G-Protein Couple Receptors, Molecular Mechanisms Of Flashcards
List 4 major targets of drugs
Enzymes
Transporters
Ion channels
Receptors
Define: Receptors
Molecules through which soluble physiological mediators produce their desired effects (i.e. agonists) = binding = conformational change = signal transduction = response
Name the 4 types of receptor
Ligand-gated ion channels e.g. nicotinic acetylcholine receptors G protein-coupled receptor e.g. adrenoceptors, opioid receptors (morphine) Enzyme coupled receptors Nuclear receptors
Where are GPCRs found?
GPCRs are normally found at the cell surface
Intracellular Ca2+ made from action
Describe the structure of a GPCR
7 transmembrane domains - single polypeptide chain crosses membrane 7 times = forms a core
How do GPCRs work?
The receptor couples to G proteins to initiate signal transduction
Activated by diverse signals e.g. photons, hormones, peptides & peptidases
What are the 2 classes of GPCR?
Olfactory = sense receptors
Non-olfactory
Why are GPCRs important?
Make up over half the current drug targets
Largest family of cell surface receptor
120 orphan receptors
How many classes of GPCR are there?
6
A, B, C, D, E, F
Classed according to primary amino structure
What is the name of class A GPCRs?
Rhodopsin-like receptors
Easily targeted by antagonists
What is the name of class B GPCRs?
Secretin-like receptors
What is the name of class C GPCRs?
Metabotropic glutamate receptors
Describe class A rhodopsin like GPCRs
7 transmembrane domains
Short N terminus
Agonists bind with extracellular loops and transmembrane domains
e.g. β-adrenoceptors, histamine receptors
Describe class B secretin-like GPCRs
7 transmembrane domains
Larger, globular N-terminus
Agonists bind to N-terminus or extracellular loops
e.g. secretin receptors, glucagon receptors
Describe class C metabotropic glutamate GPCRs
Very large N terminus
Exist at cell surface as dimers - form obligatory dimers when agonists bind
e.g. GABA(B) receptors, metabotropic glutamate receptors
How are GPCRs activated?
i.e. how are signals conveyed from one side of the plasma membrane to the other
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
What are the 3 types of G protein?
Stimulatory = Gαs
Inhibitory
q/11
Describe the signalling events triggered by a stimulatory G protein
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)
What do Gαs coupled receptors promote?
They promote the formation of cAMP and the activation of PKA
Describe the signalling events triggered by inhibitory G proteins
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
What do Gαi/o coupled receptors inhibit?
They inhibit the activity of adenylate cyclase
= decrease in cAMP
= PKA activity is shut off
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
The q/11 G protein does not act on adenylate cyclase
Stimulatory activates it, inhibitory inhibits it
Describe the signalling events triggered by q/11 G proteins
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
Therefore what do q/11 G proteins promote?
Gαq/11 coupled receptors promote release of intracellular calcium
Which channels do GPCRs modulate the function of?
GPCRs modulate the function of ion channels
How do GPCRs modulate the function of ion channels?
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
What is the desensitisation of GPCRs?
Molecular mechanism
Switches off activation of GPCRs - prevents their continuous activation
Distinct from downregulation (=the degradation of receptors)
Name the 2 types of desensitisation of GPCRs
Homologous desensitisation
Heterologous desensitisation
Define: Homologous desensitisation
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
Define: Heterologous desensitisation
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
Define: Downregulation
Degrading of receptors = fewer receptors in cell
Describe the process of desensitisation of GPCRs
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
Define: Tolerance
Tolerance is the progressive reduction in the effectiveness of a drug
Usually occurring over days or hours
e.g. morphine at μ-opioid receptors
Explain if tolerance is developed at GPCRs when there has been no pre-treatment
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
Explain the development of tolerance at GPCRs (μ-opioid receptors) to chronic morphine
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
Why do enkephalins and morphine cause different responses from the μ-opioid receptor?
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
