Module 2 Enzymes and Cell Biology 5 of 6

Biochemical Signalling - 2
G proteins and GPCRs

Shoba Ranganathan
Dept. of Molecular Sciences
4 Wally’s Walk (Building F7B), Room 121
T: 02 9850 6262; E: shoba.ranganathan@mq.edu.au
Objectives

• Receptors that bind hormones (continued)

 G Proteins and their receptors, GPCRs
 cAMP Pathway
 Phosphoinositide Pathway
 Drugs and Toxins affecting Cell Signalling
• Complexity of Hormonal Signalling

VVP5 Chap.13

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Main classes of hormonal receptors
A. G-Protein-linked receptors or G-
Protein Coupled Receptors (GPCRs)
– this lecture

B. Enzyme-linked receptors: e.g.

Receptor Tyrosine Kinases.

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Signalling by G Proteins – 3
components
1. The extracellular signalling molecule binds
to G-protein-coupled receptors (GPCRs),
which are transmembrane proteins.
2. G proteins are heterotrimers that are
anchored to the cytoplasmic side of the
membrane.
3. Adenylate cyclase is a transmembrane
enzyme that generates the secondary
messenger, cAMP (cyclic AMP).

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Steps in G protein signalling
1

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What are GPCRs?
• Largest known protein families with >800 members in
the human genome.
• Essential sensory functions: sight, smell and taste.
• Ligands include:
 nucleosides, nucleotides,
 Ca2+,
 catecholamines (adrenalin and nonadrenalin), other
bioactive amines (e.g., histamine and serotonin) and a
variety of peptide and protein hormones, and
 Lipids.

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7 transmembrane helices (7TM) in
GPCRs

Human
β2‐adrenergic 
receptor
PDBid 2RH1

Ligand binding 
site is located
between 
the helices

Helices move
when ligand 
binds.

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The α-subunit of G protein binds to the
intracellular part of GPCR

Heterotrimeric G protein
PDBid 1GP2

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Activation of G protein
• In the inactive state, the α-subunit binds GDP
(guanosine diphosphate).
• When an extracellular ligand binds to the GPCR, the
GPCR structure twists (allostery).
 The α-subunit exchanges its bound GDP for GTP and
gets activated.
 The bound trimeric G protein gets detached and the
subunits separate.
 The activated Gα-subunit then binds and activates
adenylate cyclase, for making cAMP.
 The G protein then hydrolyses the bound GTP back to
GMP.

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 Adenylate Cyclase (AC) Synthesises
the Secondary Messenger, cAMP
• Activated Gα-
subunit binds to
AC.
• AC gets
activated and
converts ATP to
cAMP.
• Inactivation of
the Gα-subunit
makes AC
inactive.

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cAMP

• cAMP targets soluble protein kinases and activates

them to phosphorylate other proteins – this is the end
result of GPCR signalling.
• These phosphorylated proteins are inactivated by
phosphatases.

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Ending cAMP signalling
• The secondary messenger, cAMP, has to be hydrolysed to
AMP, to end the signalling process.
• Enzymes, cAMP-phosphodiesterases (PDEs), step in to do
this. PDEs are activated by
 Ca2+

 Phosphorylation by PKA and insulin-stimulated PKA.

• PDEs provide crosstalk between cAMP-based and other

signalling systems.
• Drugs that are PDE inhibitors used for asthma, congestive
heart failure, depression, inflammation, retinal degeneration
and erectile dysfunction (Viagra).

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Drugs affect Cell Signalling

• Adenylate cyclase system affected by

adenosine analogues in coffee, tea and
chocolate (xanthine alkaloids).
• Antagonise adenosine
receptors, through
inhibitory G proteins
• Lead to excess cAMP
production
• Results in downstream
PKA activation.

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Toxins affect Cell Signalling
• Bacterial toxins affect G protein function
• A fragment of cholera toxin causes a side
chain modification on Gα, forming Gsα.
 cAMP is produced.
 GTP cannot be hydrolysed by Gsα.
 Fluid influx in intestines, due to uncontrolled cAMP
levels.
 Dehydration.
• Similar but less dramatic effect from E. coli
through heat-labile enterotoxins produced.

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Toxins affect Cell Signalling
• Pertussis toxin (from whooping cough agent)
causes a diffierent side chain modification on
Gα, forming Giα.
 Giα cannot exchange bound GDP for GTP
 Once again cAMP levels go unchecked.
 ~400,000 infant deaths worldwide.
 (prevented by triple antigen, DTP vaccine).

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 The adenylate cyclase signalling
system

• Toxins affect
G protein
function

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Alternative to cAMP Signalling: the
Phosphoinositide Pathway
• Ingredients: GPCR, heterotrimeric G protein,
a specific phospholipase (PLC) and a
phosphorylated glycerophospholipid.
• Three secondary messengers generated:
 IP3: inositol-1,4,5-triphosphate
 Ca2+ and
 DAG: diacyl-glycerol. This could differ, based on
the phospholipid that is hydrolysed.
 Lipid soluble and activated membrane bound
PKC, which triggers a cascade.
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 Phosphoinositide Signalling

• DAGs are produced from

glycerophospholipids by
hydrolysis (Lecture 9)
• Important for lipid
synthesis and breakdown
(later lectures) H

• Implications for obesity

and fat burning
• Involved in several
cancers, especially in
melanoma.

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Phosphoinositide Signalling System

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Complexity of hormone signalling

• Hormones have multiple roles in the cell

• They can trigger multiple signalling pathways,
involving
 metabolism as well as
 cellular growth and differentiation
• Example: insulin
 Glucose transport
 Glycogen synthesis
 DNA/RNA/Protein synthesis

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 Insulin signalling cascade

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GP/GPCR Signalling Summary
• G protein–coupled receptors (GPCRs) contain seven
membrane-spanning helices and undergo
conformational changes when a hormone binds.
• Ligand binding to a GPCR induces the α subunit of
the associated G protein to exchange GDP for GTP
and dissociate from the β and y subunits.
• Adenylate cyclase is activated to produce cAMP,
which in turn activates protein kinase A.
• Signalling activity is limited through the action of
phosphodiesterases that act on cAMP and cGMP.
• GPCRs can also signal through the phosphoinositide
pathway.

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Spot test: Quiz on iLearn!

You can always do the Practice quizzes as

well or instead of the Spot tests.

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