2 - Membrane Permeability (and Ion Transporters) Flashcards
What is a semi-permeable membrane
a layer through which only allowed substances can pass
- Pores, channels and transporters contribute to this
- Hole needs to be big enough for molecule to pass through
- Charge / potential difference needs to allow for molecule to pass
- Occurs until solute equilibrium has been reached
Black film + membranes
not on ILOs
- Phosphatidylserine (lipids) mixed + spread onto glass or plastic plates with pinhole
- Submerged into hydrophilic solution
- Phospholipid bilayer formed in pin hole
- Appears black because light is reflected back
Relative permeability of a synthetic lipid bilayer to different classes of molecules
- small gases / small uncharged eg O2, CO2, N2, H2O + urea. These pass straight through the membrane, and then back out again
- large uncharged polar molecules / ions eg glucose, sucrose, K+. Cannot diffuse through the membrane easily and therefore need a transport mechanism.
- Therefore can estimate permeability of a membrane
What is passive transport dependent on (Fick’s Law – equation will be given in exam)
J = P (C1 – C2)
Where J = net rate of transport (ie cm/s)
P = permeability coefficient (dependent on distance travelled + viscosity of medium)
C1 and C2 = concentration gradients on side 1 and 2
passive transport is dependent on permeabilty and concentration gradient
☞ rate of passive transport increases linearly with increasing concentration gradient
How is water transported through the phospholipid bilayer
passive diffusion aka osmosis
- Over entire surface of cell membrane
- Bi-directional flow
- Water crosses to reach solute equilibrium
- This is the main method of transporting water in and out of erythrocytes
aquaporins
- Integral membrane proteins
- Bi-directional flow
- Reaches equilibrium quickly
Passive vs active transport
passive = no energy required as long as there is a concentration gradient. Solute moves by diffusion down concentration / electrical gradients
active = cellular energy required. Solute moves across membrane against it’s concentration gradient.
Non-gated and gated pores
non gated eg connexins. Pores… they are always open.
gated are conditionally open. These are channels. Movement of molecule is due to its interaction with the protein → conformational change
Both have hydrophilic centres.
The CFTR transporter in cystic fibrosis vs diarohea
☞ both involve the CFTR (cystic fibrosis transmembrane conductance regulator transporter)
☞ in cystic fibrosis, CFTR not working → no Cl- outside cell → no water moving out with Cl- → produces sticky, thick mucus
☞ in diarrhoea, protein kinase A phosphorylates CFTR → loads of Cl- being pumped out of cell → loads of water moving with it → diarrhoea
Facilitated diffusion via gated channels (and examples)
- Can be ligand gated where a molecule like ACh binds → conformational change → allowing Na+ to flow through (nicotinic acetylcholine receptor in neuromuscular junction)
- another ligand gated ion channel is when ATP binds (not hydrolysed, just binds) → conformational change → closes channel → K+ cannot flow out of cell (ATP-sensitive K+ channel in insulin-secreting beta cell)
- Can be voltage gated where a membrane depolarisation occurs → sensed by voltage sensor → conformational change → opens channel → Ca2+ can flow into cell (Ca2+ channel)
Facilitated diffusion via carrier proteins
- Sequential binding and conformational change
- Allows different control mechanisms
- Limited number of substances can be moved (reaches saturation point)
- Facilitated diffusion is usually much slower, depending on number of carrier proteins at membrane
- Central pore is usually aqueous
Types of carrier (transport proteins)
one substance being transported
☞ uniport
coupled transport (co-transport)
☞ symport = two or more substances moving in the same direction
☞ antiport = two or more substances moving in opposite directions
Uniport vs co-transport
uniport
- Only one molecule transported at a time per cycle down the concentration gradient
- Can be a channel or a carrier protein
- Sometimes no energy needed (just kinetic energy of particles) eg GLUT3
- Sometimes ATP is needed to transport substance against it’s concentration gradient (eg proton pump)
co-transport
- Usually two or more substances transported on a membrane transporter per reaction cycle
- One substance could be moving down its concentration gradient, whereas the other might be carried against its concentration gradient
Symporter vs antiporter
symporter = co-transport, where two or more substances are being carried in the same direction
antiporter = co-transport, where two or more substances are being carried in different directions
What are some of the general physiological roles of transport processes
- Maintenance of ionic composition
- Maintenance of cellular pH
- Regulation of cell volume
- Concentration of metabolic fuels + building blocks
- Expulsion of metabolic waste products and toxic substances
- Generation of ion gradients necessary for the electrical excitability of nerve and muscle
What is active transport
- the transport of ions or molecules against a concentration gradient and/or electrical gradient
- Energy directly (or indirectly) comes from ATP hydrolysis
- Can be primary (energy derived directly from breakdown of ATP) or secondary (co-transporters aka symporters)
What is the difference between primary and secondary active transport
primary directly uses a source of chemical energy (eg ATP) to move molecules against their concentration gradient
secondary (aka co-transport) uses an electrochemical gradient (generated by primary active transport) to move molecules against their gradient, and therefore doesn’t directly require a chemical source of energy such as ATP
What are the primary active transporters
- Sodium potassium pump
- Calcium ATPases (PMCA and SERCA)
- Potassium/proton ATPase, aka proton pump
- F1F0 ATPase (aka ATP synthase)
these can also be used to set up an electrochemical gradient for secondary active transport
Sodium-potassium pump
3Na+ bind → conformational change → allows ATP to bind → CC → Na+ released out of cell → CC → allows 2K+ to bind → K+ released into cell
- Key concept: 3Na+ out for every 2K+ that enter
- Very important for generating action potential
- Driven by conformational change
- Antiporter
- Consists of an α and β subunit (more detail on next card)
- Every cell has one of these
What do the α and β subunits of the sodium-potassium pump do
α has K+, Na+ and ATP binding sites
β is responsible for directing pump to cell surface membrane
What are the main functions of the sodium-potassium pump
forming Na+ and K+ gradients which are necessary for electrical excitability
driving secondary active transport
- pH control
- cell volume regulation
- absorption of Na+
- nutrient uptake eg glucose
Free ion distribution across the cell membrane
- there are a number of cations and anions present in cells + outside the cell membrane
- this means that there is a concentration gradient present
- these concentrations need to be carefully controlled in order to ensure proper function of the membrane
- K+ there is a much greater concentration of this inside the cell than out. Too low = coma. Too high = cardiac arrest.
- Ca2+, Cl- and Na+ all have a greater concentration outside the cell than in.
- Ca2+ is toxic to cells in high quantites, due to the high phosphate concentration inside cells → form calcium phosphate → calcified cells
Ca2+ - ATPase (aka PMCA)
excludes Ca2+ ions from cell
- Does this by ATP → ADP + Pi (inside cell)
- Helps control the [Ca2+] with other transporters
- Uniporter
Control of resting Ca2+ concentration (in general)
primary active transport
- PMCA (expels Ca2+ out of cell, removing residual Ca2+ as high affinity + low capacity)
- SERCA (accumulates Ca2+ into the SR/ER as high affinity + low capacity)
secondary active transport
NCX (low affinity + high capacity so removes most Ca2+)
facilitated diffusion
Mitochondrial Ca2+ uniports (operate at a high [Ca2+] to buffer potentially damaging high concentrations)
Sodium-calcium exchanger (NCX)
one Ca2+ out for every 3Na+ in
- Works with PMCA primarily to control [Ca2+]
- Antiporter
- Activity is membrane potential dependent (depolarised membrane potential reverses mode of operation)
- Ie when inside cell is more positive, 3Na+ out and one Ca2+ in
What is the main difference between NCX and PMCA
both control [Ca2+] in the cell
NCX is low affinity, high capacity (doesn’t bind to calcium well but can transport it very fast). Antiporter
PMCA is high affinity, low capacity (binds a small amount of calcium well). Uniport
F1F0 – ATPase (ATP synthase)
- Sits between the inner + outer membrane (intermembranous space) in mitochondria
- Active transport in reverse
- For every 3H+ pumped from intermembranous space → mitochondrial matrix, one molecule of ATP is formed
- ADP + Pi → ATP (in matrix)
- The H+ are from the electron transport chain
- Cause conformational change
Secondary active transporters
- Co-transporters
- Usually using an electrochemical gradient already set up by primary active transport
- symporters in same direction, antiporters move two or more substances in opposite directions
- Usually electroneutral (no change in electrical charge across the plasma membrane)
Definition: electroneutral
No change in electrical charge across the plasma membrane
How many Na+ does SGLT1 and SGLT2 move?
SGLT1 moves 2
SGLT2 moves 1
(from extracellular space → cytosol)
Mechanism of ischaemia (involving NCX and Na+-K+-ATPase)
sodium-potassium ATPase
ATP is depleted → sodium pump inhibited → less Na+ being pumped out of cell → more Na+ accumulates inside cell → cell is depolarised (ie becomes more positive inside cell than out)
NCX
Depolarised cell is energetically unfavourable → NCX reverses → now 3Na+ out and one Ca2+ in → this means more Ca2+ is entering the cell → high Ca2+ is toxic
Na+/H+ exchanger (NHE)
one Na+ into cell, while one H+ out
- Works using Na+ gradient set up by sodium-potassium pump
- Electroneutral due to 1:1 ratio
- Antiporter
- Regulates pH and cell volume
- Activated by growth factors
- Can be inhibited by amiloride which is a potassium-sparing diuretic
Control of cell pH
- Na+-K+-ATPase is in all cells, creates an Na+ gradient for other transporters
- NHE involved in Na+ and H+ exchange (acid extrusion)
- NBC Exchanger is also acid extrusion, and alkali influx
- **sodium-bicarbonate co-transport* is involved in alkali influx
- anion exchanger (aka band 3 in erythrocytes) are involved in alkali extrusion
Bicarbonate transporters (NBC + AE) – details on separate cards
- NBC aka Na+-bicarbonate-chloride co-transporter alkalinises cell
- AE aka band 3 in erythrocytes acidifies cell
- Both are involved in cell volume regulation
NBC (sodium-bicarbonate-chloride exchanger)
- One Na+ and one HCO3 in
- One Cl- and one H+ out of cell
- Alkalinises cell
- Involved in both acid extrusion + alkali influx
- Also involved in cell volume regulation
Anion exchanger (AE)
- Aka band 3 in erythrocytes
- One HCO3- out and one Cl- in
- Acidifies cell by alkali extrusion
- Antiporter
- Electrogenic due to 1:1 ratio
Sodium-bicarbonate co-transport
- 3HCO3- and one Na+ into cell
- Involved in cellular pH regulation
- Alkali influx into cell
Sarco(endo)plasmic reticulum ATPase (SERCA)
- One H+ from inside SR/ER → intracellular fluid
- One Ca2+ from intracellular fluid → SR/ER
- One ATP → ADP + Pi (intracellular fluid)
- This is the equivalent of the PMCA but on the sarco(endo)plasmic reticulum
- Responsible for replenishing Ca2+ stores in the SR/ER
- Antiporter
Hydrogen-potassium ATPase
- One H+ into gastric lumen, where one K+ into cytoplasm of parietal cell
- Antiporter
- Responsible for acidification of stomach contents
- Found in parietal cells (highly specialised epithelial cells located in the inner stomach lining)
- electroneutral due to 1:1 ratio, so no effect on membrane potential
Mitochondrial Ca2+ uniporters
- Pump Ca2+ into mitochondrial matrix from the intracellular fluid
- Form of facilitated transport
- Used at high [Ca2+] to buffer damaging Ca2+ concentrations
Why is high [Ca2+] damaging to cells
- There is a high phosphate concentration inside cells
- Calcium phosphate formed
- Leads to calcified cells
- May end up interfering with organ function or cause other health issues eg cancer
Km is half of Vmax – how do we make this estimate more accurate
Use a Lineweaver-burk plot
(it’s a graphical representation of enzyme kinetics)
Glucose transport mechanisms – uptake in the gut
uses SGLT-1 and GLUT2
- Microvilli express SGLT1 (co-transporter of Na+ and small-no polar molecules)
- Therefore Na+ drawn into enterocyte with glucose from the ileum lumen
- Na+ recycled using sodium-potassium ATPase
- Some of the glucose remains in the enterocyte to be used
- 60% of the glucose is transported immediately to interstital space, and then into blood by GLUT2
- A small amount of the glucose slowly leaves by passive diffusion (glucose can cross membranes, but extremely slowly)
Glucose absorption using H+
- Hydrogen-potassium ATPase (found in parietal cells of the inner stomach lining) are responsible for acidification of the stomach contents
- H+-K+-ATPase pump K+ into the cytoplasm of parietal cell, and H+ into the gastric lumen (antiporter)
- This creates a proton differential
- Can now use sucrose-H+ symporter
- Proton and glucose transported into enterocyte simultaneously
- Sucrose → glucose + fructose
- Glucose transported into blood using GLUT2
Glucose re-absorbtion in the kidney
the proximal convoluted tubule has 3 segments
☞ to get glucose from PCT → epithelial cell
in segment 1, SGLT2 + 1Na+ used
in segment 2+3, SGLT1 + 2Na+ used
☞ to get glucose from epithelial cell → bloodstream
In segment 1, GLUT2 used
In segment 2+3, GLUT1 used
Na+ ions are recycled using Na+-K+-ATPase
What are the GLUTs used to transport glucose in liver, muscle and adipose
GLUT2 + GLUT4
Insulin and its receptor regulate GLUT synthesis and intracellular transport
What are MCT proteins
Monocarboxylate transporters
Symporters that transport H+ ion and lactate ion (-) from astrocyte to neurone in CNS
Glucose transport in the CNS
☞ Pathway 1 key molecules = GLUT1, MCT1, MCT2 and MCT4
Glucose into cell by GLUT 1 → glucose converted to lactate (via pyruvate) by astrocyte → lactate transported to neurone by MCT
☞ Pathway 2 key molecules = GLUT1, GLUT3, MCT1 and MCT4
Glucose transported by GLUT1 (in apical + basal surface of epithelial cell) to interstitial space → glucose transported from interstitial space to neurone by GLUT3 in neurolemma
(in both pathways, glucose → G6P so it’s trapped inside cell)
In general, what is the mechanism of GLUT
facilitated diffusion
Insulin promotes GLUT containing vesicles from cytoplasm → plasma membrane
This is an example of regulated secretion
☞ glucose transport
In general, what is the mechanism of SGLTs
secondary active transport
Na+-K+-ATPase generates the sodium electrochemical gradient that drives the transport of glucose
