GPCR will continue to be highly important in clinical medicine because of their large number, wide expression and role in physiologically important responses. Future discoveries will reveal new GPCR drugs, in part because it is relatively easy to screen for pharmacologic agents that access these receptors and stimulate or block receptor-mediated biochemical or physiological responses.

1. G-Protein Coupled Receptors and Secondary
Messenger Pathways
Presentation By;
Mr. Saikat Polley
M. Pharm 1st Semester
Registration No. 211592320210002 of 2021-22
Roll No. 15920221005
Department of Pharmacology
Calcutta Institute of Pharmaceutical Technology & AHS
Banitabla, uluberia,howrah-711316, West Bengal

2. Contents
Introduction
Structure of GPCR
Signal Transduction by GPCR
Inactivation or Termination of Response
Secondary Messenger cAMP Pathway
Phosphoinositide Signaling Pathway
Disease linked to G-protein Pathways
Conclusion
References

3. Introduction
 G proteins, also known as guanine nucleotide-binding proteins, are a family of
proteins that are involved in transmitting signals from a variety of stimuli outside
a cell to its interior. G proteins belong to the larger group of enzymes called GTPases.
 Heterotrimeric G proteins were discovered, purified, and characterized by Martin
Rodbell and his colleagues at the National Institutes of Health and Alfred Gilman and
colleagues at the University of Virginia.
 G protein-coupled receptors (GPCRs) are so named because they interact with G
proteins. Members of the GPCR superfamily are also referred to as seven-
transmembrane (7TM) receptors because they contain seven transmembrane helices.
 Humans express over 800 GPCRs that make up the third largest family of genes in
humans.
 Thousands of different GPCRs have been identified in organisms ranging from yeast to
flowering plants and mammals that together regulate an extraordinary spectrum of
cellular processes.

4.  GPCRs share a common structural signature
of seven hydrophobic transmembrane (TM)
segments, with an extracellular amino
terminus and an intracellular carboxyl
Terminus
 The structure of a GPCR can be divided into
three parts:
1. The extra-cellular region, consisting of
the N terminus and three extracellular
loops (ECLI1-ECL3).
2. The TM region, consisting of seven α
helices(TM1-TM7).
3. The intracellular region, consisting of
three intra-cellular loops (ICL1-ICL3),
an intracellular amphipathic helix
(H8), and the C terminus .
Structure of GPCR
Fig 1: General Structure of GPCR

5.  Basic signal transduction cascade
 G protein coupled receptors contain
Seven membrane spanning helices.
 When bound to their ligand, the receptor
interacts with a trimeric G protein, which
activates an effector, such as adenylyl
cyclase.
 The alpha and gamma subunits of the G
protein are linked to the membrane by
lipid groups that are embedded in the
lipid bilayer.
Fig 2: A GPCR and a G-Protein

6. The mechanism of receptor-mediated activation (or
inhibition) of effectors by means of heterotrimeric G
proteins.
 The ligand binds to the receptor, altering its
conformation and increasing its affinity for the G
protein to which it binds.
 The alpha subunit releases its GDP, which is replaced
by GTP.
 The alpha subunit dissociates from the G(beta and
gamma) complex and binds to an effector (in this case
adenylyl cyclase), activating the effector.
 Activated adenylyl cyclase produces cAMP.
 The GTPase activity of alpha subunit hydrolyzes the
bound GTP, deactivating alpha subunit.
 G(alpha) re-associates with G(beta and gamma),
reforming the trimeric G protein, and the effector
ceases its activity.
 The receptor has been phosphorylated by a GRK.
 The phosphorylated receptor has been bound by an
arrestin molecule, which inhibits the ligand bound
receptor from activating additional G proteins.
 The receptor bound to arrestin is likely to be taken up
by endocytosis.
Fig 3: Activation of GPCR

7. Inactivation or Termination of Response
 Desensitization, the process that blocks active
receptors from turning on additional G
proteins, takes place in two steps. In the first
step, the cytoplasmic domain of the activated
GPCR is phosphorylated by a specific type of
kinase, called G- protein-coupled receptor
kinase (GRK )
 GRKs form a small family of serine-threonine
protein kinases that specifically recognize
activated GPCRs.
 Phosphorylation of the GPCR sets the stage
for the second step, which is the binding of
proteins, called arrestins. Arrestins form a
small family of proteins that bind to GPCRs
and compete for binding with heterotrimeric G
proteins.
 As a consequence, arrestin binding prevents
the further activation of additional G proteins.
Fig 4: Termination of response

8.  Cyclic AMP formation
Secondary Messenger cAMP Pathway
Cyclic AMP formation
usually depends upon the
activation of G-protein-
coupled receptors (GPCRs)
that use heterotrimeric G
proteins to activate the
amplifier adenylyl cyclase
(AC), which is a large
family of isoforms that
differ considerably in both
their cellular distribution
and the way they are
activated. There are a
number of cyclic AMP
signalling effectors such as
protein kinase A. Fig 5: Formation of cyclic AMP

9.  Adenylyl Cyclase or AC
 Adenylyl cyclase is an integral polyphyletic
protein of the plasma membrane, with its
active site on the cytosolic face.
 The enzyme synthesizes cyclic adenosine
monophosphate or cyclic AMP from adenosine
triphosphate (ATP). Cyclic AMP functions as a
second messenger
 The best known class of AC is class III.
 Protein Kinase
 Protein kinase, is an kinase type of enzyme that
modifying other proteins by adding phosphate
group to them (phosphorylation).
 PKA is composed of two regulatory (R)
subunits and two catalytic (C) subunits.
 There is two types of PKA, protein kinase A
(PKA) I is found mainly free in the cytoplasm
and has a high affinity for cyclic AMP, whereas
protein kinase A (PKA) II has a much more
precise location by being coupled to the A-
kinase-anchoring proteins (AKAPs).
Fig 6: Structure of adenylyl cyclase
Fig 7: Protein kinase A activation

10. Phosphoinositide Signaling Pathway
 Phospholipids of cell membranes are converted into
second messengers by a variety of enzymes that are
regulated in response to extracellular signals.
 Phospholipases are enzymes that hydrolyze specific
ester bonds that connect the different building blocks
that make up a phospholipids molecule.
 Phospholipase C( PLC), which splits the
phosphorylated head group from the diacylglycerol.
Fig 9: IP3-DAG Pathway
Fig 8: Structure of a phospholipid

11.  Phospholipase C
 When acetylcholine binds to a
smooth muscle cell, or an antigen
binds to a mast cell, the bound
receptor activates a heterotrimeric
G protein, which, in turn, activates
the effector phosphatidylinositol-
specific phospholipase C-β (PLC
β) (step 4).
 PLC β is situated at the inner
surface of the membrane, bound
there by the interaction between its
PH domain and a phosphoinositide
embedded in the bilayer.
 PLC β catalyzes a reaction that
splits PI(4, 5)P2 into two
molecules, inositol 1,4,5-
trisphosphate (IP3 ) and
diacylglycerol (DAG) (step 5) both
of which play important roles as
second messengers in cell
signaling.
Fig 10: Activation of phospolipase C

12.  Diacylglycerol (DAG)/protein kinase C (PKC) signalling
 The diacylglycerol (DAG) that is formed when PtdIns4,5P2 is hydrolysed by
phospholipase C (PLC) remains within the plane of the membrane where it acts as a lipid
messenger to stimulate some members of the protein kinase C (PKC) family.
 The protein kinase C family has been divided into three subgroups.
• Conventional PKCs (cPKCs: α, β1, β2 and γ) contain C1 and C2 domains
• Novel PKCs (nPKCs: δ, ε, η and θ) contain a C1 and a C2-like domain
• Atypical PKCs (aPKCs: ζ , ι and λ) contain a short C1 domain the regulation of
specific cellular processes.
Fig: Protein Kinase C family

13.  Inositol 1,4,5-trisphosphate (InsP3)/Ca2 + signalling
 Inositol 1,4,5-trisphosphate
receptor (IP3R) is a widely
expressed channel for
calcium stores.
 After being activated by
inositol 1,4,5-trisphosphate
(IP3) and calcium signalling
at a lower concentration,
IP3R can regulate the
release of Ca2+ from stores
into cytoplasm, and
eventually trigger
downstream events
Fig: IP3 signalling

14. Disease linked to G-protein Pathways

15. Conclusion
 The GPCR family includes receptors that are responsible for the recognition of light, taste,
odour, hormones, pain, neurotransmitters and many other things. Or in other words, most
physiological processes are based on GPCR signaling. This is why the GPCR family is of
huge pharmaceutical importance.
 Nearly 40% of the drugs approved for marketing by the FDA target GPCRs
 800-1,000 different GPCRs and the drugs that are marketed target less than 50 GPCRs
 GPCR will continue to be highly important in clinical medicine because of their large
number, wide expression and role in physiologically important responses
 Future discoveries will reveal new GPCR drugs, in part because it is relatively easy to
screen for pharmacologic agents that access these receptors and stimulate or block receptor-
mediated biochemical or physiological responses

16. References
1. Fredriksson R, Lagerstorm M.C et al. The G protein-coupled receptors in the human
genome from five main families. Phylogenetic analysis, Mol. Pharmacol. 2003; 63:
1256-72.
2. Rang HP, Dale MM, Ritter JM, Flower RJ. PHARMACOLOGY. Churchill
livingstone elsevier. 2008; 6: 35-40.
3. Arac D, Boucard A, Bollinger M.F. et al. A novel evolutionary conserved domain
of cell-adhesion GPCRs mediated auto-proteolysis. The EMBO Journal. 2012; 31:
1364-78.
4. Karp Gerhald. CELL BIOLOGY. International student version. 2014; 7: 622-34.
5. Costanzi S. Homology Modelling of Class A G Protein-Coupled Receptors. Methods
in Molecular Biology. 2012; 857: 259-79.
6. AbdAlla, S., Lother, H., el Missiry, A., Langer, A., Sergeev, P., el Faramawy, Y., et al.
Angiotensin II AT2 receptor oligomers mediate G-protein dysfunction in an animal
model of Alzheimer disease. J. Biol. Chem. 2009; 284: 6554–65.