Cyclic GMP Flashcards
What are the two types of guanylyl cyclase?
Soluble GC (cytosolic and NO sensitive) Particulate GC (membrane bound)
How many isoforms of pGC exist?
7 (A-G)
What domains does pGC have?
Extracellular Ligand binding domain Regulatory domain Catalytic domain Dimerisation domain (pGC exists as homodimers)
There are 3 groups of pGC what are they based on?
Their ligands
PGC A+B = bind to natriuretic peptides such as BNP which decrease TPR and increase diuresis
PGC C = binds to intestinal peptides
PGC D,E,F,G are orphan receptors
What is sGC formed from?
Hetrodimers of 1 alpha and 1 beta subunit
Multiple isoforms of the subunits exist
Catalytic activity of sGC is dependant on what?
The presence of both and alpha and beta subunit
SGC contains a heme group…why is this important?
NO (or CO) can bind here which increases endure activity by 100-200 fold
What are some functions of NO?
SM relaxation Neurotransmission Platelet aggregation Stimulates sGC Nitrosylation of proteins Neurotoxicity Induction of protein ADP ribosylation
How is NO formed?
Via Nitric oxidase synthase (nNOS, eNOS, iNOS)
This is a homodimeric protein
Why is calcium calmodulin important for NOS function?
Needed to aid/ complete the electron transport chain in the enzyme important for conversion of arginine + O2 -> citrulline +NO
What does PKG do?
Phosphorylates serine/threonine
Dependant on upstream amino acid sequence
Is PKG a monomeric protein?
No it is a homodimer
What are the types of PKG?
Type 1 (cytosolic) Type 2 (membrane bound)
What are the isoforms of type 1 PKG?
1alpha
1beta
Formed from alternative splicing
Structure of PKG monomer
N terminus
Regulatory domain - contains site of leucine zipper, cGMP binding and pseudosubstate
Hinge
Catalytic domain - contains ATP binding site and kinase area
C terminus
What does the pseudo-substrate of PKG do?
Autoinhibition via bind to kinase domain in catalytic domain area
What is the importance eof the leucine zipper?
Allows dimerisation for from PKG homodimer
What happens when cGMP binds to PKG?
4 cGMP bind to PKG dimer (2 for each regulatory domain)
Causes conformational change which releases pseudo-substrate from kinase area as monomers unfold.
Allows substrates to enter kinase domain and be phosphorylated
Name come PKG functions
Cardiac protection
Endothelial permeability
Smooth muscle relaxation
Neuronal plasticity
How does NO modulate vascular smooth muscle?
NO is made in endothelial cells in response to endogenous ligands such as bradykinin an d shear stress NO diffuses into VSMC Activates sGC Makes cGMP which activates PKG Relaxation
How does PKG mediate relaxation?
Phosphorylates myosin light kinase phosphatase which activates it and promotes dephosphorylation of myosin light chain (relaxation)
Inhibits RhoA. Since RhoA activates ROCK which intern inhibits MLCP PKG releases MLCP from inhibition
PKG activates RGS proteins which attenuate Gaq signalling via increasing their intrinsic GTPase activity thus decreasing calcium release
Phosphorylates IRAG which regulates the IP3 R on the SER = inhibits calcium release
Where is cGMP important for cycling nucleotide regulated ion channels?
Retinal rod cells
CGMP bound channels are open
Occurs during the dark state current
What isoforms of PDE degrade cGMP?
Specifically 5,6, and 9
Non specifically 1,2,3,10,11
How does cGMP regulate phosphodiesterase?
Alter rate of hydrolysis via competition for active site (PDE 1,2 and 3)
Allosteric modulation of activity (2,5, and 6)
What is the drug which inhibits PDE5?
Sildenafil
What is sildenafil used for?
Impotence
2-3 fold increase from 40 - 70
What are other causes of erectile dysfunction?
Diabetes
Post prostectomy
Hypertension medication
Trauma
How does sildenafil cause an erection?
Causes vasodilation of arterial vessels within the corpus cavernosum which allows cavernous sinuses to fill with blood
This then compresses the low pressure venous outflow which permits blood to remain in the penis 8====D
Where does the No come from which initiates the erection and prolongs the erections?
Initiation - Neural NO
Prolongation - endothelial NO
In terms of cellular processes how does an erection occur?
Cholinergic neurons activate M3 receptors on endothelial cells in repsonce to psychogenic and relfexogenic
M3 receptors are couple to Gq therefor increase in IP3 which causes calcium release
Calcium/calmodulin actives eNOS which makes NO
No defuses into SMC and forms cGMP via activation of sGC
cGMP does on to increase PKG which causes vasodilation
In terms of cellular mechanisms how does sildenafil work?
In VSMC cGMP is broken down by PDE5.
Sildenafil competes for the active site for cGMP in catalytic domain of PDE5 thus preventing it breakdown. Thus, protentates amount of PKG - more vasodilation
How can cGMP regulate PDE allosterically?
CGMP binds to allosteric binding sites on PDEif this occurs PKG can phosphorylate PDE at serine 92
This will increase activity of PDE so more is degraded
Mechanism by which levels of cGMP can promote its own degradation if levels get on high.
But how much as allosteric modulation increase PDE activity?
50-70%
How does PDE exist?
As a homodimer
What has higher affinity for cGMP in PDE 2,5 and 6…the allosteric or catalytic site?
Why is this important?
Allosteric (10-50 times greater)
Means that when cGMP levels subsequently rise again cGMP will still be bound to allosteric site. Thus PDE will be available more quickly = desentisation
What is another use of PDE5 inhibitors?
Pulmonary hypertension
PDE5 inhibition will cause redirection of blood flow to well ventilated areas for optimal ventilation perfusion ration
