W2&3 Membrane Transports + Na/K ATP-ase (Dustin) Flashcards
The phospholipid bilayer is most permeable to _______ and least permeable to ______
The phospholipid bilayer is most permeable to hydrophobic molecules (O2, H2O, H2) which freely diffuse. Least permeable to ions (Cl-, K+, etc)
Simple diffusion is what?
Transport through a membrane driven by concentration gradient, no transporters.
Must be lipid-soluble
Facilitated diffusion is what?
Transport driven by concentration gradient or electrochemical gradient, but a transporter molecule assists. Still, it doesn’t require energy
Primary active transport is what?
primary active transport needs energy to go against concentration or electrochemical gradient
as in Na/K-ATPase
What is secondary active transport?
secondary active transport uses concentration gradient made by primary active transport to drive further movement
usually involves sodium gradient created by the Na/K-ATPase
P-type ATPases are what?
A transport ATPase (active transporter),
“P” for “phosphorylation, they are reversibly phosphorylated by ATP (on an Asp residue) as part of the transport cycle
Most ATPases are P type
These are integral plasma membrane proteins
What are V-type ATPases?
A transport ATPase (active transporter),
located within the cell in the membranes of vacuoles/vesicles
Function is to create a proton gradient (proton pumps),
which may serve some other kind of transport that is done secondarily or acidify lysosomes etc.
What are F-type ATPases?
A transport ATPase (active transporter),
Active in mitochondrial inner membrane
Also a proton pump. Includes ATP synthase.
What does the ABC in ABC transporters stand for?
What are they?
A transport ATPase (active transporter),
ATP-binding casette transporter
Make specific transporters in the membranes responsible for the transport of cholesterol, bile acids, and many other things against a concentration gradient. Responsible for extruding some of the foreign molecules from cell, i.e. some drugs
The multidrug transporter MDR1 aka P glycoprotein is responsible for resistance against anti-tumor drugs
With P-type ATPases there are 4 domains mentioned in the alpha subunit
What are the functions of these domains:
T
N
P
A
- T: transport. Could also see it as “t” for transmembrane; its conformation enables some molecule to be transported through the protein
- N: ATP/ADP binding
- P: phosphorylation. Contains the Asp residue that is phosphorylated
- A: phosphatase activity
Note that the beta subunit projects extracellularly (not seen in image)
SERCA ATPase and K, H ATPase are examples of what type of ATPase?
P-type
Where is the K, H ATPase?
What does it do?
Stomach parietal cells
Makes for potassium reasborption, H excretion
Helps create the acidity of the stomach
What does SERCA stand for?
There are 3 isoforms of it, where are they?
(I doubt the test will be detailed enough to require you to correctly match them)
SERCA = Smooth Endoplasmic Reticulum Calcium ATPase,
Fills the sarcoplasmic reticulum with cytosolic calcium
- SERCA1: striated muscle
- SERCA2: smooth muscle, striated muscle, heart muscle (all types) - phospholamban inhibits it
- SERCA3: platelets, endothelial cells, other non-muscle
What does PMCA stand for?
What are PMCA ATPases?
PMCA = Plasma Membrane Calcium ATPase
These are P-type ATPases that remove calcium from the cell
What is the ratio of sodium-potassium transported by Na/K ATPase, and in what direction are the ions transported?
This pump is responsible for some of the negative resting membrane potential of a cell, what is that feature of the pump called?
3 Na+ out, 2 K+ in
This makes it an “electrogenic” pump
What are the approximate values of
intracellular and extracellular Na+, K+, and Ca2+
- K+: i.c. = 140 mM, e.c. = 5 mM
- Na+: i.c. = 10 mM, e.c. = 140 mM
- Ca2+: i.c. = 100nM, e.c. = 2 mM
What is the function of intracellular calcium level being so low?
Calcium is used for signal transduction, where a small increase in calcium can lead to muscle contraction, neurotransmission, etc.
Describe the postulated mechanism for the Na+/K ATPase
- Starts in E1 position. First the 3 Na+ bind intracellularly, then ATP is hydrolyzed
- Conformational change from E1 to E2 form
- the 3 sodium molecules now face the extracellular side, then diffuse away
- 2 Potassium from the outside can now fit inside the pump, which then loses its phosphate
- Return to original E1 conformation as K+ is released in the cytosol
In which position is Na+/K+ ATPase when inhibitors bind to it?
Which position binds better to ATP?
E2 position (open facing extracellularly for potassium) binds to the inhibitor Ouabain
E1 position has high affinity for ATP
With the SERCA pump, how does it work in comparison to the Na+/K+ ATPase?
What are the important domains?
Mostly similar
- There is a transmembrane domain with 2 Ca2+ binding sites internally
- Has a nucleotide binding domain with the ATP-binding site
- Phosphorylation domain with similar Asp binding site
- Actuator domain
What is the function of the beta subunit of Na+/K+ ATPase?
The beta subunit is mostly extracellular with only one transmembrane segment (there are 10 TM segments in alpha subunit, which is also where all the action of Na/K ATPase takes place)
The beta subunit stabilizes the alpha subunit, which involves the highly glycosylated structure of the beta subunit that is not able to be pushed around the membrane as easily as the alpha subunit.
What is the function of the FXYD subunit of Na+/K+ ATPase (sometimes called the gamma subunit)
Regulates the transport kinetics of the alpha subunit, adding to the diversity of how the different isoforms of Na+/K+ ATPases throughout the body function (Essentially, creates tissue-specific sodium potassium pumps)
The N-terminal segment has a conserved Phe-X-Tyr-Asp motif (hence the shorter amino acid abbreviations of FXYD)
Particularly important in heart and kidney
What are the therapeutic effects of Ouabain/ Strophantine/ digitalis?
(many names to be familiar with for this compound from the foxglove plant)
This is a cardiac glycoside and the most important inhibitor of Na+/K+ ATPase
- Slight inhibition of sodium potassium pump in the heart muscle -> increase in intracellular sodium (bc it isn’t pumped out)
- Increase in intracellular sodium means the sodium-calcium exchanger reverses direction, bringing calcium into the cell
- Increased intracellular calcium makes for more forceful contractions (increased ionotropy)
The heart’s sodium-potassium pump contains alpha 2 subunits, which are more sensitive to Oubain/ Strophantine than the other types of alpha subunits, which are not expressed as much in other tissues
Why does the structure of Ouabain look so similar to steroid hormones when it is produced by plants?
Because the body produces an endogenous form of ouabain, which the plant has mimicked because it works as a protective toxin. The endogenous form is released by the zona fasciculata and increases vascular tone -> increased blood pressure
Na-K ATPase is regulated by intracellular _____ concentration, which is sensed by Na-K ATPase and initiates the cycle
Na-K ATPase is regulated by intracellular sodium concentration, which is sensed by Na-K ATPase and initiates the cycle
The potassium binding site is saturated at physiological levels
How much of total ATP consumption goes to the sodium potassium pump?
How much in neurons?
In severe hypoxia, what happens to it?
30% of total
50% in neurons
Even in severe hypoxia, it keeps working
What is unique about the heart’s FXYD subunit of its sodium potassium pump?
What other name does it have?
This is FXYD1, which = phospholemman
When dephosphorylated, it decreases the sodium affinity for the alpha subunit
Beta1 adrenergic receptor stimulation (Gs) -> PKA -> phosphorylation of phospholemman -> intracellular sodium and calcium decrease -> prevention of arrythmia, protective of the heart
What is unique about the kidney’s FXYD subunit of its sodium-potassium pump?
This is FXYD2
This creates a moderate increase of Na/K-ATPase ATP affinity in the kidney medulla, which is useful because the medulla has lower oxygen concentrations + thus less ATP.
(Increase is moderate because too-high affinity would use up too much of the already low medullary ATP supply)
What is the difference between short term and long term regulation of the sodium potassium pump by hormones?
Short term -> direct effects or changes in translocation of the Na+/K+ ATPase
Long term -> de novo synthesis of Na+/K+ ATPase is influenced
What are the effects of aldosterone on Na+/K+ ATPase?
Long term effect: increase expression of Na+/K+ ATPase
Short term effect: increase activity, translocation to plasma membrane, and affinity to sodium
If there is decreased sodium intake, then aldosterone attempts to keep as much sodium as possible in the body by increasing sodium reabsorption
(Remember that Na+/K+ ATPase is always on the basolateral side in the nephron. Aldosterone increases the intake of sodium on the luminal side, which is then reabsorbed by Na+/K+ ATPase on the basolateral side)
What is the effect of dopamine on Na+/K+ ATPase in the kidney?
Inhibits Na+/K+ ATPase in the proximal tubules and collecting tubules, leading to decreased sodium reabsorption
Important when there is high salt intake
What are the effects of epinephrine and norepinephrine on Na+/K+ ATPase?
What happens with epi in skeletal muscle?
With norepi in the kidney and brain?
Stimulation of Na+/K+ ATPase
Epinephrine in skeletal muscle helps reuptake more potassium, reducing hyperkalemia after exercise
Norepinephrine in the kidney is a dopamine antagonist, which allows for more sodium reabsorption, while in the brain it helps reestablish the ion gradients after a nerve impulse
What are the effects of insulin on Na+/K+ ATPase?
Short term and long term
Short term: stimulation, translocation to the plasma membrane in slow-twitch muscles
Long term: complex, either increased or decreased expression
Glucose uptake, amino acid uptake, the sodium-calcium exchanger, choline and neurotransmitter uptake, and the sodium-proton exchanger are all able to transport because of what?
Thus, what can inhibit them all?
Secondary active transport via co-transport with sodium
Thus they can be inhibited with the Na+/K+ ATPase inhibitors, like ouabain
What is the relationship between the different SGLT’s in the kidney loop?
Sodium Glucose Transporter 2 (SGLT2) is more proximal because it has a lower affinity to glucose, yet has a larger capacity. This is able to absorb most of the glucose in the tubule. More distally is the SGLT1 transporter, which has a higher affinity and lower capacity, which absorbs the remaining low-concentration glucose.
~100% of glucose is typically reabsorbed, except in very hyperglycemic states
The sodium proton exchanger can be found where?
What is its role?
Found in virtually all cells, functions to reduce the proton concentration (raise pH)
It’s particularly important on the luminal side of the kidney collecting duct, where it helps regulate pH balance of the body
What is a normal pH value inside of a vesicle that contains neurotransmitters?
How does it get this way?
What is the function of this pH?
~6
Uses a proton ATPase to pump in hydrogens using 1 ATP
This means that there is a proton gradient built up that can be used to drive in the neurotransmitter
How many nucleotide binding domains do ABC transporters have?
How many transmembrane domains?
2 NBD’s
2 TMD’s
The two TM domains alternate between an inward and outward facing orientation, and the alternation is powered by the hydrolysis of ATP
What is the ABCA1 transporter, and what’s its significance?
Functions in reverse cholesterol transport, from organs to HDL (helps reduce cholesterol)
What is the ABCB1 transporter, and what is its function?
aka MDR1 - multidrug resistance transporter
It transports lipophilic compounds, providing protection against toxins
It may have increased expression in tumor cells, making them harder to kill with drugs
Different PMCA (Plasma Membrane Calcium ATPase) types:
Where do you find PMCA 1, 2, 3 and 4?
PMCA 1 general
PMCA 2 neuronal - higher affinity for cAMP than PMCA 4
PMCA 3 striatal muscle
PMCA 4 general
