Lecture 3 Part 2 Flashcards
Gs and other G proteins use a molecular mechanism that involves what?
the binding and hydrolysis of GTP
what does GDP and GTP stand for?
GDP = guanosine diphosphate
GTP = guanosine triphosphate
when the GPCR (G protein coupled receptor) is NOT ACTIVATED by a ligand, what is it associated with?
guanosine diphophate (GDP)
when the GPCR IS activated by a ligand, what happens?
GDP is released and GTP enters the nucleotide binding site of the G protein
what happens after the GPCR has been activated by a ligand and GDP has been released and GTP has entered the nucleotide binding site of the G protein?
the activated G protein (now bound to GTP) migrates from the receptor and goes to regulate the activity of an EFFECTOR ENZYME or an ION CHANNEL
what are the potential effectors of an activated G protein (one bound to GTP)
-an effector enzyme
-an ion channel (E)
as mentioned, an activated G protein can regulate the activity of either an effector enzyme or an ion channel (E)
what stops the signal?
the hydrolysis of GTP stops the signal and E active –> E
the G protein is now again bound to GDP and couples back with the receptor to await activation
(1 inorganic phosphate is released from GTP-> GDP)
true or false
when the G protein is bound to GDP, it is said to be in an inactive state
true
true or false
once the ligand has activated the G protein to release its GDP and bind to GTP, it doesn’t matter whether or not the receptor ligand is still bound or free
TRUE
what is the rate limiting step in the termination of effector activation in the case of GPCRs?
the hydrolysis of GTP to GDP
what determines how long the effect produced by a GPCR will last?
the rate limiting step – hydrolysis of GTP to GDP
true or false
cGMP is not a second messenger
false - it is
true or false
NE (norepinephrine) works via a G-protein coupled receptor
true - it’s a cetecholamine
explain how the rate limiting step for the effect of a G protein is not determined by how long the ligand is bound to the receptor
a neurotransmitter like norepinephrine may only encounter its receptor for milliseconds.
but, when the Gs protein coupled to the NE receptor binds GTP and is activated, it may remain active for TENS OF SECONDS – enormously amplifying the original signal
which 3 subfamilies of G proteins are most important and usual for drugs?
Gs
Gi
Gq
Gs = G stimulatory
Gi = G inhibitory
Gq
name the ligands for Gs
beta-adrenergic amines
glucagon
histamine
serotonin
other HORMONES
Further classify Gi subfamily
Gi1
Gi2
Gi3
name the ligands for Gi
alpha2-adrenergic amines
acetylcholine (muscarinic)
opioids
serotonin
others
name the ligands for Gq
acetylcholine (muscarinic)
serotonin (5-HT1C)
many others
state the effector/signaling path for Gs
increased adenylyl cyclase activity
increased cAMP and calcium channel activation
state the effector/signaling path for Gi
decreased adenylyl cyclase
decreased cAMP
cardiac K+ channels open, decreases heart rate
what are the ligands for Golf
odorants
state the effector/signaling path for Golf
decreased adenylyl cyclase and decreased cAMP
what are the ligands for Go
neurotransmitters in the brain
state the effector/signaling path for Go
not clear
State the effector/signaling path of Gq
increased cytoplasmic calcium
all 2nd messengers increased
DAG
IP3
phospholipase c activity
what are the ligands for Gt(1-2)
photons
rhodopsin - retinal rods
color opsonins - cone cells
state the effector/signaling path of Gt(1-2)
increased cGMP phosphodiesterase
decreased cGMP (phototransduction)
some GPCR’s are also called ___________. why?
Serpentine receptors bc the receptor polypeptide chain resembles a snake
true or false
“serpentine” receptors are GPCR’s
true
serpentine receptors are GPCR’s and are often called….
7 transmembrane spanning receptors
explain the structure of serpentine receptors
extracellular amino terminus (N) NH2
intracellular carboxy terminus (C) COOH
spans the membrane 7 times
consists of extracellular and intracellular loops
true or false
all GPCR’s are serpentine receptors
false - only some
in serpentine receptors, is the amino group extracellular or intracellular?
extracellular
carboxy group (COOH) is intracellular
name 2 potential ligands for serpentine receptors.
where does the ligand bind?
catecholamines and acetylcholine
binds within the pocket formed by the 7 transmembrane regions
after the ligand binds within the pocket of the serpentine receptor, what happens?
there is a conformational change in the receptor.
this is sensed by the loops within the cytoplasm and the G protein gets activated (GDP becomes GTP) and dissociates from the receptor
GTP gets hydrolyzed back to GDP and the G protein-GDP complex is INACTIVE and couples again with the receptor
in serpentine GPCR, what may be associated with diminished interaction between the receptor and G protein?
the cytoplasmic tail of the GPCR contains SERINE and THREONINE residues.
these serine and threonine residues can be phosphorylated and this phosphorylation may be associated with diminished interaction between receptor and G protein
_________ responses to drugs and hormonal agonists often DESENSITIZE with time
receptor-mediated responses
true or false
desensitization is usually irreversible
false - usually reversible
define desensitization
occurs when even though the receptors are saturated with agonist, the response gradually decreases
true or false
many kinds of receptors undergo desensitization
true
true or false
although many kinds of receptors undergo desensitization, the mechanism is not clear in most cases
TRUE - like the nicotinic acetylcholine receptor
the beta-adrenoceptor desensitization mechanism, however, has been worked out in some detail
explain the mechanism of beta-adrenoceptor desensitization
when the ligand binds, Gs-GTP and bARK (b-adrenergic receptor kinase) is activated.
bARK PHOSPHORYLATES the cytoplasmic tail on the serine and threonine residues. this increases the affinity of b-arrestin to bind
b-arrestin (protein) then binds to the cytoplasmic tail. this results in a diminished ability for the receptor to interact with the G protein. THUS, THE RESPONSE TO THE AGONIST IS REDUCED
In the desensitization mechanism of the b-adrenoceptor, what happens when the agonist is removed?
-phosphatases cleave the phosphates from the cytoplasmic tail (originally placed there by bARK)
-bARK activity is decreased (stops phosphorylating the tail of the receptor)
-receptor has now “reset” itself and is now ready to fully respond to the agonist
in the desensitization mechanism for b-adrenoceptor, why is it that even though the ligand (agonist) is bound, the response will be decreased
the G protein-GDP can’t be activated because it can’t bind to the 3rd intracellular loop of the receptor – it is occupied by b-arrestin
true or false
once the ligand dissociates from the receptor, bARK is inactivated
true
when agonist is NOT bound to the receptor, explain what is happening to the beta-adrenoceptor
the inorganic phophates are released, beta-arrestin dissociates from the cytoplasmic tail, and the G protein-GDP complex can now couple with the receptor
the receptor has now “reset” itself and is ready again to fully respond to the agonist
name 3 well established second messengers
cAMP
Ca2+
cGMP
what does cAMP stand for
cyclic adenosine monophosphate
true or false
cAMP is a second messenger
true
when and where is cAMP released?
released in the cytoplasm due to hormone binding
explain the function of 2nd messengers
they can activate many cell activities leading to a large scale, coordinated response.
2nd messenger mechanism - cAMP
-agonist binds
-this activates Gs and Adenylyl Cyclase. ATP converted to cAMP by Adenylyl cyclase.
cAMP converted to AMP by PDEs (phosphodiesterases)
cAMP kinase: consists of catalytic region and regulatory region. cAMP binds to the regulatory sites and the catalytic regions are released.
catalytic regions + ATP will phosphorylate the substrate leadinf to a CELLULAR RESPONSE:
-enzyme activation
-protein synthesis
-muscle relaxation
-nerve stimulation
-hormone secretion
what is the function of AC in the mechanism of second messengers
AC = adenylyl cyclase
converts ATP to cAMP
what does cAMP get converted to and by what
cAMP is converted to AMP by PDEs (phosphodiesterases)
explain the structure and role of cAMP kinase
consists of 2 regulatory regions and 2 catalytic regions.
cAMP binds to the regulatory sites which releases the catalytic regions.
these catalytic regions + ATP phosphorylate the substrate to produce a cellular response
explain the cellular response produced by cAMP
-enzyme activation
-protein synthesis
-muscle relaxation
-nerve stimulation
-hormone secretion
true or false
cAMP causes muscle stimulation
false - muscle relaxation
true or false
cAMP causes nerve stimulation
true
true or false
cAMP causes enzyme activation
true
true or false
cAMP causes protein degradation
false - protein synthesis
true or false
cAMP causes hormone secretion
true
what is the mechanism of viagra
phosphodiesterase inhibitor.
this will prevent the conversion of cAMP to AMP
this will increase cAMP causing an increase in downstream signaling and vasodilation (muscle relaxation)
in the mechanism of cAMP, what happens when the agonist dissociates?
the intracellular actions of cAMP are terminated by 3 mechanisms:
-GTP hydrolyzed back to GDP
-the activation of cAMP kinase is inhibited. the Tetramer is re-established
-the phosphorylated substrate (generated by cAMP kinase) is dephosphorylated (BY PHOSPHATASES), causing a DIMINISHED CELLULAR RESPONSE
What is the effect of caffeine on cAMP levels?
caffeine acts as a competitive inhibitor of phosphodiesterases, thus INCREASING CAMP LEVELS
Besides caffeine, what drugs increase cAMP levels?
theophylline (bronchodilator used for asthma treatment)
methylxanthine drugs
what substances are COMPETITIVE INHIBITORS of PDEs? (phosphodiesterases)
caffeine and theophylline
methylxanthine drugs are non competitive inhibitors of phosphodiesterases
true or false
caffeine, theophylline, and methylxanthine drugs all increase intracellular cAMP levels
true
name 5 effects of methylxanthine drugs
- increased CNS activity (increased alertness, deferral of fatigue)
-increase in inotropic effect of the heart (increase force)
-increased gastric acid secretion and increased enzymes (in GI)
- decreased Na reabsorption and increased glomerular filtration (thus they are weak diuretics!)
-increased bronchodilation (used for asthma treatment!)
true or false
methylxanthines decrease heart contraction
false - increase
true or false
methylxanthines increase gastric secretion and enzymes in the GI tract
true
true or false
methylxanthines decrease sodium reabsorption
true
also increase glomerular filtration – weak diuretics
what effect do methylxanthines have on the CNS?
increased CNS activity leading to increased alertness and deferral of fatigue
what is(are) the second messenger(s) involved in Gq coupled receptors
calcium and phosphoinositides
name 4 receptors that are Gq coupled
acetylcholine (muscarinic)
alpha1-adrenergic
platelet activating factor
serotonin (5-HT1C and 5-HT2)
When a ligand is bound to Gq, what 2nd messenger is likely involved?
calcium
explain the mechanism of calcium as a second messenger
agonist binds, Gq protein is stimulated and binds GTP.
PLC activates PIP2 which activates DAG and IP3. IP3 releases CALCIUM. this calcium binds and activates CALMODULIN.
calmodulin (with bound calcium) then stimulates signaling through calcium/calmodulin dependent protein kinases to PHOSPHORYLATE THE SUBSTRATE AND PRODUCE A CELLULAR RESPONSE
meanwhile, while IP3 released calcium, DAG remains in the membrane where it binds and activates PKC (protein kinase C) which phosphorylates certain substrates to produce a cellular response – INCREASED GLAND SECRETION, CELL GROWTH AND DIFFERENTIATION, AND INCREASED METABOLISM
how does PKC become active and what happens when it is activated
DAG binds and activates PKC.
PKC can then phosphorylate certain substrates to produce a cellular response
what cellular responses occur when PKC phosphorylates its substrates
increase gland secretion
increased cell growth and differentiation
increased metabolism
what does cGMP stand for
cyclic guanosine-3’,5’-monophosphate
explain where cGMP plays a role as a second messenger
only in a few cell types like intenstinal mucosa and vascular smooth muscle
explain whne cGMP-based signal transduction is initiated.
in which cells does this occur?
in intestinal mucosa and vascular smooth muscle when:
-ligand binds to the extracellular domain of the receptor
-ligand binding stimulates intracellular guanylyl cyclase activity
-cGMP activates cGMP-dependent protein kinases
what is cGMP produced from and how?
GTP becomes cGMP through guanylyl cyclase
guanylyl cyclase activity is stimulated when the ligand binds to the extracellular domain of the receptor
what does cGMP do
it activates cGMP dependent protein kinases
these activated kinases then phosphorylate the substrate
increased cGMP causes….
relaxation of vascular smooth muscle
HOW does cGMP cause the relaxation of vascular smooth muscle?
by a kinase-mediated mechanism that results in the DEPHOSPHORYLATION of myosin light chains
name 2 specific things that will result in increased cGMP accumulation
-ANP binds to its receptor which increases guanylyl cyclase activity. guanylyl cyclase converts GTP to cGMP
-lipid souble gas NO (nitric oxide) is released by nearby vascular endothelial cells which DIRECTLY ACTIVATES guanylyl cyclase (binds to it and activates)
several _____ drugs mimic NO (nitric oxide)
vasodilator
