Intro to Pharm and Tox Topics
Guanine nucleotide binding proteins (G proteins) serve to transduce a signal initiated by the binding of an agonist to its receptor into an intracellular effector molecule. G-proteins must be activated by binding to GTP and have intrinsic GTPase activiy which hydrolyzes GTP to GDP, returning the G-protein to its inactive state. There are two types of G-proteins: heterotrimeric and small G-proteins.
Heterotrimeric
G-proteins contain three different subunits: alpha, beta and gamma, and bind
to and are affected by 7-membrane spanning regions receptors. The alpha subunit
binds GTP and contains the intrinsic GTPase activity. The alpha subunit usually
binds the effector, although beta-gamma also seems to interact with other proteins.
There are at least 17 distinct alpha proteins ranging between 39 and 46 kD. The beta subunit binds to alpha and gamma. There are at least 4 types of beta proteins ranging in size between 35 and 36 kD. The gamma subunit ranges from 8 to 10 kD.
Inactive
G-proteins contain bound GDP and are activated when GTP is exchanged for GDP,
a process catalyzed by the active receptor. The GTPase activity slowly hydrolyzes
the bound GTP, catalyzing their own inactivation.
Active heterotrimeric G-proteins dissociate into their respective alpha and beta/gamma subunits. The dissociated alpha subunit binds to an effector molecule to start the signaling cascade. While the alpha subunit is bound to the effector, the receptor is in a low affinity state, therefore agonist molecules are less likely to activate it. After the signal is transmitted to the effector, the alpha subunit hydrolyzes GTP to GDP and returns to its resting state, bound to beta/gamma, and making the receptor have a high affinity again.
Small G-proteins are small molecules between 20 and 30 kD, with GTP binding sites and low intrinsic GTPase activity. They need guanine nucleotide exchange factors (GNEFs) to activate their GTPase activity.
In
their resting state, small G-proteins like Ras are bound to GDP and are activated
by guanine nucleotide exchange factors (GNEFs), for example Sos. GNEFs allow
Ras to bind to GTP. In the active state, Ras phosphorylates another kinase,
usually Raf, or other mitogen-activated phosphokinase kinase kinase (MAPKKK),
which in turn activates MAP kinases like ERKS, JNKS and p-38. The MAP kinases
will in turn do more phosphorylation or activate transcription factors. Since
the protein kinase activity in Ras is low, it needs a GTPase activation protein
(GAP) to activate its intrinsic GTPase activity.
There are several criteria to show that a G-protein participates in transduction:
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[Q3] [Q4] [Q5]
[Q6].
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Need more practice? Answer the review questions below (after sponsor).
1- What is the function of a G-protein
2- What is the role of GTP in G-proteins?
3- List two types of G-proteins
4- What is an heterotrimeric G-protein?
5- What are the roles of the subunits in a heterotrimeric G-protein?
6- What are the sizes of the subunits of a heterotrimeric G-protein?
7- What are small G-proteins?
8- How is a G-protein activated?
9- How is a G-protein deactivated?
10- What happens to a heterotrimeric G-protein after activation?
11- What happens to the G-protein receptor while alpha is associated with the effector?
12- What happens to a heterotrimeric G-protein after the signal is transduced?
13- How are small G-proteins activated?
14- What is Ras?
15- Describe the Ras patway.
16- What is a MAPKKK? Give one example.
17- List 3 MAP kinases activated by the Ras patway
18- How are small G-proteins deactivated?
19- List 5 criteria to show that a G-protein participates in a transduction patway.
Continue scrolling to answers below (after sponsor).
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Answers:
1- What is the function of a G-protein
Transduce a
signal initiated by agonist binding to its receptor into an intercellular effector
molecule.
2- What is the role of GTP in G-proteins?
G-proteins are activated by binding to GTP and have intrinsic
GTPase activity which hydrolyzes GTP to GDP, returning the G-protein to its
inactive state.
3- List two types of G-proteins
heterotrimeric G-proteins
small G-proteins
4- What is an heterotrimeric G-protein?
Contains 3 different subunits: alpha, beta and gamma.
Binds to and is affected by a 7-membrane spanning regions receptor.
5- What are the roles of the subunits
in a heterotrimeric G-protein?
Alpha binds GTP and contains intrinsic GTPase activity.
Alpha usually binds the effector, although beta/gamma may also interact with
proteins. Beta binds to alpha and gamma.
6- What are the sizes of the subunits
of a heterotrimeric G-protein?
About 17 distinct alpha proteins range between 39 and
46 kD, about 4 distinct beta proteins range between 35 and 36, and gamma proteins
range between 8 and 10 kD.
7- What are small G-proteins?
Small molecules between 20 and 30 kD with GTP binding
sites and low intrinsic GTPase activity.
8- How is a G-protein activated?
The inactive protein contains bound GDP and is activated
when GTP is exchanged, a process catalyzed by activated receptor.
9- How is a G-protein deactivated?
The G-protein slowly hydrolyzes the bound GTP, catalyzing
its own inactivation.
10- What happens to a heterotrimeric
G-protein after activation?
It dissociates into alpha and beta/gamma subunits. Alpha
then binds to an effector molecule to start the signaling cascade.
11- What happens to the G-protein
receptor while alpha is associated with the effector?
The receptor is in a low affinity state, agonists molecules
are less likely to activate it.
12- What happens to a heterotrimeric
G-protein after the signal is transduced?
The alpha subunit hydrolyzes GTP to GDP and returns to
its resting state bound to beta/gamma, making the receptor have high affinity
again.
13- How are small G-proteins activated?
In their resting state, small G-proteins are bound to
GDP, and are activated by guanine nucleotide exchange factors (GNEFs) that allow
the small G-protein to bind GTP.
14- What is Ras?
Ras is a small G-protein activated by SOS (a GNEF).
15- Describe the Ras patway.
Active Ras phosphorylates another kinase, which in turn
activates MAP kinases. MAP kinases in turn do more phosphorylation or activate
transcription factors.
16- What is a MAPKKK? Give one example.
Mitogen activated phosphokinase kinase kinases (MAPKKK)
are activated by Ras. Raf is an example.
17- List 3 MAP kinases activated
by the Ras patway
ERKS
JNKS
p-38
18- How are small G-proteins deactivated?
Since their intrinsic GTPase activity is low, a GTPase
activation protein (GAP) is needed to activate the GTPase and allow deactivation
of the small G-protein.
19- List 5 criteria to show that
a G-protein participates in a transduction patway.
- Resistant analogs of GTP weaken ligand binding
- GTP binding is stimulated by agonist
- Activity of an effector is altered by agonist and GTP
- Pertussin toxin blocks some G-proteins
- Cholera toxin activates some G-proteins
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