Lecture 5: Membrane Function 2 - Membrane transport, channels

Question 1

Learning objectives

Answer 1

• Understand the selectivity mechanisms of channels
• Understand the concept of gating
• Appreciate the molecular mechanisms of different channels
• Aquaporins and K+ channels

Question 2

What is a selectivity filter?

Answer 2

The narrowest point of a pore which determines which molecules can pass through. The size and charge of the ion/molecule is important.

Question 3

Give some of the characteristics of channels.

Answer 3

• narrow and highly selective
• Open and close rapidly (very responsive to environmental changes)
• Transport inorganic ions
• Passive transport (no energy required)
• Much, much faster transport than transporters

Question 4

What kinds of gating are there?

Answer 4

• Voltage-gating
• Ligand-gated (intracellular/extracellular)
• Mechanical-gating (responds to pressure change)

Question 5

How does voltage-gating work?

Answer 5

If the charges are changed or reduced, the channel acts to rectify this back to resting state by opening the pore.

Question 6

How does mechanical-gating work and what is the purpose?

Answer 6

If there is a change in pressure, the pore is opened. This prevents rupture of the membrane.

Question 7

How does ligand-gating work?

Answer 7

Ligand gating channels (also called receptors) have a binding site on one surface (cytosolic or internal side). When a ligand binds to the binding site, the channel opens. These receptors are specific to the ligand that opens them and the molecule/ion that they transport once opened.

Question 8

What can also activate specific channels?

Answer 8

Protein phosphorylation

Question 9

Give two important applied roles of ion channels in organisms.

Answer 9

• Electrical excitability of muscle cells
• Leaf closing response
  Both of these require very rapid cellular response

Question 10

Which are the commonest ion channels and what is their most important role?

Answer 10

K+, potassium ion, channels, which are very important in maintaining membrane potential.

Question 11

What does an aquaporin transport?

Answer 11

Water. Aquaporins are responsible for water secretion in tears, sweat and saliva and water reabsorption in the kidneys.

Question 12

How many different aquaporins are there in a) mammals and b) the model plant organism, A. thaliana? Explain the difference.

Answer 12

a) 11
b) 38
The different aquaporins have different functions in different parts of the body.
Plants have more aquaporins because they need better water regulation as they cannot move to fins water like animals can.

Question 13

Discuss the flow rate and direction of aquaporins.

Answer 13

Aquaporins have the highest flow rate of any biological molecule.
They are very easily opened and closed - no energy required.
The direction of the flow of water is determined by the osmotic gradient.

Question 14

Describe how water moves through an open aquaporin.

Answer 14

Water molecules move single-file through the pore.

Question 15

What is a charge relay and how it is caused/prevented by the aquaporin?

Answer 15

There is a charge relay: protons can hop from one water molecule to another and move across the pore. This charge relay is prevented by Asn192 and Asn76 which form transient interactions with water molecules, but not with H3O+ (formed if a proton has hopped onto that water molecule)

Question 16

Describe the structure of an aquaporin, focusing on the features which make it specific to H2O.

Answer 16

• An aquaporin is a monomer which contains 6 transmembrane a-helices.
• Two half helices make up the selectivity filter. Each half helix has Asn-Pro-Ala at its end, to prevent charge relay of protons. This means that the selectivity filter allows the passage of H2O but not H3O+.
• The pore narrows to 2.8 A, which is just wide enough to allow H2O through.
• Positively charged amino acid residue (Arginine) prevents passage of positively charged ions such as H3O+ (which is a similar size to H2O).

Question 17

Why is it so important that aquaporins only transport H2O, not H3O+?

Answer 17

Cells must maintain a membrane potential. Aquaporins must therefore prevent the movement of charged molecules, such as H3O+.

Question 18

Describe the structure of a K+ channel.

Answer 18

The K+ channel has comparatively simple structural for an integral membrane protein. It is made up of 4 polypeptide chains each with 2 a-helices and a third shorter helix-pore region. The opening of the channel has many negatively charged amino acid residues, which attract K+ and other positively charged ions. The ion selectivity filter allows the passage of K+ (1.33 A) but not Na+ (0.95 A).

The first part of the pore is a large water-filled channel, which allows the ions to enter still in hydrated form (hydration shell).

Question 19

Describe the selectivity filter of the K+ channel.

Answer 19

Each of the four subunits in the K+ channel has a selectivity loop, which can carbonyl functional groups projecting from it into the pore. These carbonyls stabilise the K+ ion better than the water molecules in the hydration shell so that it is energetically favourable for K+ ions to shed their hydration shell and form transient interaction with the oxygen molecules of the carbonyl projections.

Na+ is too small to form transient interactions with the carbonyl oxygens, so the carbonyls cannot stabilise Na+ more than its hydration shell and so it is not energetically favourable for it to shed its hydration shell.

There are four potential binding sites, only two of which can be occupied at any one time (not positions next to each other, like 1 and 2, though), because the K+ ions repel each other. This pushes the K+ ion already in the pore out into aqueous region on other side of membrane.

Transport through the K+ channel is unidirectional.

Question 20

Describe the structure of voltage gated channels.

Answer 20

They have 6 transmembrane domains and have a soluble region extended into the cell called S4. The core of the voltage-gated channel is similar to the K+ channel.

Question 21

Describe how voltage-gating works.

Answer 21

The extra domains (S4) are responsible for the voltage gating. They sense changes in the membrane potential and move in response to a change, causing the opening or closing of the channel.

Normally the transmembrane electrical potential (negative inside) pulls positive S4 to cytosolic side of membrane. Depolarisation reduces the pull towards the cytosol and reversal of the potential pulls S4 to extracellular side of membrane. The movement of S4 is coupled to the opening/closing of the channel. There is not a lot of conformational change between the open and closed forms. When the pore is closed it is too narrow for K+ to pass through.

Question 22

Discuss channel-blocking compounds.

Answer 22

Compounds that block channels are effectively poisons. They can be used as antibiotics as long as they only block the pathogen’s channels and no human ones.

Poisons which are channel blockers include dendrotoxin (from black mamba snake), tubocurarine, cobrotoxin and bungratoxin. These poisons all cause paralysis and can be fatal if the antidote isn’t administered quickly.