Structure, Function and Pharmacology of the Na+/K+ ATPase pump Flashcards
- Describe what is the pump-leak hhypothesis - Recall evidence that the Na pump is an ATPase - Describe what energises the pump - Describe the nature of the cation transport system - Describe the pump reaction mechanism - Define the five operational modes of the pump - Recall the molecular structure of the Na pump, tissue distribution of pump isoforms and pharmacological interactions with the alpha1 subunit
What is the Na-K ATPase pump
- member of the P-type ATPases
- pumps Na into the cell and K out of the cell
What does an electrochemical gradient do
critical for maintaining osmotic balance and a stable membrane potential, and the excitable properties of muscle/nerve cells
What does the Na+ gradient do
provides the energy that fuels Na-coupled transporters mediating the translocation of ions and substrates across the plasma membrane
Role of Na-K-ATPase pump in the kidney
- drives the reabsorption of Na+ and water
- essential in maintenance of body fluid and electrolyte homeostasis
How can cation gradients be abolished
- cell death
- withholding glucose
- use of specific inhibitors
What is the pump-leak hypothesis
- active extrusion of Na and uptake of K ions is balanced by a passive leak of these ions in opposing directions
- transmembrane potential exists -> Na takes place against chemical and potential gradients
Proof that the movement of Na and K across the membrane requires metabolic energy
cation transport is still active in red blood cells without oxidative metabolism -> suggests ATP rather than glycolysis/Kerb cycle events energise pump
How does the Na/K-ATPase pump work
- can be stopped at any time and is reversible
STEP 1: affinity of sodium for three binding sites
STEP 2: phosphorylation occurs
STEP 3: conformational change alters protein shape and diminishes affinity of sodium for binding site
STEP 4: potassium binding sites exposed -> high affinity for potassium
STEP 5: hydrolysis, detachment of phosphate group from residue
What is the transition step
transport step -> ratio of Na release and K binding corresponds to the ratio of 3 Na ions expelled for 2 K ions taken up
What does the Na pump do
- helps maintain normal trans-membrane electrochemical gradients of membrane permeability properties
- maintains stable membrane potential
- energy in gradient used to drive secondary active transport systems for sugars and amino acids
What does the catalytic subunit do
- possesses binding sites for Na, K, ATP and Mg and cardiac glycosides
- intrinsic ATPase activity
- 4 isoforms
- molecular szie ~ 112kDa
- developmentally regulated and expressed in tissue-specific manner
What does the regulatory subunit do
- three isoforms
- heavily glycosylated (28% w/w)
- crucially required for full enzyme activity, but also for enzyme assembly, intraacellular transport, and stability of alpha subunit
What is the small auxillary protein
- belongs to the FXYD protein family - named after invariant motif
- single-spanning membrane peptide
Evidence for multiple isoforms of the Na,K-ATPase
- different curve in brain and kidney, as sensitivity to cardiac glycoside are different when comparing tissues
- tissue specific antibodies against purified enzyme
- biphasic curve indicates drug binding to receptor
Expression on alpha 1 subunit
ubiquitous expression -> contains binding site for drugs such as digoxin
Expression on alpha 2 subunit
excitable tissues / insulin responsive tissues
Expression on alpha 3 subunit
excitable tissues
Expression on alpha 4 subunit
only expressed spermatozoa
Use of Digoxin clinically
- treat cardiac arrythmias
- narrow therapeautic window
- inhibitor of Na/K-ATPase pump
- binds to alpha subunit
- increase in intracellular Na
- reduced action of Na/Ca exchanger
- increased intracellular Ca in SR
- Ca release during cardiac AP, increasing force of contraction
Mechanism of action of Potassium-sparing diuretics
- block Na reabsorption via ENaC leading to Na excretion
- reduced Na reabsoprtion leads to reduced K excretion
What does the alpha subunit of Na-K pump do
ATP binds to alpha subunits and causes conformational change, allowing for closure so that the opening of cavity faces extracellular compartment
What is the role of the beta subunit of the Na-K pump
generally glycosylated
