Membranes (Williamson)

Question 1

From an evolutionary perspective, why did membranes arise?

Answer 1

To define a barrier between ‘inside’ and ‘out’.

Question 2

What is the key function of membranes that all cells perform?

Answer 2

Transport of chemicals in/out (nutrients/waste)

Question 3

What functions of membranes evolved later?

Answer 3

Conversion of membrane potential to energy. Cellular recognition. Signalling from outside to inside. Molecule trafficking (internal).

Question 4

Why do Eukaryotic cells need to compartmentalise?

Answer 4

They are so much larger than Prokaryotic cells. Different functions must be enclosed, molecules must be transported to different places.

Question 5

Membrane lipids have hydrophobic and hydrophilic ends. What does ths cause?

Answer 5

Spontaneous aggregation.

Question 6

In vitro, what structures can membrane lipids spontaneously aggregate into?

Answer 6

Lipid bilayer. Liposome. Vesicle.

Question 7

What does FRAP stand for?

Answer 7

Fluorescence Recovery after Photobleaching.

Question 8

What did using FRAP on membrane proteins demonstrate?

Answer 8

That membrane proteins are laterally mobile.

Question 9

What does Atomic Force Microscopy of a membrane show?

Answer 9

That the bilayer is tightly packed with proteins.

Question 10

AFM showed membranes to be tightly packed with proteins. What does this imply?

Answer 10

That membranes may not be as fluid as the the Fluid Mosaic Model implies.

Question 11

What type of lipids make up most of the membrane?

Answer 11

Phospholipids (phosphoglycerides)

Question 12

Phospholipids bind the tails with Os. What do sphingolipds use?

Answer 12

NH

Question 13

What double bond configurations are often found in sphingolipids?

Answer 13

Trans

Question 14

How do cells alter membrane fluidity?

Answer 14

My changing the lipid composition of the membrane

Question 15

How does lipid composition affect fluidity?

Answer 15

Cis double bonds take the lipid chains off at angles, whereas trans double bonds allow the chain to extend linearly.

Question 16

What lipids does cholesterol pack against?

Answer 16

Trans double bond lipds.

Question 17

What affect does cholesterol packing have on the lipid bilayer?

Answer 17

It flattens the lipids making them longer (increasing the bilayer length)

Question 18

PE (Phosphatidylethanolamine) has a smaller head group than PC (Phosphatidylcholine). What effect does this have on the bilayer?

Answer 18

PE in the bilayer caues it to curve.

Question 19

Describe 5 ways in which lipids are used to control membrane curvature.

Answer 19

1) Lipid composition - head group and acyl chain 2) Membrane proteins - shape and oligomerisation 3) Cytoskeleton - Internal actin control/external motor control 4) Scaffolding - Indirect/Direct +ve, -ve 5) Amphipathic helix insertion

Question 20

Which orientation are GPI anchored proteins in?

Answer 20

Outside

Question 21

Which orientation are lipid anchored protein in?

Answer 21

Inside

Question 22

What are flippases and what do they do?

Answer 22

Flippases are ATP dependant enzymes that flip lipids in the bilayer.

Question 23

What is phase separation in membranes?

Answer 23

Groups of similar lipds coming together to form clusters.

Question 24

What is a membrane raft?

Answer 24

Thicker membrane regions with more cholesterol and sphingolipids.

Question 25

Proteins also form groups based on similarities. Describe some group types.

Answer 25

Proteins with TM helices. Proteins with GPI anchors. Proteins with palmitoyl anchors. Proteins with prenyl anchors.

Question 26

Is membrane raft formation spontaneous?

Answer 26

No.

Question 27

Why is membrane raft formation controlled by the cell?

Answer 27

As this can form a control mechanism for various systems. eg Bringing signalling systems together. Starting endocytosis. T Cell activation.

Question 28

Membrane rafts can be used to bring proteins together or to move them apart. What kind of proteins can be added/removed from these rafts?

Answer 28

GPI linked, prenylated etc - any covalently linked protein.

Question 29

What major proteins are involved in ligand-mediated endocytosis?

Answer 29

Caveolins

Question 30

How do caveolins lead endocytosis?

Answer 30

They insert halfway into the membrane, causing it to curve inwards.

Question 31

What is patch clamping used for?

Answer 31

Measuring conductance across a membrane (ion channels)

Question 32

What does the patch clamping data show? (There are two ion channels)

Answer 32

* Ion channels have the same current when open * Opening/closing is random

Question 33

What types of ion transporters are found in an axon?

Answer 33

* Voltage-gated Na⁺ channel * Voltage-gated K⁺ channel * Na⁺/K⁺ pump * K⁺ leak channel

Question 34

What is the typical resting membrane potential?

Answer 34

-60mV

Question 35

At what membrane potential does the Na⁺ channel begin to open?

Answer 35

-40mV

Question 36

What is the Na⁺ channel plug, and when is it 'used'?

Answer 36

It plugs the open channel after it has been open for ~1ms. It remains closed until normal resting potential has been re-established.

Question 37

What is the main difference between the K⁺ channel and the Na⁺ channel?

Answer 37

The K⁺ channel opens and closes slower than the Na⁺ channel.

Question 38

What does the Na⁺/K⁺ channel do?

Answer 38

It pumps 3Na⁺ out for every 2K⁺ in. It is ATP dependant.

Question 39

What to K⁺ leak channels do?

Answer 39

They are always open, allowing small amounts of K⁺ to leak out, giving the cell membrane a negative potential.

Question 40

What are nerve impulses?

Answer 40

Transient changes in membrane potential.

Question 41

Since nerve impulses are 'all or nothing', how is a stronger signal created?

Answer 41

More rapid impulse firing.

Question 42

An action potential initally depolarises the membrane to the threshold potential of -40mV. What happens at this point?

Answer 42

Na⁺ channels open, causing Na⁺ to rush in to the cell. This causes the membrane potential to rush up to around +35mV.

Question 43

After the peak of the action potential (+35mV), what happens?

Answer 43

The delayed K⁺ channels start to open. Potassium leaves the cell reducing the membrane potential. It becomes hyperpolarised ( \> -70mV)

Question 44

What is a useful consequence of membrane hyperpolarisation in action potentials?

Answer 44

Action potentials can only travel forwards.

Question 45

How many sodium and potassium ions are moved in/out of the cell, per μm² of membrane during an AP?

Answer 45

10⁵ of each ion.

Question 46

During an AP, what percentage of total cell ions are used?

Answer 46

1% of all cell sodium/potassium.

Question 47

What is the consequence of the percentage of ions used in an AP?

Answer 47

APs are very energy efficient.

Question 48

What is the speed of a action potential down an unsheathed axon?

Answer 48

1ms^-1

Question 49

Why is the myelin sheath of vital importance?

Answer 49

It speeds up nerve impulses 100x. 1ms^-1 is far too slow for any useful kind of nerve signalling.

Question 50

How long are the myelin sheath nodes, and how often do they occur?

Answer 50

They are 1μm long and occur every 100μm of nerve.

Question 51

What do myelin sheath nodes do?

Answer 51

They allow the nerve impulse to jump, speeding the impulse up.

Question 52

What are the four major pathways of signal transduction into the cell?

Answer 52

1. Hydrophobic molecule diffusion 2. Ion Channel 3. GPC Ligand Receptor 4. Ligand Enzyme Receptor

Question 53

What are G-proteins?

Answer 53

Molecular switches

Question 54

How do G-proteins change between on/off states?

Answer 54

They are on when bound to GTP and off when bound to GDP

Question 55

How does GTP being bound maintain the G-protein in an on state?

Answer 55

The extra phosphate in GTP (compared to GDP) forms H bonds with Thr35 and Gly60 in the 'spring sections'.

Question 56

In the cell, which is higher - [GTP] or [GDP], and by how many times?

Answer 56

[GTP] is 10x higher than [GDP]

Question 57

What are the two factors that aid G-proteins, and what do they do?

Answer 57

1. GEF - Guanine Exchange Factors - it swaps GDP for GTP 2. GAP - GTPase-activating protein - helps the G-protein to hydrolyse GTP

Question 58

What mechanism do receptor-linked kinases use to transduce signals?

Answer 58

Dimerisation

Question 59

When a ligand binds to a R-l kinase, and dimerisation occurs, what happens next?

Answer 59

The R-l kinase autophosphorylates.

Question 60

What are Grb2 and Sos proteins?

Answer 60

Modular signalling-adaptors

Question 61

What domains does Grb2 contain, and what do they do?

Answer 61

* SH2 - binds phosphorylated R-l kinases * SH3 - recognises polyproline helices

Question 62

What does Sos bind to?

Answer 62

The polyproline helix in Sos binds to SH3.

Question 63

Once Sos is complexed, what does it activate\>

Answer 63

It brings GEF to the cell surface to activate a cell surface linked Ras (G-protein)

Question 64

Once Ras becomes active, what does it activate?

Answer 64

A kinase called Raf

Question 65

What is the general structure of GPCRs?

Answer 65

7 transmembrane helices

Question 66

GPCR intracellular loop 3 binds to what?

Answer 66

A heterotrimeric G protein (Gα, Gß, Gγ)

Question 67

In the trimeric G-protein, which subunit bind GDP/GTP?

Answer 67

Gα

Question 68

Once Gα is activated, what does it do?

Answer 68

Moves along the membrane looking for a target to activate.

Question 69

What is a typical Gα target?

Answer 69

Adenylyl cyclase

Question 70

What does adenylyl cyclase do?

Answer 70

It converts ATP to cAMP, which acts as a secondary messenger.