re always open, they don’t open and close
Water moves freely across the plasma membrane to equalize osmotic/solute pressure across the membrane
58
What kind of transporters are GLUTs?
Facilitated diffusion
Conformational change of membrane protein with *noncovalent* glucose binding - can function bidirectionally but always with the concentration gradient
59
What are GLUTs characterized by?
- affinity for glucose
- rate of transport
- tissue expression sites
NOT ENZYMES they’re proteins
GLUT4 is insulin regulated-whether glut4 is on the surface or not is glucose regulated
60
How is GLUT2 an example of bidirectional transport?
If blood glucose is high, GLUT2 receptors on the liver will transport glucose into the liver
If you’ve been fasting and blood glucose is low, GLUT2 receptors in the liver will transport glucose from the liver to the blood stream
In each case though it still follows the gradient!!
61
Nancy has a BMI is 42 and her passing glucose is 510. She is fatigued, always thirsty, lots of vaginal yeast infections, dark and velvet skin folds, non-compliant with medication since her last visit. She has uncontrolled type II diabetes. One of the reasons her blood sugar is high is that some of her cells no longer respond to insulin to take up glucose. How many of the GLUT proteins are insulin-responsive?
One....GLUT 4
62
What best describes transport of glucose by GLUT4?
Transport does not involve hydrolysis of ATP
63
How are ions transported across biological membranes?
1. Ion channels
2. Active transport mechanisms
For ion channels, transport is down the gradient because they’re passive transport
Active transport is used when ions must be moved against their gradient, NOT ion channels
64
What are ion channels?
Integral, membrane-spanning proteins that when open permit the passage of certain ions but not others; usually made of multiple domains but can also be single polypeptide chain
- always with the gradient
- if the channel is open, ions will flow through
- highly selective
- very rapid passage so they need rapid on/off signaling
Ions coming in/out change local signaling so they have
65
What does it mean that ion channels are gated? What are the different types?
Ion channels are usually closed but open briefly in response to a signal then rapidly resets itself to the closed position
Ex. Of signals: voltage gated, ligand gates, mechanically gated, messenger gated
Exceptions are K+ leak channels that are always open
66
Why are ion channels highly selective?
Selectivity filter
All ions exist in a hydrated water shell and the hydrated form is larger than the free ion - initially the size of the hydrated form determines if the ion can attempt to enter the channel
Energetically favorable interactions of the ion with the water shell have to be replaced by favorable interactions with atoms of the channel selectivity filter for ion flow to occur
Only the correct ion has these favorable atomic interactions with the selectivity filter
67
What is a big example of a ligand-gated ion channel?
Acetylcholine receptor
it’s actually an ion channel at the neuromuscular junction where acetylcholine is the ligand
Binding of acetylcholine opens this ion channel allowing for a large influx of Na+ into the cell followed by depolarization of post-synaptic membrane, resulting in action potential leading to muscle contraction
Then acetylcholinesterase is produced to degrade acetylcholine so you get rid of the signal for the opening
Inside of the acetylcholine receptor is leucine residues. These bulky chains get moved away from the center and small polar amino acids get moved to the middle so that ions can move through
This receptor is present on the cell surface, composed of multiple subunits, ions will travel through this protein only down their gradient, has a selectivity filter region on its overall structure
68
What does ligand binding do to receptors in ligand-gated ion channels?
Favorable conformational change in receptor
69
Dysfunction of acetylcholine receptors leads to which disease?
Myasthenia gravis
Autoimmune disease
70
What is myasthenia gravis?
Autoimmune disease
Acquired not inherited because patient develop antibodies that recognize acetylcholine receptor as foreign and they attack it
Impaired binding of acetylcholine or destruction of receptor by autoantibodies
Skeletal muscle weakness with increased activity, eye drooping, mouth drooping
Symptoms can be treated with acetylcholinesterase inhibitors because acetylcholesterase breaks down acetylcholine
71
What causes myasthenia gravis?
Autoimmune disease where acetylcholine receptors in the NMJ are destroyed or there’s impaired binding
Nerve ending usually releases acetylcholine into synaptic cleft but now the antibodies are binding to the receptor that block its function
72
What is the difference between primary and secondary active transport?
If the transporter protein itself hydrolyzes ATP = primary active transport
If unfavorable uphill flow of one molecule is coupled to favorable downhill flow of another = secondary active transport
73
What are the three types of primary active transporters?
1. P-class or P-type
2. ABC type
3. F&V type (bacteria, plants, not covered!!)
74
What are P-class transporters? Give examples
Primary active transporters (also called ATP-Powered pumps)
Ex. Na/K ATPase (all cells)
Ca ATPase (all cells)
H/K ATPase (stomach acid)
Transporter itself hydrolyzes ATP and the transporter is phosphorylated
2 subunits of ONE transporter (alpha subunit usually does the work and gets phosphorylated)
75
What is the most valuable pump?
Na+/K+ ATPase pump = active primary transport
Uses 75% of all the body’s ATP!! Generates the Na/K gradient for:
- establishing and maintaining gradients in all cells
- driving transport of sugars and AA (Secondary transport mechanism)
- maintains membranes potential (-60 mV, negative cytoplasm)
- controls cell volume
76
What is one of the first signs of hypoxia? What happens with hypoxia?
Cell swelling
It occurs early in hypoxia because oxygen is needed for oxidative phosphorylation and the ETC!!! Oxygen is the final electron acceptor in the ETC!
- Hypoxia inhibits ATP production because ETC doesn’t work
- with less ATP, there’s not enough for Na/K ATPase pump
- without the pump working, intracellular Na concentration increases which results in water influx and cell swelling
77
What does the Na/K ATPase pump do?
Pumps Na out against its gradient
Pumps K in against its gradient
That’s why you have to use ATP to make it happen
78
What determines the conformation of the Na/K ATPase pump? What are the steps in the pump?
Phosphorylation of the Na/K ATPase via ATP
3 Na sites and 2 K sites in the transporter
1. 3 Na+ bind to transporter
2. Phosphorylation makes a conformational change from E1 to E2 in the transporter
3. Na+ can be expelled outside of the cell
4. 2 K+ can bind from the outside
5. After K+ binding, dephosphorylation of the pump
6. 2 K+ atoms are pumped inside
79
What cardiac drug targets the Na/K ATPase pump?
Digoxin
Cardiotonic steroid drug that inhibits function of the Na/K ATPase pump by inhibiting the dephosphorylation of the E2P form of the pump (when K+ is loaded in the transporter and ready to be sent inside the cell) —> this locks the pump in a nonfunctional state
What we want to do is increase cytosol Ca+2 in myocytes because this increases heart contractility and function
Cardioglycoside drug inhibit the pump so we get increased Na+ inside the cell too. There’s another transporter called the Na-Ca exchanger (2nd active transport) that exchanges Na and Ca. Normally the exchanger pumps Na in and Ca+2 out but if the Na/K ATPase pump is broken then there’s a buildup of Na in the cell and the Na/Ca exchanger stops pumping Na in/Ca out so there’s a buildup of Ca inside the cell which increases heart contractility and function!
80
What is another important P-type ATPase? What drug accompanies it?
H+/K+ ATPase (gastric) = “proton pump”
Present on parietal cells in gastric mucosa
They pump H+ from the ICF of parietal cells into lumen of stomach
Acidified the gastric contents
Prilosec inhibits this pump which reduces the level of stomach acid and relieves heartburn = proton pump inhibitor
81
Too much of a certain ion in blood due to digoxin is called?
Hyperkalemia
Too much potassium in the blood
Because the Na/K ATPase pump isn’t working so no K+ is getting pumped into the cell
82
What are ABC pumps?
Type of primary active transport pump
ABC = ATP-Binding Cassette
- Distinct from P-type
- transports ions and small molecules
- binds and hydrolyzes TWO ATP
- transporter itself is NOT phosphorylated
Ex. MDR1 and CFTR
83
What is MDR1?
ABC-type pump
Causes drug failure in some chemo therapies in cancer
These pumps are usually present in the liver
Lots of chemo drugs are small planar drugs which diffuse through membrane without transporter (adriamycin, doxorubicin, vinblastine)
MDRI is a type of pump called a drug efflux pump that pumps drugs out of the cell normally (also called P-glycoprotein)
The problem with cancer is that some cells survive and become resistant because MDR1 gene is amplified so there’s 1000’s of copies which means more efflux pumps of the membranes of the cancer cells so they just pump out the chemo drugs ):
84
What causes cystic fibrosis?
Mutation of an ABC-Type chloride channel called cystic fibrosis transmembrane regulator (CFTR)
CFTR is located on apical plasma membrane of epithelial cells and pumps Cl- across the cell membrane
Most common deletion is a 3 base-pair deletion which causes the loss of AA F508 in the CFTR protein
CFTR protein effects exocrine secretions like sweat, digestive and respiratory so abnormal chloride transport leads to abnormal Na transport in all exocrine cells so sweat is extra salty
85
What are the effects of cystic fibrosis?
Pulmonary disease (main cause of mortality)
CFTR protein pump usually pumps Cl- out of the airway epithelial cells which sets up an equilibrium with Na+ entering cells via a different transporter protein (ENaC)
In CF proteins, CFTR doesn’t make it to cell surface so Cl- isn’t pumped out of the cell and it builds up in the cell. Because Cl- is building up, Na+ influx increases through ENac and water follows it to equalize solute concentration. So water is moving out of the mucus in the airway into the cell which causes dehydrated mucus
Prone to serious respiratory infections because mucus isn’t clearing stuff out like it should
86
Common mechanism associated with chloroquine -resistance in malaria patients and multi drug resistance in cancer patients involves what?
An ATPase on the cells (MDR1)
Chloroquine is a treatment for malaria and MDR1 works to pump it out of cells too to prevent it from working
87
What best describes the mutated protein in cystic fibrosis?
The mutation leads to increased Na influx into chronically airway cells through a type of secondary active transporter termed ENaC
88
How much of our ATP does the Na/K ATPase pump use?
75%
89
What’s an example of a secondary transporter?
Transport of glucose into the cell by coupling it with the gradient set up from the Na/K ATPase pump
Glucose goes in against its gradient as Na+ goes in with its gradient
90
What are symporters? Antiporter?
Used for secondary active transport
A and B are the molecules going against the gradient and with the gradient
If A and B are going in opposite directions then it’s antiporter = exchanger
If A and B are going in the same direction then it’s a symporter = co-transporter
B is usually Na
A can be sugars, AA, other ions etc.
91
How do Na and glucose transport function together in enterocytes? What are the 3 transporters and the steps to getting glucose from the intestinal lumen to the bloodstream?
3 transporters:
1. Na/K ATPase (primary active transport)
2. GLUT 2 (facilitated diffusion)
3. Na/glucose symport protein (Secondary active transport)
Steps
1. Na/K ATPase is pumping Na out so low ICF Na+ so now Na can flow down to gradient from the intestinal lumen into the enterocyte and bring glucose with it
2. Now glucose in the enterocyte cell is building up
3. Glucose has now built up a gradient and can go through GLUT2 from the enterocyte to the blood
