G-Protein Couple Receptors, Molecular Mechanisms Of

Question 1

List 4 major targets of drugs

Answer 1

Enzymes
Transporters
Ion channels
Receptors

Question 2

Define: Receptors

Answer 2

Molecules through which soluble physiological mediators produce their desired effects (i.e. agonists)
= binding
= conformational change 
= signal transduction
= response

Question 3

Name the 4 types of receptor

Answer 3

Ligand-gated ion channels
e.g. nicotinic acetylcholine receptors
G protein-coupled receptor
e.g. adrenoceptors, opioid receptors (morphine)
Enzyme coupled receptors
Nuclear receptors

Question 4

Where are GPCRs found?

Answer 4

GPCRs are normally found at the cell surface

Intracellular Ca2+ made from action

Question 5

Describe the structure of a GPCR

Answer 5

7 transmembrane domains - single polypeptide chain crosses membrane 7 times = forms a core

Question 6

How do GPCRs work?

Answer 6

The receptor couples to G proteins to initiate signal transduction
Activated by diverse signals e.g. photons, hormones, peptides & peptidases

Question 7

What are the 2 classes of GPCR?

Answer 7

Olfactory = sense receptors

Non-olfactory

Question 8

Why are GPCRs important?

Answer 8

Make up over half the current drug targets
Largest family of cell surface receptor
120 orphan receptors

Question 9

How many classes of GPCR are there?

Answer 9

6
A, B, C, D, E, F
Classed according to primary amino structure

Question 10

What is the name of class A GPCRs?

Answer 10

Rhodopsin-like receptors

Easily targeted by antagonists

Question 11

What is the name of class B GPCRs?

Answer 11

Secretin-like receptors

Question 12

What is the name of class C GPCRs?

Answer 12

Metabotropic glutamate receptors

Question 13

Describe class A rhodopsin like GPCRs

Answer 13

7 transmembrane domains
Short N terminus
Agonists bind with extracellular loops and transmembrane domains
e.g. β-adrenoceptors, histamine receptors

Question 14

Describe class B secretin-like GPCRs

Answer 14

7 transmembrane domains
Larger, globular N-terminus
Agonists bind to N-terminus or extracellular loops
e.g. secretin receptors, glucagon receptors

Question 15

Describe class C metabotropic glutamate GPCRs

Answer 15

Very large N terminus
Exist at cell surface as dimers - form obligatory dimers when agonists bind
e.g. GABA(B) receptors, metabotropic glutamate receptors

Question 16

How are GPCRs activated?

i.e. how are signals conveyed from one side of the plasma membrane to the other

Answer 16

Agonist binds to the GPCR
Conformational change occurs = new surface charge (new AAs, new charges uncovered, new shape)
Attracts heterotrimeric G proteins which is formed of 3 sub-units (α, β, γ)
Receptor can now bind G proteins at C terminal as the receptor shape has changed
GTPase activity activated (part of heterotrimeric protein, α) = exchange of GDP to GTP = GDP is displaced (affinity)
GTP dissociates (hydrolysed by α sub-unit)
Molecules then signal downstream to effectors = response

Question 17

What are the 3 types of G protein?

Answer 17

Stimulatory = Gαs
Inhibitory
q/11

Question 18

Describe the signalling events triggered by a stimulatory G protein

Answer 18

The Gαs sub-unit is now activated and GTP-bound
Adenylate cyclase (very large protein) is present in the plasma membrane (cell surface) and has a catalytic domain
The stimulatory G protein has a positive effect on the catalytic site = promotes the activity of the adenylate cyclase
This converts ATP to cAMP
Rapid build up of cAMP in cell
This activates protein kinases e.g. protein kinase A (PKA)

Question 19

What do Gαs coupled receptors promote?

Answer 19

They promote the formation of cAMP and the activation of PKA

Question 20

Describe the signalling events triggered by inhibitory G proteins

Answer 20

Gαi/o sub-unit is now activated and GTP bound
It inhibits the activity of the catalytic domain of adenylate cyclase
So ATP is not converted to cAMP
Therefore PKA activity is shut off

Question 21

What do Gαi/o coupled receptors inhibit?

Answer 21

They inhibit the activity of adenylate cyclase
= decrease in cAMP
= PKA activity is shut off

Question 22

Name the main difference between the signalling events triggered by the stimulatory and inhibitory G proteins and the events triggered by the q/11 G protein

Answer 22

The q/11 G protein does not act on adenylate cyclase

Stimulatory activates it, inhibitory inhibits it

Question 23

Describe the signalling events triggered by q/11 G proteins

Answer 23

Gαq/11 causes recruitment of phospholipase C to plasma membrane
PIP2 lipids exist in the membrane (substrate for phospholipase C)
PIP2 is cleaved into 2 smaller fragments
- 1 stays in the plasma membrane = diacylglycerol (DAG)
- 2 smaller fragment = IP3
The 2 fragments activate different signalling pathways
DAG activates protein kinase C(β) (recruited to plasma membrane)
IP3 binds to receptors present on the endoplasmic reticulum = causes eflux of intracellular calcium into cell
Calcium rapidly brought back into ER by transporters

Question 24

Therefore what do q/11 G proteins promote?

Answer 24

Gαq/11 coupled receptors promote release of intracellular calcium

Question 25

Which channels do GPCRs modulate the function of?

Answer 25

GPCRs modulate the function of ion channels

Question 26

How do GPCRs modulate the function of ion channels?

Answer 26

Gβγ sub-units have a negative effect on the activation of the 1/11 loop in a voltage-gated calcium channel Induces a loss of function in the voltage-gated calcium channel and therefore there is a reduction in neurotransmitter release e.g. GABA receptors

Question 27

What is the desensitisation of GPCRs?

Answer 27

Molecular mechanism Switches off activation of GPCRs - prevents their continuous activation Distinct from downregulation (=the degradation of receptors)

Question 28

Name the 2 types of desensitisation of GPCRs

Answer 28

Homologous desensitisation | Heterologous desensitisation

Question 29

Define: Homologous desensitisation

Answer 29

Desensitisation effects are restricted to agonists acting through a specific receptor i.e. if receptor A is desensitised, all receptor A's will become desensitised

Question 30

Define: Heterologous desensitisation

Answer 30

Desensitisation effects can affect receptors that share a component of the same signalling cascase i.e. if receptor A is desensitised, so is receptor B

Question 31

Define: Downregulation

Answer 31

Degrading of receptors = fewer receptors in cell

Question 32

Describe the process of desensitisation of GPCRs

Answer 32

Agonist binds to GPCR Conformational change to receptor Therefore GPCR kinases (GRK) joins bottom of receptor = promotes phosphorylation of receptor Phosphorylation occurs on serine and threonine within the intracellular loop and C terminal tail The phosphorylated form of the GPCR is now attractive to β-arrestins (stop signal transduction) which facilitate internalisation β-arrestins bind and recruit scaffolding proteins = form a cage around multiple GPCRs and promote its internalisation into internal vesicles (endosomes) Once in endosome, ligand, agonist and arrestins are released Receptor becomes dephosphorylated by phosphatases Receptor can then be recycled and returned back to surface = resensitisation OR Receptor can be trafficked to a lysosome where they are degraded = downregulation

Question 33

Define: Tolerance

Answer 33

Tolerance is the progressive reduction in the effectiveness of a drug Usually occurring over days or hours e.g. morphine at μ-opioid receptors

Question 34

Explain if tolerance is developed at GPCRs when there has been no pre-treatment

Answer 34

Endogenous agonist (e.g. enkephalins) binds to the receptor and activates it Agonist promotes the phosphorylation of receptor by GPCR kinases (GRKs), the recruitment of arrestins and therefore internalisation of receptors Receptors are recycled and so ready to be activated again Can keep activating with enkephalins and tolerance is not developed

Question 35

Explain the development of tolerance at GPCRs (μ-opioid receptors) to chronic morphine

Answer 35

Morphine is a partial agonist so does not cause efficient internalisation of receptor = does not activate GRK-β arrestin pathway Morphine binds to receptor Activates protein kinase C (PKC) So receptor is phosphorylated and desensitisation is caused The receptor is not internalised very well and so cannot go through a resensitisation and recycling process Receptor sits at surface of cell still desensitised for longer periods of time Morphine can still bind but because of the different phosphorylation, it cannot activate the pathway = pain relief

Question 36

Why do enkephalins and morphine cause different responses from the μ-opioid receptor?

Answer 36

Both activate the receptor but activate different signalling pathways Enkephalins = phosphorylation by GRK, arrestins, internalisation - no tolerance developed Morphine = PKC activated, β-arrestin pathway not activated (inefficient internalisation) - tolerance developed