Transmembrane Receptors that Activate G Proteins - Cyclic AMP Signalling Flashcards
What is so different about G protein structure compared to other receptors we’ve covered?
They have 7 transmembrane domains. whilst the others are all in the single membrane-pass category
What are G protein involved in?
light-detection/visual system (rhodopsin) taste, smell (odorants and pheromones)
regulation of cognition, behaviour and mood (in brain bind serotonin, dopamine, glutamate, GABA)
autonomic nervous system transmission: responsible for control of blood pressure, heart rate, and digestive processes
How often are GPCRs target?
~40% of all modern drugs target them
How many genes in humans are estimated to encode GPCRs?
800
Generic structure of GPCRs
N-terminus outside the cell binds ligands/signalling molecules that can’t cross the membrane
7 transmembrane helices
C-terminus inside the cell that interacts with Heterotrimeric guanine nucleotide-binding proteins (G proteins)
Heterotrimeric guanine nucleotide-binding protein
(G protein)
Alpha subunit (binds to GPCR)
Beta subunit
Gamma subunit
Alpha & gamma have lipid modifications to interact in the membrane + are always associated with the membrane
What is the G protein doing when there’s no signal?
Resting
May be associated with the membrane or nearby ready to go
Bound to GDP
2 different types of G protein
Heterotrimeric guanine nucleotide-binding protein (what we’re talking about here)
Small G proteins (no beta and gamma subunits, part of the alpha subunit missing, don’t interact with GPCRs)
How are G proteins activated?
Some kind of input causes it to lose affinity for GDP and take up GTP instead, causing a conformational change that creates an interface perfectly shaped to fit an effector. The effector becomes activated and leads to the outputs.
How are G proteins deactivated?
Alpha subunit has ow levels of enzymatic activity; it starts hydrolysing terminal phosphate on GTP to turn it back into GDP, switching the G protein off with another conformational change
Signalling by G protein coupled receptors
Binding of ligands on extracellular region of GPCR induces changes in the tight ‘packing’ of transmembrane helices leading to changes on the intracellular face of membrane
Induces exchange of GDP for GTP on a subunit of associated heterotrimeric G protein
Activated (GTP-bound) a subunit dissociates from activated bg subunits and interacts with effectors
GPCRs act as guanine nucleotide exchange factors
Guanine nucleotide exchange factors (GEFs) deform the nucleotide-binding regions of the G protein, promoting the release of GDP
GTP then binds to the nucleotide-binding site because of the much higher concentration of GTP over GDP in the cell (x 10 more)
Molecular basis of G protein activation
GDP & GTP differ only by presence of a terminal phosphate on GTP
When present, this terminal phosphate interacts with 2 loops to form hydrogen bonds with the main chain atoms of conserved Gly and Thr residues
Signalling via cyclic AMP
Ligand binding to GPCRs coupled to Gs activates adenylyl cyclase which synthesises cyclic AMP from ATP
Cyclic AMP is degraded by phosphodiesterases – restricts cAMP diffusion in cell
4 known effectors of cyclic AMP
1) activation of protein kinase A
2) activation of EPAC
3) activation of cyclic nucleotide-gated ion channels
4) Popeye domain containing-(popdc-) proteins
Activation of protein kinase A by cyclic AMP
Binding of cyclic AMP to two sites on each R subunit causes the R subunits to dissociate from the C subunits.
C subunits now active and can phosphorylate target proteins. PKA is a serine/threonine kinase
The problem with cyclic AMP
Little, can potentially move unimpeded through the cell, and could potentially activate anything -> so how do we get specific responses?
Signalling specificity
A limited number of small signalling mediators are used to generate many diverse biological responses
A single mediator can be generated by many upstream inputs and can potentially activate many downstream effectors
A key question is how small signalling mediators can yield a specific output (yellow pathway)
How so scaffold proteins increase specificity?
By organising interacting molecules into complexes
What are AKAPs?
scaffold proteins that contain multiple protein binding sites and act to anchor PKA and its target (effector) close to where cyclic AMP is generated
How do AKAPs control cyclic AMP signalling?
anchor PKA and target close to where cyclic AMP is generated so that when a receptor is activated and cyclic AMP generated – activated PKA ‘finds’ and phosphorylates its target quickly and specifically
The AKAP complex may also contain phosphodiesterases, which destroy cyclic AMP, and phosphatases that dephosphorylate the target protein so the signal can be quickly and specifically switched off
