Superfamily 2

Question 1

What is the molecular structure of G protein coupled receptors?

Answer 1

Single polypeptide chain
Typically 400-500 amino acids long
7 transmembrane spanning alpha helices
Ligand binding site embedded in membrane surface
Long 3rd cytoplasmic loop interacts with G protein

Question 2

What are the features of the intracellular C-terminal?

Answer 2

Phosphorylation (involved in receptor desensitisation)
Palmitoylation to anchor in lipid membrane
Interacts with scaffolding proteins to orientate in membrane and localise micro-domains
Interacts with other enzymes such as tyrosine kinases to influence signalling (upon agonist binding)

Question 3

Give some interesting facts about G protein coupled receptors

Answer 3

Largest superfamily
1-3% of total genome
Targets for >30% of all drugs
>900 genes

Question 4

Give some examples of G Protein coupled receptors

Answer 4

Muscarinic cholinoceptor
Adrenoceptors
Histamine, dopamine, serotonin (amine receptors)
Endothelin and angiotensin (peptide mediators)
Prostaglandin receptors
Rhodopsin and olfactory chemoreceptors

Question 5

What are the three subgroups of G protein coupled receptors?

Answer 5

Group A
Group B
Group C

Question 6

Describe Group A GPCR

Answer 6

Rhodopsin / beta2 adrenoceptor like
Short N terminal
Binding site embedded in TM alpha helices
Includes eicosanoid, purine and amine receptors

Question 7

Describe Group B GPCR

Answer 7

Glucagon/ VIP/ CT receptor like
Large N terminal
Several cys-cys bridges
Many peptide receptors

Question 8

Describe Group C GPCR

Answer 8

Metabotropic neurotransmitter / calcium receptors
Very large N terminal
GABA A, pheromone and taste receptors

Question 9

Describe heterotrimeric G proteins

Answer 9

Consist of 3 subunits
Alpha, beta and gamma

Question 10

Describe small G proteins

Answer 10

5 subfamilies
RAS, Rho, ARF, Rab, Ran
Molecular weight 20-30kDa
Regulate wide variety of cellular processes

Question 11

What do heterotrimeric G proteins do?

Answer 11

Communicate between receptor and effector enzymes or ion channels
Require GTP for activation

Question 12

What is the mechanism of action for GPCR?

Answer 12

Agonist binding destabilises intra molecular interactions
Alpha helixes TM3/TM6 move apart during activation
Expose cytoplasmic sequences that facilitates G protein interaction
2nd cytoplasmic loop confers efficiency of G protein activation
3rd cytoplasmic loop confers specificity of G protein interaction

Question 13

What are the basic steps of GPCR?

Answer 13

Agonist
Receptor
G protein
Channel/ enzyme

Question 14

Describe what stimulatory G proteins do

Answer 14

Induce a conformational change in the target to a more active state
To enhance activity

Question 15

Describe what inhibitory G proteins do

Answer 15

Induce a conformational change in the target to a less active state
To reduce activity

Question 16

What are the properties of G protein alpha subunits?

Answer 16

Different receptor interaction sites
Different target interaction sites
Similar GTP/ GDP site and GTPase activity
Variable sensitivity to bacterial toxins

Question 17

What effectors are regulated by G proteins?

Answer 17

Adenylate cyclase
Phospholipase C-beta
Voltage gated potassium channels
Voltage gated calcium channels

Question 18

What are the differences between Galpha proteins?

Answer 18

GTP affinity
GTPase kinetics
Regulation
Localisation

Question 19

What are the four different types of Galpha proteins?

Answer 19

G alpha (s/ olf)
G alpha (I/ o/ t/ Z)
G alpha (q / 11)
G alpha (12 / 13)

Question 20

What does the G alpha (s/olf) do?

Answer 20

Increase adenylate cyclase
Increase calcium channels

Question 21

What does G alpha (I/o/t/z) do?

Answer 21

Decreases adenylate cyclase
Increases potassium channels
Decreases calcium channels
Increases cGMP-specific PDE

Question 22

What does G alpha (q/11) do?

Answer 22

Increases phospholipase C-beta and others

Question 23

What does G alpha (12/13) do?

Answer 23

Increase chloride channels
Regulates protein Ras, Rho activity

Question 24

Explain the example of cholera on GPCR

Answer 24

‘Faulty off switch’
Arg amino acid impairs GTPase activity
Uncontrollable increased activity of Gs/ cyclic AMP
Increased transluminal movement of water and electrolytes
Severe diarrhoea

Question 25

Explain the example of whooping cough on GPCR

Answer 25

Pertussis toxin Gi family
C terminal cys residue prevents transduction between receptor and effector
Stabilises G protein in inactive state
Decreases activity of some Gi, Go, Gt
Immobilises cilia prompting cough
Interferes with signalling in phagocytes and their ability to counter infection

Question 26

What is the clamshell model?

Answer 26

Agonist receptor induces changes in G protein confirmation
Exposes G alpha and G beta gamma interfaces
Act as scaffolds to direct interaction with respective effectors
G alpha remains closely associated with G beta gamma
Not interchangeable between other G alpha subunits

Question 27

Describe some of the features of regulators of G protein signalling proteins

Answer 27

Largely inhibitory
Cytosolic proteins adjacent to membrane G proteins
Roughly 20- 25kDa
Bind directly to activated G alpha subunits
Interact with C terminal of receptor and with specific domains of effector

Question 28

What is the mechanism of action of regulating proteins?

Answer 28

GTPase activating proteins - increases the inactivation rate of G alpha, promotes heterotrimer reformation so also blocks G beta gamma signalling
Competitive inhibitor of G alpha effector or G beta gamma effector interactions
Impedes GDP releases/ prevents GTP binding/ G alpha activation