G Protein Coupled Receptors Flashcards
G-protein coupled receptors
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GTP-binding protein pool
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G-protein cycle
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Downstream effectors
G proteins couple to specific effectors:
Gq/11 = increased phospholipase C activity
Gs = increased adenylyl cyclase activity
Gi = decreased adenylyl cyclase activity
G12/13= regulates GTP exchange factors
A role for beta gamma subunits
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Simple view:
1)Inert receptor waiting
2)Agonist arrives
3)Biological effect
4)Termination (then 1)
Signal Adaption
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Desensitisation
*G-protein receptor kinases (GRK1-3)
*High affinity for free bg subunits
*Phosphorylate GPCR
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Role of G-protein-linked receptor kinase
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b-arrestin associates with many proteins
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Biased signalling definition
The ability to direct a G protein–coupled receptor to
selectively signal through a G protein–mediated pathway or a β-arrestin–mediated pathway.
GPCRs may NOT always signal through G proteins!
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Producing b-arrestin biased ligands for COVID
Hyperactivation of the Ang II type 1 receptor (AT1R) is major contributor to adverse outcomes in patients with COVID-19–related acute respiratory distress syndrome (ARDS),
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Receptor activity
modifying proteins (RAMPs)
Functional GPCRs are not always a single polypeptide.
Calcitonin family
The calcitonin family of peptides comprises :
-calcitonin
-amylin
-two calcitonin-gene-related peptides (CGRP1 and
CGRP2)
-adrenomedullin (ADM)
Effective modulators of biological activity
Receptors uncloned
CGRP receptors
Early studies revealed the existence of two “receptors”: CGRP1 and CGRP2.
The linearized CGRP analog [Cys(ACM)2,7]hCGRP was
found to be a potent agonist in the rat vas deferens bioassay, but not guinea pig atrial preparations.
Truncated peptide fragments hCGRP12–37 and CGRP8–37 had antagonist properties in atrial preparations but not in the vas deferens.
CGRP receptors
Discovery of a “calcitonin receptor-like receptor” (CLR)
unresponsive to CGRP stimulation when expressed in cell lines.
In human embryonic kidney (HEK293) cells cloned CRLR
CGRP produced a 60-fold increase in cAMP generation following blocked by CGRP8–37.
HEK293 cells endogenously express a protein that, when coexpressed with CLR, responded to CGRP.
Receptor activity modifying protein or RAMP
small (148-175-amino acid) proteins
single predicted membrane-spanning domain
large extracellular domain
short cytoplasmic domain
Summary of reported roles for RAMPS
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RAMP determines receptor
pharmacology
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The RAMP/Receptor Paradigm for AM and
CGRP Signaling
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dimerization
Functional GPCRs are not always a single polypeptide
Dimer formation in GPCRs
In neurones GABA produces rapid responses
through GABAA R and slower responses
through GABAB.
But expression of the gene encoding a GABAB
receptor (GABABR1) = fails to recreate native receptor
Dimer formation in GPCRs pt 1
*immature glycoprotein on intracellular membranes
*low affinity for agonists compared with the
endogenous receptor
However co- expression with GABABR2 in HEK293T cells =
*GABABR1 is terminally glycosylated
*expressed at the cell surface.
*binds GABA with a high affinity equivalent to that
of the endogenous receptor.
Dimer formation in GPCRs pt 2
The type C or III class of GPCR includes metabotropic glutamate, GABAB, calcium sensing receptor, pheromone and taste receptors.
In addition to the typical GPCR HD (heptahelical
domain), class-III GPCRs possess a large ECD
(extracellular domain) responsible for ligand
recognition
All members need to form DIMERS to be fully
functional
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Calcium sensing receptor
Type 1 GPCR
eg adrenoceptor
a multimodal modulator protein
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Dimer formation in GPCRs- Not just C type
AngII augments PGF contraction.
PGF augments Ang II contraction
Convergence of downstream Gαq-PLC-Ca2+ pathway?
L158, 809 = AngII Receptor Antagonist
AL-8810 & AS604872 = FP Receptor Antagonist
Symmetrical and asymmetrical regulation of
AT1R/FP signaling likely through the agency
of a heterodimer.
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Functional GPCRs agonists derived from the receptor protein
Soluble agonist
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Receptor Activator
PAR1 th > tryp
PAR2 trypsin
PAR3 thrombin
PAR4 tryp > thr
tethered ligand
SFLLRN (h) or SFFLRN (m, r)
SLIGKV (h) or SLIGRL (m, r)
TFRGAP (h) or SFNGGP (m, r)
GYPGQV (h) or GYPGKF (m, r)
Constitutive activity
The work on inverse agonist has led to the acceptance that many , if not ALL GPCRs are actually active in the absence of ligand.
The ligand tips the balance towards greater
turnover / greater coupling
