Membrane Proteins and Function

Question 1

What three ways can membrane proteins attach?

Answer 1

1. A groove/polar pocket that binds a lipid or recognises a specific ligand
2. An ion binding site
3. A shape that is complementary to the membrane, a curved surface of head groups with charges that interact with the membrane

results in peripheral membrane proteins - binding at the periphery and do not penetrate through the membrane

Question 2

Fun fact!

Answer 2

~50% of surface of synaptic vesicle is proteins

Question 3

Attachment of peripheral proteins to membranes is permanent. True or False?

Answer 3

False. Different peripheral proteins have differing degrees of transiency; some being more permanently attached than others but still reversible

Question 4

Describe the binding of ENTH type domains to membranes and give an example protein with this motif.

Answer 4

ENTH motif binds PI(4,5)P2 ligand, more permanent because of a hydrophobic protrusion into the membrane binding a head group - but still can be reverted.

Question 5

Describe the binding of BAR type domains to membranes and give an example protein with this motif.

Answer 5

BAR motif binds complementary structure to membrane strongly as large surface, e.g. in Amphiphysin, but interactions with ions or exchanging phospholipid head groups can relieve the binding of the domain and the membrane.

Question 6

Describe the binding of Pleckstrin Homology type domains to membranes and give an example protein with this motif.

Answer 6

PH motifs bind phosphatidyl inositols; PIP2, PIP3 e.g. in PLC.

Question 7

Describe the binding of C2 type domains to membranes and give an example protein with this motif.

Answer 7

C2 binds Ca2+ and anionic phospholipids, e.g. in PKC

Question 8

Describe the binding of Ankyrin-repeat type domains to membranes and give an example protein with this motif.

Answer 8

Ankyrin repeat domains bind phosphatidylserine, e.g. Ankyrin

Question 9

Describe the binding of FERM type domains to membranes and give an example protein with this motif.

Answer 9

FERM domains bind PI(4,5)P2, e.g. in Ezrin, radixin and moesin.

Question 10

Describe the binding of FYVE type domains to membranes and give an example protein with this motif.

Answer 10

FYVE domains bind PI(3)P e.g. EEA1

Question 11

Describe the binding of PX type domains to membranes and give an example protein with this motif.

Answer 11

PX domains bind PI(3)P, e.g. sorting nexins

Question 12

The association of peripheral proteins with membranes is dynamic and depends on what 4 things?

Answer 12

1. The type of membrane
2. Ca2+ concentration
3. Availability of lipid species
4. Shape of membrane

Question 13

What three ways can proteins cause membrane deformation?

Answer 13

1. Amphipathic helix - polar and hydrophobic amino acids on opposite sites of helix
2. Loop insertion - loops with hydrophobic amino acids
3. Curved lattices - lattices form a curved polymer that bind cargo proteins, forming vesicles of tight curvature.

(Or by proteins just binding themselves to complementary regions of the membrane, e.g, BAR domains)

Question 14

How do proteins become anchored in the membrane?

Answer 14

Through lipidation - addition of a lipid tail to the protein, which is often reversible.

Some lipid anchors are tucked away when the protein is in the cytosol.

Question 15

Give examples of proteins that are anchored to the membrane through lipidation.

Answer 15

G-proteins, C-terminus of GPCRs, SNAREs… etc.

Question 16

Give 3 examples of lipidation modifications allowing proteins to be anchored to a membrane.

Answer 16

1. Palmitoyl group on internal Cys or Ser
2. N-Myristoyl group on amino-terminal Gly
3. Farnesyl or Geranygeranyl group on carboxyl-terminal Cys

These lipid modifications result in the protein being tucked on the inside facing the cytosolic environment

Question 17

What are GPI anchors?

Answer 17

GPI = Glycosylphosphatidylinositol
A phosphoglyceride attached to the C-terminus of a protein during post-translational modification anchoring of proteins to plasma membranes.

The phosphate connects glycerol to an inositol group, which is attached to sugars, e.g. GlcNAc, Mannose chain. Attaches to protein by phosphate at C-terminus.

Question 18

Where are GPI anchors found?

Answer 18

Exclusively in plasma membranes and primarily in lipid rafts, facing outside of the cell

Question 19

Where are GPI-anchors synthesised?

Answer 19

In the ER and Golgi

Question 20

What are integral membrane proteins?

Answer 20

AKA Transmembrane/TM proteins, are proteins that cross the membrane.

Question 21

How are integral membrane proteins different from peripheral and GPI-anchored proteins?

Answer 21

Peripheral and GPIs can be cleaved and then become soluble to be released from the membrane. TM proteins cannot.

Also cannot be removed because of a hydrophobic helix which would not be stable outside the membrane environment.

Question 22

How many types of TM proteins are there?

Answer 22

4

Question 23

Describe the structure of Type I transmembrane proteins.

Answer 23

A single transmembrane domain;
C-terminus of protein is in cytosol.
N-terminus is facing extracellular environment, or lumen of intracellular compartments.

Question 24

Describe the structure of Type II transmembrane proteins.

Answer 24

A single transmembrane domain;
N-terminus is in cytosol
C-terminus is facing extracellular environment/lumen of intracellular compartments

Question 25

Describe the structure of Type III transmembrane proteins.

Answer 25

Tail-anchored protein;
N-terminus on outside,
C-terminus on inside, with a very short tail. (Only the N-terminus is in the outside)

Question 26

Describe the structure of Type IV transmembrane proteins.

Answer 26

Multiple TM domains unlike TI-TIII;
Polytopic TM proteins.
N or C terminus on either side.

Question 27

Give examples of Type I TM proteins.

Answer 27

Glycophorin, LDL receptor, Influenza HA, Insulin receptor, growth hormone receptor

Question 28

Give examples of Type II TM proteins.

Answer 28

Asialoglycoprotein receptor, Transferrin receptor, Sucrase-isomaltase precursor, Golgi galactosyltransferase, Golgi siayltransferase, influenza HN protein

Question 29

Give examples of Type III TM proteins.

Answer 29

Cytochrome P450

Question 30

Give examples of Type IV TM proteins.

Answer 30

GPCRs, GLUT1, VGCCs, ABC small molecule pumps, CFTR Cl- channels, Sec61, connexin.

Question 31

Why are TM domains mostly alpha helical?

Answer 31

In an alpha helix, the hydrophilic part - the peptide backbone - of an amino acid, can be buried in the middle of an alpha helix so that all the hydrophobic amino acid side chains are pointing toward the fatty acid layer of the membrane.

Question 32

How long are the alpha helices in TM domains?

Answer 32

~20 AAs long as have the length of the membrane but depending on the membrane specifically.

Question 33

Where are aromatic amino acids found in transmembrane protein alpha helices?

Answer 33

Contain lipid head groups; found at edges of TM helices

Question 34

Give a role for non-hydrophobic amino acids in TM alpha helices.

Answer 34

Positive charges can be used in voltage sensors for channel proteins

Question 35

What is the hydropathy index?

Answer 35

The hydropathy index, as plotted by a hydropathy plot, peaks where the average hydrophobicity is highest - in the middle of a TM alpha helix.

Question 36

TM domains of integral proteins project hydrophobic side chains into the bilayer. True or False?

Answer 36

True.

Question 37

How can TM proteins be solubilised?

Answer 37

Not soluble naturally - detergents must be added which add themselves into the bilayer.

Excess detergent breaks up the membrane and removes lipids into micelles.

Hydrophobic TM domains are covered by detergents becoming soluble.

Question 38

What are the names of membrane regions that resist solubilisation by detergents?

Answer 38

Detergent-Resistant Membranes/DRMs - or Lipid Rafts

Question 39

What are lipid rafts?

Answer 39

Detergent-Resistant Membranes

Question 40

Describe how properties of lipid rafts aid its function

Answer 40

Less fluidity in lipid raft - ideal for localisation of signalling molecules.
Less conformational flexibity - proteins that need to be more rigid are localised in lipid rafts

Useful in immune signalling but also host/pathogen interactions, providing a structured environment for pathogen binding and viral budding.

Question 41

What are the three classes of transport proteins?

Answer 41

1. Pumps
   - Primary active transport by ATP
   - High affinity for ligand, used to accumulate solutes against large gradients; quite slow relatively.
2. Carriers
   - Facilitators, not directly energised, transport along concentration gradient or using a secondary solute.
   - Intermediate affinity, faster than pumps slower than channels.
3. Channels
   - Diffusion pores, passive but very fast transport along a concentration gradient, a regulating open/shut gate.
   - Ion specificity, transport water across membranes, only depend on gradient and transport only on gradient.

Question 42

On which face of the ER does synthesis of most phospholipids occur?

Answer 42

The cytoplasmic face
Phosphatidylethanolamine (PE) and Phosphatidylserine (PS) are on the correct leaflet of the plasma membrane, but not Phosphayidylcholine (PC). -> leads to lipid asymmetry.

Question 43

What is a role of phosphatidylserine (PS)?

Answer 43

A marker of cells that are undergoing apoptosis -> membrane potential is lost, so serine cannot be transported and serine is deposited on the outside

Question 44

What are flippases?

Answer 44

Phospholipid transport protein - moving phospholipids from outside to inside e.g. phosphatidylserine

Question 45

What are floppases?

Answer 45

Phospholipid transport protein - moving phospholipids from inside to outside e.g. phosphatidylcholine or cholesterol.

Question 46

What is common between flippases and floppases?

Answer 46

Both transfer phospholipids across membranes
Both are pumps
Both use ATP to get the phospholipid head-group across the membrane.

Question 47

What are aquaporins?

Answer 47

Water transport channel proteins.

Very high conductance and very selective for water.

Question 48

Describe 1992 experiment by Peter Agre demonstrating the significance of aquaporins.

Answer 48

Frog oocytes with aquaporin knockouts do not expand in hypoosmotic solution.
WT oocytes do expand in hypoosmotic solution.