Flashcards in Signal Transduction II Deck (40):
What does an agonist do?
It mimics the physiological ligand- it turns on a receptor
Synonyms for GCPR
heptahelical, serpentine (due to topography in the membrane)
How many transmembrane segments do GCPR have?
7 (sit as a helical bundle)
What sits down in the middle of a GCPR bundle?
G proteins that interact with GCPR are of what conformation?
What does triheteromeric mean?
Three subunits that are not identical (a, B, y)
In the basal state, what is bound to the a subunit of G proteins?
In the basal state, are the a,b, and y subunits complexed together?
Which subunit has intrinsic GTPase activity?
Which subunit has lipid modification?
a and y
Is the y subunit hydrophobic or hydrophilic?
What do lipid modifications do on G protein subunits?
tether them to the membrane (thus, only a and y are tethered to the membrane because b does not have lipid modification)
How does GCPR signal transduction begin?
An agonist comes in and binds to a receptor, promotes a conformational change in the receptor, and begins to interact with the G protein
What is the rate limiting step in GCPR signal transduction?
GDP release from the a subunit
When is GDP released from the a subunit?
When the GCPR binds to the G protein
What happens after GDP is released from the a subunit?
GTP (at high conc in the cell) and immediately binds to the empty GDP binding site on the a subunit- this process is unidirectional
Binding of GTP to the a subunit causes what?
separation of the a subunit from the B/y subunit - this activates both subunits (a and By)
What does the a subunit do once its activated?
it can now act upon a target protein or effector and activate it
What causes inactivation of the a subunit?
GTPase activity on the a subunit auto-hydrolyzes the GTP to GDP, causing dissociation of the a subunit.
NOTE: The a subunit then re-combines with the B/y subunit to 'reset' the process
Do all GCPR work the same?
Yes. Same steps
Do adrenaline, glucagon, and adrenocorticotropic hormone work by cAMP transduction?
Does cAMP diffusion act quickly?
Yes. 100x increase in cAMP in some tissue in as little as 20 sec.
When are secondary messengers like cAMP release/synthesized?
When the a subunit becomes activated and binds with an effector protein. The effector protein is responsible for synthesizing/releasing cAMP, not the activated a subunit
What synthesizes cAMP? From what substrate?
adenylyl cyclase. from ATP
What breaks down cAMP? Into what?
cyclic AMP phosphodiesterase; into AMP
Does the cell want high levels of cAMP?
No, if you force the cell to maintain high levels of cAMP (i.e. block cyclic AMP phosphodiesterase), it will die
What are the main steps of adenylyl cyclase activation?
1) normal steps of activation of G protein
2) binding of activated G protein with adenylyl cyclase (thus, activating it)
3) adenylyl cyclase makes cAMP from ATP
4) cAMP activates PKA (protein kinase A)
-PKA has many targets in the cell cytoplasm and nucleus
What kind of kinase is PKA?
serine/threonine (only phosphorylates on serine or threonine in substrates)
How can cAMP affect glycogen breakdown in the cell?
activated PKA (from cAMP) uses ATP hydrolysis to activate phosphorylase kinase. Activated phosphorylase kinase uses ATP to activate glycogen phosphorylase, which causes glycogen breakdown
How can cAMP affects gene expression?
activated PKA goes through the nucleopore and activates/phosphorylates a transcription factor, thus up regulating target gene expression
How does GPCR regulation of ion channels using B subunits work?
acetylcholine binds to GCPR
(skip ahead a couple of steps) the activated B/y subunit complex (which at this point is dissociated from the a subunit due to GTP binding to the a subunit) binds to the K+ channel and opens it
inactivation comes when GTP on a subunit is hydrolyzed to GDP, causing re-assembly of subunits to inactive form
Phospholipase C breaks which 'head' region off phospholipids?
PIP2- Phosphatidylinositol 4,5, biphosphate
fatty acid chain breakdown by Phospholipase C makes what two products?
diacylglyercol and IP3
What is diacylglycerol?
The remnant body of the fatty acid chain that remains embedded in the cell membrane following cleavage of the head by phospholipase C (activated PKC)
What is IP3?
The 'head' region of the fatty acid cleaved by phospholipase C
What does IP3 do?
It binds to receptors on the membrane of the ER and causes release of Ca2+ stores from inside the ER to the cell cytoplasm
What does the Ca2+ do once released from the ER by IP3?
1) directly binds to some enzymes and activates them
2) activates calmodulin-a Calcium sensor
3 helps in activation of PKCs
How does Ca2+ affect calmodulin and what is the action of calmodulin?
Calmodulin is originally sitting in the cell doing nothing until Ca2+ levels rise. In the absense of calcium, calmodulin sits in an almost linear shape. When Ca2+ binds to it, it reconforms its structure to wrap around any calmodulin target (must bind Ca2+ in all four of calcium binding sites)- once it has those holes filled, it wraps around target
Once calmodulin binds to target enzyme, it releases the inhibitory domain of enzyme from catalytic domain. The enzyme can then auto-phosphorylate to fully activate itself. Calmodulin then falls off if Ca2+ levels drop, leaving an activated, Ca2+ independent enzyme
Does calmodulin have enzymatic activity?
No. it is just a Ca2+ sensor that, when activated by Ca2+, can affect the activity of target enzymes in the cell