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Moderator:
Dr Mohan Amberkar
G-PROTEIN COUPLED
RECEPTORS(GPCR)
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Introduction
StructureClassifcationActivation and inactivationStructure and role o G-proteins
Classifcation o G-proteinsTarets o G-proteins
-Aden!l!l c!clase
-"hospholipase C
-Ion channels -#ho A$#ho kinase
-MA" kinase#ecent developments in G"C# biolo!Conclusion#erences
PROTOCOL
%
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&rian 'obilka and #obert (eko)it*
+crucial or understandin ho) G"C#unction,
Cell surace proteinsMaor class o dru tarets
.-TM receptors/0eptahelicalreceptors/Serpentine receptors/G"(#1s22
G"C# amil!Senses the outside molecules and activates
inside the sinal trasduction path)a!cellular responses2
INTRODUCTION
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&eta adrenoreceptor in 4567
All the receptors identifed have no) been cloned
Aspirin receptor has to be cloned
Charecteristic structure:
- . transmembrane alpha helices - e8tracellular 9-terminal
- intracellular C-terminal
STRUCTURE
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;
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7
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Family Receptors Structural Features
A: Rhodopsin family -Thelargest group.
- Receptors for most amine
neurotransmitters, many
neuropeptides, purines,
prostanoids, cannabinoids, etc.
-Shortextracellular (N terminal)
tail.
-Ligand binds to transmembrane
helices (amines) or to extracellular
loops (peptides)
B: Secretin/Glucagon receptor
family
Receptors for peptide hormones,
including secretin, glucagon,
calcitonin,gastrin,CC,leptin etc
-Intermediateextracellular tail
incorporating ligand-binding
domain
C: Metabotropic glutamate
receptor/calcium sensor family
-Smallestgroup
-!etabotropic glutamate receptors,
"#$#$receptors, Ca%&-sensing
receptors
-Longextracellular tail
incorporating ligand-binding
domain
Others :D: ungal mating pheromonereceptors
!: Camp receptors
: ri""led/smoothened
receptors .
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Glutamate receptor amil!
#hodopsin receptor amil!
Adhesion receptor amil!
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5
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Sinle site mutaenesis liand mappin
desin s!nthetic
liands
X ray crystallography-molecular structureo G"C#1s
Fluorescence methods-kinetics o liandbindin
4=
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Aonist bindin"rotease activated receptors>"A#1s?
- thrombin snips o the 9-terminal tail
TET#ERED GONIST
Activation o "A#1s
"A# molecule can be activated onl! once
$et%o&s o' ctiatio"
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INCTITION
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GT" GD"?
G Protein
4
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Consists o 3 subunits-alpha/beta and amma
Alpha subunit:
-Guanine nucleotide binds to the alpha
subunit -0as en*!matic activit!
Beta and gamma subunit:
-#emain toether as a comple8
Anchored throuh an alpha att! acid chainBPREN*LTION+
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SU,T*PE SSOSITED RECEPTORS $IN EFFECTORS
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SU,T*PE SSOSITED RECEPTORS $IN EFFECTORS
Gs Man! amine and other receptors>e22 catecholamines/ histamine/serotonin?
Stimulates aden!l!l c!clase/ causinincreased cAM" ormation
Gi As or Gs/ also opioid/ cannabinoid
receptors
Inhibitsaden!l!l c!clase/ decreasincAM" ormation
Go As or Gs/ also opioid/ cannabinoid
receptors
(imited eects o subunit >eectsmainl! due to EF subunits?
G Amine/ peptide and prostanoidreceptors
Activates phospholipaseC/ increasinproduction o second messenersinositol trisphosphateanddiac!ll!cerol
GEF subunits
All G"C#s Activate potassium channelsinhibit voltae-ated calciumchannelsactivate G"C# kinasesactivate mitoen-activated proteinkinase cascade
4.
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#eceptor and e@ector selectivit!
Molecular variations )ithin the alpha subunits
Specifc reconition domains
en*!mes?
Cholera to8in "ertussis to8in
SPECIFICITY
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Cholera to8in
Acts onl! on Gs
"ersistent activation
S!mptoms o cholera
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"ertussis to8in
&lock Giand Go
"revents Gi to bind to aden!l!l c!clase
Increases cAM"
%=
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Activation o G"C#
Conormational chane in the receptor
Assosiaton o G-protein )ith the receptor
GD"-GT" e8chane
Dissosiation o the trimer
Active orms o G-protein
Di@use and assosiate )ith various en*!mes and ions channels
Activation o TA#GHT$H
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4? Aden!l!l c!clase
%? "hospholipase C
3? Ion channels
? #ho A$#ho kinase;? Mitoen-activated protein kinase>MA"
kinase?
Tarets 'or G-protei"s
%%
l l l
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Sutherland and his colleaues
+second messeners in sinal transduction,
AT" cAM" ;AM"
There are 5 di@erent molecular isoorms oaden!l!l c!clase2
e"ylyl cyclase.c$Psystem
AC "DH1s
%3
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cAM"
"rotein kinases
Jarious e@ects
Celldivision
Celldi@erentiatio
n
Iontranspo
rt
Ionchannels
Contractile
proteins
Hner!
metabolism
%
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%;
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cAM"
"rotein kinase
Increase activit! o voltae ated calcium channels in heartmuscle cells
"hosphor!lation o these channels
Increase calcium entr!
Increasin the orce o contraction o the heart
%7
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cAM"
"rotein kinases
Inactivation o m!osin liht chain kinase
Smooth muscle rela8ation
%.
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cA!P
Decrease I"crease
M% receptor o 42 "DH?
pioid receptors 2 Milrinone>"DH3?
;2Sildenafl>"DH;?
%6
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In 45;=1s b! 0okin and 0okin
Michell and &erride
- Increase in ree Ca increase "I
turnoverEg:
- muscarinic aonists
- alpha aonists actin on smooth musclesand salivar! lands
- vasopressin actin on liver cells
"I"%/member o "I amil!/ pla!s a ke! role
P%osp%olipase C.I"ositolp%osp%ate system
%5
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3=
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G"C# can act on ion channels directl!orindirectl!
Direct G-protein channel interaction )as frst
sho)n on cardiac muscleH: - mACh#s enhances 'L permeabilit!
- In neurons/ opioid analesics
open 'L channels inhibit Cachannels
ION C#NNELS
34
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Activated b! certain G"C#1s )hich couple to the G-proteinso G4%$43t!pe2
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Activated b!:
- c!tokines
- ro)th actors
- G"C# liands
Hnd result cell prolieration
"rovin to be a pathoenic path)a! or cancer
Man! compounds inhibitin this path)a!
potential anti cancer drus
H: Soraenib/ dabraenib and vemuraenib #akinase
Selumetinib/ trametinib MH' inhibitors
$P 0i"ase system
33
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Involves % main processes:
- receptor phosphor!lation
- receptor internalisation>endoc!tosis?
#esidues in the C-terminal c!toplasmic tailets phosphor!lated
Involvement o protein kinase A/protein kinaseC and G"C# kinases
DESENSITISTION
3
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3;
Rece"t &eelopme"ts i"
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GPC" dimerisation#
GA&Ab #4
H:GA&A b
GA&Ab #%
- Dopamine %
-Somatostatin receptors
Constitute$ely acti$e receptors:
- &eta adrenoreceptor:mutation in the 3rdintracellular loop or overe8pression o the receptor2
-0istamine>03? sho)s constitutive activit! in vivo2
Rece"t &eelopme"ts i"GPCR
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"eceptor acti$ating modi%yingproteins&"A!P's(#
- amil! o membrane proteins that associate)ith G"C# and alter the unctional
characteristics2H:
- 9europeptide CG#"NC#(#>G"C#? L #AM" 4
- Adrenomedullin N C#(#>G"C#? L #AM" %
3.
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"egulators o% G)protein Signalling&"GS(
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Cell penetratin peptides
Acts as intracellular modulators o sinal transmission rom aG"C# to G-proteins2
#O1 IT CTS22
It utili*es the lipid raments o intracellular G"C# loops tomodulate the G"C# action2
1#ERE IT CTS22
Anchorin to the lipid bi-la!er and thereb! taretin the interace
"epducins or over 4; di@erent G"C#1s have been successull!produced
Ananti "A# pepducin-e8tendedbleedin time in mice andprotected aainst platelet activation and thrombosis
A COC# aonist pepducin mobil*es hematopoetic stem cells tobone marro)
PEPDUCINS
35
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C*+C,-SI*+
=
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42 #an 0"/Dale MM/#itter PM/?244;5%=2
32 &arnadUttir T'/ Gloriam DH/ 0ellstrand S0/ 'ristiansson 0/ September %==7?2 Comprehensive repertoire and
ph!loenetic anal!sis o the G protein-coupled receptors in human andmouse2 Genomics33>3?: %73.32
2 Tressel S(/ 'oukos G/ Tchern!chev &/ Pacues S(/ Covic (/ 'uliopulos A>%=44?2 "harmacolo!/ biodistribution/ and eWcac! o G"C#-basedpepducins in disease models2 Methods in Molecular Biology (Clifton,N.J.)2 Methods in Molecular &iolo! 536: %;5.;
;2 Dimond "/ Carlson '/ &ouvier M/ Gerard C/ Ou (/ Covic (/ Aar)al A/ Hrnst"/ Pan* PM/ Sch)art* TR/ Gardella TP/ Millian G/ 'uliopulos A/ Sakmar
T"/ 0unt SR >Ma! %=44?2 G protein-coupled receptor modulation )ithpepducins: movin closer to the clinic2Annals of the New or! Academyof "ciences7885: 35
4
#erences
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72 "enela "/ #ibas C/ Ma!or < >9ovember %==3?2Mechanisms o reulation o the e8pression and unctiono G protein-coupled receptor kinases2 Cell."ignal.74>44?: 5.3642
.2 (uttrell (M/ (eko)it* #P >
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