BioSci M160 / MolBio255 Structure-Function Relationships of Integral Membrane Proteins Lecture 6

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BioSci M160 / MolBio255Structure-Function
Relationships of Integral Membrane
ProteinsLecture 6
Hartmut Hudel Luecke Biochemistry, Biophysics
Computer Science Email hudel_at_uci.edu
http//bass.bio.uci.edu/hudel
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G Protein - Coupled Receptors (GPCRs)

• GPCRs constitute the largest gene family in the
  human genome (950 genes), 500 of which are
  sensory.
• They are the target of the majority of
  best-selling drugs (40-50 of all prescription
  pharmaceuticals on the market). Examples
• Zyprexa (bipolar disorder schizophrenia, Eli
  Lilly)
• Clarinex (antihistamine against seasonal
  year-round allergies, Schering-Plough)
• Zantac (treat and prevent ulcers in the stomach
  and intestines histamine receptor antagonists,
  GlaxoSmithKline)
• Zelnorm (severe, chronic, irritable bowel
  syndrome (IBS) increases the action of serotonin
  in the intestines, Novartis)

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Receptor Activation
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GPCR Ligands

• Three types of ligands
• agonists are ligands which shift the equilibrium
  in favor of the active state.
• inverse agonists are ligands which shift the
  equilibrium in favor of inactive states.
• neutral antagonists are ligands which do not
  affect the equilibrium.

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Receptor Activation
If a GPCR (red) in an active state (activated by
binding of orange ligand) encounters an inactive
G protein (blue, with bound GDP), it may activate
the G protein. Activated G proteins have a
reduced affinity for GDP and an increased
affinity for GTP, leading to GXP exchange.
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Receptor Activation II
The enzyme adenylate cyclase (green) is an
example of a membrane protein that can be
regulated by a G protein. Adenylate cyclase is
activated when it binds the alpha subunit of the
activated G protein. Activation of adenylate
cyclase ends when the G protein returns to the
GDP-bound state.
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GPCR Function

• The visual sense the opsins use a
  photoisomerization reaction to translate
  electromagnetic radiation into cellular signals.
  Rhodopsin, for example, uses the conversion of
  11-cis-retinal to all-trans-retinal for this
  purpose.
• The sense of smell receptors of the olfactory
  epithelium bind odorants (olfactory receptors)
  and pheromones (vomeronasal receptors).
• Behavioral and mood regulation receptors in the
  mammalian brain bind several different
  neurotransmitters, including serotonin and
  dopamine.
• Regulation of immune system activity and
  inflammation chemokine receptors bind ligands
  that mediate intercellular communication between
  cells of the immune system receptors such as
  histamine receptors bind inflammatory mediators
  and engage target cell types in the inflammatory
  response.
• Autonomic nervous system transmission both the
  sympathetic and parasympathetic nervous systems
  are regulated by GPCR pathways. These systems are
  responsible for control of many automatic
  functions of the body such as blood pressure,
  heart rate and digestive processes.

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GPCR Membrane Topology
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GPCR Classes
Class A Rhodopsin like Class B Secretin
like Class C Metabotropic glutamate /
pheromone Class D Fungal pheromone Class E cAMP
receptors Frizzled/Smoothened family Putative
families Ocular albinism proteins
Insect odorant receptors Plant Mlo
receptors Nematode chemoreceptors
Vomeronasal receptors (V1R V3R) Taste
receptors T2R Orphans Putative /
unclassified GPCRs non-GPCR families
Class Z Archaeal/bacterial/fungal opsins
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GPCR Classes
Class A Rhodopsin like o Amine
o Peptide o Hormone protein
o (Rhod)opsin o Olfactory o
Prostanoid o Nucleotide-like
o Cannabinoid o Platelet activating
factor o Gonadotropin-releasing
hormone o Thyrotropin-releasing hormone
Secretagogue o Melatonin o
Viral o Lysosphingolipid LPA (EDG)
o Leukotriene B4 receptor o
Class A Orphan/other Class B Secretin like
o Calcitonin o Corticotropin
releasing factor o Gastric inhibitory
peptide o Glucagon o Growth
hormone-releasing hormone o Parathyroid
hormone o PACAP o Secretin
o Vasoactive intestinal polypeptide
o Diuretic hormone o EMR1
o Latrophilin o Brain-specific
angiogenesis inhibitor (BAI) o
Methuselah-like proteins (MTH) o
Cadherin EGF LAG (CELSR) o Very large
G-protein coupled receptor
Class C Metabotropic glutamate / pheromone
o Metabotropic glutamate o
Calcium-sensing like o Putative
pheromone receptors o GABA-B
o Orphan GPRC5 o Orphan GPCR6
o Bride of sevenless proteins (BOSS) o
Taste receptors (T1R) Class D Fungal
pheromone o Fungal pheromone A-Factor
like (STE2,STE3) o Fungal pheromone B
like (BAR,BBR,RCB,PRA) o Fungal
pheromone M- and P-Factor Class E cAMP
receptors Frizzled/Smoothened family
o frizzled o Smoothened Putative
families Ocular albinism proteins
Insect odorant receptors Plant Mlo
receptors Nematode chemoreceptors
Vomeronasal receptors (V1R V3R) Taste
receptors T2R Orphans Putative /
unclassified GPCRs non-GPCR families
Class Z Archaeal/bacterial/fungal opsins
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GPCR Ligands
Rhodopsin family amine receptors
Acetylcholine (muscarinic) Adrenaline
Dopamine Histamine Serotonin
Octopamine Trace amine Rhodopsin family
peptide receptors Angiotensin Apelin
Bombesin Bradykinin C5a anaphylatoxin
CC Chemokine CXC Chemokine CX3C
Chemokine C Chemokine Cholecystokinin
Endothelin fMet-Leu-Phe Galanin
Ghrelin KiSS1-derived peptide
Melanocortin Motilin Neuromedin U
Neuropeptide FF Neuropeptide S
Neuropeptide Y Neuropeptide W / neuropeptide
B Neurotensin Orexigenic neuropeptide
QRFP Opioid Orexin Oxytocin
Prokineticin Somatostatin Tachykinin
Urotensin II Vasopressin
Protease-activated (thrombin) Adrenomedullin
(G10D) GPR37 / endothelin B like
Chemokine receptor like Melanin-concentrating
hormone Follicle stimulating hormone
Lutropin-choriogonadotropic hormone
Thyrotropin
Rhodopsin family other receptors Rhodopsin
Olfactory Prostaglandin Prostacyclin
Thromboxane Adenosine Purine /
pyrimidine Cannabinoid Platelet
activating factor Gonadotropin-releasing
hormone Thyrotropin-releasing hormone
Melatonin Lysosphingolipid and LPA (EDG)
Leukotriene B4 receptor SREB Mas
proto-oncogene Mas-related (MRGs) RDC1
EBV-induced Relaxin LGR like Free
fatty acid G protein-coupled bile acid
Nicotinic acid GPR GPR45 like
Cysteinyl leukotriene Putative / unclassified
Class A GPCRs Secretin family Calcitonin
Corticotropin releasing factor Gastric
inhibitory peptide Glucagon Growth
hormone-releasing hormone Parathyroid
hormone PACAP Secretin Vasoactive
intestinal polypeptide EMR1 Latrophilin
Brain-specific angiogenesis inhibitor (BAI)
Methuselah-like proteins (MTH) Cadherin EGF
LAG (CELSR) Putative / unclassified Class B
GPCRs
Metabotropic glutamate family Glutamate
(metabotropic) Extracellular calcium-sensing
GABA-B Pheromone (V2R) Taste receptors
(T1R) Orphan GPRC5 Orphan GPCR6 Bride
of sevenless proteins (BOSS) Putative /
unclassified Class C GPCRs Other families
Frizzled / Smoothened family Ocular albinism
proteins Vomeronasal receptors (V1R)
Taste receptors (T2R) Insect odorant
receptors Nematode chemoreceptors Plant
Mlo receptors Fungal pheromone cAMP
(Dictyostelium) Bacterial rhodopsin
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G ProteinCoupled Receptor Mutational Data
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
The rhodopsin crystal structure provides a
structural basis for understanding the function
of this and other G proteincoupled receptors
(GPCRs). The major structural motifs observed
for rhodopsin are expected to carry over to other
GPCRs, and the mechanism of transformation of the
receptor from inactive to active forms is thus
likely conserved. Moreover, the high expression
level of rhodopsin in the retina, its specific
localization in the internal disks of the
photoreceptor structures termed rod outer
segments (ROS), and the lack of other highly
abundant membrane proteins allow rhodopsin to be
examined in the native disk membranes by a number
of methods. The results of these investigations
provide evidence of the propensity of rhodopsin
and, most likely, other GPCRs to dimerize, a
property that may be pertinent to their function.
Annu. Rev. Biochem. 2006. 75743767
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
Annu. Rev. Biochem. 2006. 75743767
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• Rhodopsin highlights
• single photon detection
• one ROS stack contains 1000-2000 discs
• 6.4 million retinal cells (70 rods BW, lt2
  cones color)
• 50 of membrane area is rhodopsin (balance is
  phospholipids and cholesterol)
• molar ratio of rhodopsin to phospholipids is
  about 160

Annu. Rev. Biochem. 2006. 75743767
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Visual system signaling cascade
Lower cGMP leads to Na channel
inactivation, which in turn leads to
hyperpolarization
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Visual system signaling cascade
signaling state, binds GDPG????
GTPG? separates from G??
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
Annu. Rev. Biochem. 2006. 75743767
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
Annu. Rev. Biochem. 2006. 75743767
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11-cis vs. all-trans retinal
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Absorption spectrum of rhodopsin
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
Annu. Rev. Biochem. 2006. 75743767
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Bacteriorhodopsin
Bovine rhodopsin
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Annu. Rev. Biochem. 2006. 75743767
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
Annu. Rev. Biochem. 2006. 75743767
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Annu. Rev. Biochem. 2006. 75743767
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Structure of the ß2-adrenergic GPCR
Originally published in Science Express on 25
October 2007. Paper version Science 23 November
2007 Vol. 318. no. 5854, pp. 1258 -
1265. High-Resolution Crystal Structure of an
Engineered Human ß2-Adrenergic G ProteinCoupled
Receptor Vadim Cherezov, Daniel M. Rosenbaum,
Michael A. Hanson, Søren G. F. Rasmussen, Foon
Sun Thian, Tong Sun Kobilka, Hee-Jung Choi, Peter
Kuhn, William I. Weis, Brian K. Kobilka,Raymond
C. Stevens Heterotrimeric guanine
nucleotidebinding protein (G protein)coupled
receptors constitute the largest family of
eukaryotic signal transduction proteins that
communicate across the membrane. We report the
crystal structure of a human ß2-adrenergic
receptorT4 lysozyme fusion protein bound to the
partial inverse agonist carazolol at 2.4 angstrom
resolution. The structure provides a
high-resolution view of a human G proteincoupled
receptor bound to a diffusible ligand.
Ligand-binding site accessibility is enabled by
the second extracellular loop, which is held out
of the binding cavity by a pair of closely spaced
disulfide bridges and a short helical segment
within the loop. Cholesterol, a necessary
component for crystallization, mediates an
intriguing parallel association of receptor
molecules in the crystal lattice. Although the
location of carazolol in the ß2-adrenergic
receptor is very similar to that of retinal in
rhodopsin, structural differences in the
ligand-binding site and other regions highlight
the challenges in using rhodopsin as a template
model for this large receptor family.
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Structure of the ß2-adrenergic GPCR (tiff)
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Structure of the ß2-adrenergic GPCR (gif)
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Structure of the ß2-adrenergic GPCR (jpg)
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G ProteinCoupled Receptor Rhodopsin Krzysztof
Palczewski
Annu. Rev. Biochem. 2006. 75743767