Membrane Proteins

Question 0

Describe the mobility of membrane proteins.

Answer 0

Conformational change, rotation, lateral diffusion

Question 1

Describe membrane fractionation

Answer 1

Burst the cell to release cytosol, then centrifuge to form a pellet of membrane.
Gel electrophorese the pellet using SDS-PAGE to separate the different proteins.

Question 2

Describe how mobility of proteins can be restricted

Answer 2

Form rafts (large aggregates) which move slowly in the membrane
Fixed in place if they are tethered or attached to the cytoplasm. 
Avoid cholesterol-rich areas of membrane as they prefer fluidity.

Question 3

Describe two types of haemolytic anaemia.

Answer 3

Hereditary spherocytosis - spectrin depleted by 40-50% so the lattice is not totally formed. Makes the cells less resistant to lysis. To large for capillaries so shearing forces destroy the cells. Cleared by the spleen. Treat with a transfusion.

Hereditary elliptocytosis - defect to spectrin so unable to form spectrin heterodimers. Fragile elliptoid cells formed.

Question 4

Describe how a ribosome and protein become associated with the ER membrane during translation.

Answer 4

Leader sequence formed (hydrophobic)
Signal recognition particle binds the sequence and ribosome. No further translation in cytosol.
Recognised by docking protein
Releases signal to direct through ER membrane
Signal peptidase removes the signal.

Question 5

What sequence stops a protein moving through the membrane during translation?

Answer 5

Stop transfer sequence

Question 6

Describe renal control of circulating sodium in the thick ascending limb.

Answer 6

NKCC2 on the lumenal surface moves sodium, potassium and 2 chloride ions into the cell, utilising a gradient formed by a Na/K ATPase on the capillary membrane.
This creates a gradient which allows KClCT to move potassium and chloride into the capillary. Cl also moves into the capillary through a ClC-Kb channel.
Potassium can move back into the lumen by the K+ channel ROMK.

Question 7

How do loop diuretics work?

Answer 7

NKCC2 in the thick ascending limb.

They decrease osmotic pull and causes more water to be excreted, decreasing blood pressure.

Question 8

Describe how thiazide diuretics work.

Answer 8

Inhibit NCCT in the distal convoluted tubule, decreasing osmotic pull which reduces absorption of water. This lowers blood pressure

Question 9

Describe hyperaldosteronism and it’s treatment.

Answer 9

Primarily used for hyperaldosteronism but can also be a last resort treatment for hypertension.
Aldosterone increases expression of ROMK, ENaC and Na/K ATPase, so in hyperaldosteronism there is high sodium reabsorption and potassium excretion, leading to high blood pressure.
Treat with amiloride which inhibits ENaC and NHE as well as inhibiting the action of aldosterone (potassium-sparing diuretic). Can also use spironolactone.

Question 10

Describe the difference between peripheral and integral proteins.

Answer 10

Peripheral - bound to the membrane surface by electrostatic or hydrogen bond interaction. A salt wash or changes to pH can remove the protein.

Integral - embedded in the membrane with a hydrophilic domain. Can only remove with a solvent or detergent.

Question 11

What is a hydropathy plot used for?

Answer 11

To show hydrophobic/hydrophilic regions of a protein

Question 12

What type of receptor is a nicotinic receptor?

Answer 12

Ligand-gated Na+ channel (ligand is ACh)

Question 13

Describe how an ATP-sensitive K+ channel work and give a cell which it can be found in.

Answer 13

ATP binds, blocking the channel. This causes K+ to build up in the cell, increasing the membrane potential.
Found in beta cells of the pancreas

Question 14

Describe how voltage-gated channel proteins work

Answer 14

The inside of the protein is more negative, and has a voltage sensor. A change in potential causes the voltage sensor to be driven outwards, which drives the conformational change.

Question 15

Describe the role of PMCA

Answer 15

A plasma membrane calcium ATPase. Hydrolysis of ATP drives a conformational change (primary active transporter), moving calcium into the cell. They are high affinity but low capacity.

Question 16

Describe the role of F1F0 ATPase

Answer 16

Uses the hydrogen gradient on the inner mitochondrial membrane for ATP synthesis. Is essentially a primary active transporter in reverse

Question 17

Describe the two types of cotransporter

Answer 17

Symport - both in the same direction

Antiport - opposite directions

Question 18

Describe the Na/K ATPase

Answer 18

Primary active transporter, antiport. Moves 3Na into the cell and 2K out. Is a p-type ATPase so ATP phosphorylates aspartate in the protein to induce a conformational change.
Alpha subunit - K/Na/ATP binding sites. and ouabain (inhibitor)
Beta subunit - address. glycoprotein on the surface.
Used to generate Na and K gradients
Controls pH, regulates cell volume and calcium, absorbs sodium into the epithelia, nutrient uptake (e.g. glucose)

Question 19

What type of transporter is mainly responsible for membrane potential?

Answer 19

Potassium channels

Question 20

Describe NCX

Answer 20

Sodium-calcium exchanger.
Antiport, secondary active transporter.
Uses the inward sodium gradient from Na/K ATPase for the extrusion of calcium. (3Na in per sodium)
Low affinity and high capacity so it’s good when there is high calcium in the cell (protective)
Has a role in cell toxicity in ischaemia and reperfusion injury. In ischaemia, low ATP inhibits Na/K ATPase so sodium accumulates, causing depolarisation. Calcium also accumulates, which is toxic.
If a cell depolarises with high intracellular sodium the protein reverses

Question 21

Describe NHE

Answer 21

Sodium-hydrogen exchanger.

Secondary active transporter for the extrusion of hydrogen. Antiport.

Question 22

Describe Na/glucose transporter

Answer 22

Found in the intestines.

Symport, secondary active transporter

Question 23

Describe how protein kinase A can cause diarrhoea

Answer 23

It enhances the rate of Cl- transport by CFTR, which increases water in the intestine

Question 24

Describe how transporters work together in the intestinal wall to maintain ion concentrations to allow CFTR to extrude Cl- into the gut lumen.

Answer 24

NKCC2 brings Na, 2Cl and K into the epithelial cell with the help of a Na/K ATPase.
A potassium channel allows K to be removed from the cell to keep the gradient for NKCC2 to move K+ in.
This raises the Cl- concentration in the cell which builds a gradient for CFTR to move it into the lumen.

Question 25

Describe SERCA

Answer 25

Smooth endoplasmic reticulum calcium ATPase
Primary active transport, symporter.
Moves calcium into the cell and hydrogen ions out.

Question 26

Where are calcium uniporters found?

Answer 26

In the mitochondria

Question 27

Give a protein in the cytoplasm that binds to calcium to buffer it.

Answer 27

Calmodulin

Question 28

Describe how mitochondria can act as a calcium buffer

Answer 28

They have a membrane potential of -160mV and calcium uniporters so calcium moves down the electrical gradient.

Question 29

Describe a protein which is an acid extruder.

Answer 29

NHE - sodium hydrogen exchanger
Electroneutral, regulates volume and pH, found in most cells. Stimulated by growth factors and inhibited by amiloride.

NBC - sodium bicarbonate cotransporter.
Alkalinises. Involved in cell volume regulation and only found in some cells.
Moves Cl- and H+ out of the cell, Na+ and HCO3- into it.

Question 30

Describe a base extruder.

Answer 30

AE - anion exchanger
Acidifies and for cell volume regulation
Moves Cl- into the cell and HCO3- out.

Question 31

Describe bicarbonate reabsorption in the kidney and where in the nephron it occurs.

Answer 31

Proximal tubule.
In the lumen, sodium bicarbonate releases a bicarbonate ion which binds to two hydrogen ions. Hydrogen bicarbonate is then converted to water and carbon dioxide by carbonic anhydrase. They can then diffuse into the epithelia where it is converted back to hydrogen bicarbonate which dissociates to hydrogen and bicarbonate.
NHE on the lumenal side extrudes hydrogen and brings Na into the epithelial cell. The gradient for this is from both a Na/K ATPase on the capillary wall, and dissociation of hydrogen bicarbonate to release H+