Receptors 1 Flashcards
Receptor
- Refers to a protein that participates in intracellular communication via chemical signals
- Upon recognition/binding of external chemical signal (ligand)
- Receptor protein transmit signal INSIDE the cell.
- Upon recognition/binding of external chemical signal (ligand)
Receptor Lignads
- Include Endogenous signaling molecules:
- Hormones
- Neurotransmitters
- Also Include Drugs that serve as antagonist or agonist.
- Ex.
- Propranolol antagonizes #> b-adrenergic receptors
- Prazosin antagonizes #> a-adrenergic receptors
- Atropine #> AcTH on cholinergic muscarinic receptors
- Ex.
Signal Transduction
- Process of transferring the information from:
- OUTSIDE the cell –> to inside the cytoplasm
-
First messenger (ligand/drug) concentration levels are translated into a biological response
- = secondary messenger
- Most signaling across membranes occur by one of a limited set of mechanisms
Signal Transduction Mechanisms
5
- Lipid-soluble drug diffuses across the membrane
- –> intracellular receptor (enzyme / regulatory protein)
- Drug binds to transmembrane ion channel
- stimulates internal enzyme action
- 2 forms - simply converting or phosphorolating
- Drug binds to transmembrane ion channel
- regulates its opening (can be in or out)
- Drug binds to cell-surface receptor
- –> interacts with G-protein
- –> regulates an internal enzyme activity
- –> interacts with G-protein
Ligand Gated Ion Channels
LGIC
- Binding –> ions enter
- change in membrane potential
- ionic concentration change
-
VERY FAST
-
if the effect of the ligand is conformational change of the receptor
- GABAAR / AChR / Nicotinic acetylcholine
-
if the effect of the ligand is conformational change of the receptor
Tyrosine Kinase Linked Recptors
TRKs
- binding –> protein phosphorolation
- ex. Insulin receptor
- __response is slower
- since a reaction is required (phosphorolation)
G-Protein Coupled Receptors
GPCRs
- binding of cell-surface receptor–> g-protein –>
- intracellular secondary messenger (cAMP)
- –> protein posphorlation (does not always occur)
- ex. B-adrenergic receptor
- intracellular secondary messenger (cAMP)
- __slower due to reaction required (phosphoralation)
Ligand Activated Transcription Factors
eg. SERMs
- ligand –> intracellular receptor
- –> translation / transcription
- –> protein –> effect
- takes hours, takes the longest time for effects
Nicotinic AcetylCholine Receptor
Ligand Gated Ion Channel (LGIC - Na/K)
- ACh Binding –> rotation of transmembrane helices
- (M2 amphipathic helixes surround the channel)
-
–> OPENING of inner pore
- allows sodium and pot. to go through
- Ex. Varenicline (chantix)
- partial AGONIST of 1 subtype of the receptor
Varenicline
Chantix
Partial Agonist
Nicotinic Acetylcholine Receptor
- –> conformational change –> allows na/k to enter
Volted Gated Ion Channels
VGIC
- Comprised of a voltage sensor / pore / gate
-
Ion Specific
- Na / K / Ca / Cl
- Activated by changes in electrical membrane potential
-
Movement of voltage sensor
- –> conformational change (open/close)
-
Movement of voltage sensor
- Critical role in excitable cells:
- Neuronal / Muscle cells
-
Ex.
- Verapamil (Ca channel blocker)
- Amiodarone (arrhythmics)
- Carbamezapine (Na)
- Penytoin (Cl)
- Lidocaine (anesthetics NA)
Transient Voltage (TRP-type) Ion Channels
ex. TRPV
- 6 major types that differ in ligand specificity & bio fxn
- Tetramers w/ 6 transmembrane helices in each
- Most important for drugs:
- TRPV (vanniloid)
- TRPA (anykrin)
- TRPM (melastatin)
- also involved in INSECT VISION downstream from rhodopsin
Transient Voltage Ion Channels
TRPV1
-
Vanilloid receptor
-
Thermal / Noxious agent sensors
- <activated by pungent chemicals:>
<li>
<strong>capsaisin</strong> / <strong>Resinferatoxin</strong>
- <activated by pungent chemicals:>
-
Thermal / Noxious agent sensors
</li>
<li>
<strong>DkTx (</strong>spider venom toxin) / vanillotoxin</li>
</activated>
* Analgesic drug design
* DUAL GATE MECHANISM
* 2 diff ligands bind in different region of receptor
* –> conformational changes induced by each ligand
* are allosterically coupled
Transient Voltage Ion Channels
TRPA
A = ankyrin
- Ankyrin mechanoreceptor
-
activated by pungent chemicals:
- wasabi: allyl isothiocyanate
-
activated by pungent chemicals:
Transient Voltage Ion Channels
TRPM
m = melastatin
- Melastatin receptor
- diverse fxn amoung 8 species:
-
cold sensors
- < pepperment / menthol / calcium
-
Overexpression –> tumorigenesis
- = prostate cancer / melanoma
GPCRs
- Lefkovits / Kobilka Nobel Prize
- Large family of signaling proteins
- each specifically binding a unique small-molecule ligand
- Transduce chemical “signal” from OUTSIDE
- –> internal changes (biological response)
-
Integral to plasma membrane
- a-helixes transverse membrane
- characteristic 7-helix bundle structure (7-pass)
- extracellular domain = bind agonist
- intracellular domain = regulate cytosolic enzymes__
GPCR
Dopamine D3 Receptor
-
7transmembrane helices (a) + extracellular loops
- = form binding pocket for dopamine
- ex. ETICLOPRIDE (D-receptor ANTAgonist)
-
ICL2 (intracellular helix)
- –> conformational change after binding
- assumes alpha-helical structure w/ 2-3 helical turns
- communicates w/ G-proteins in the cell
- –> activate enzyme (effector molecule)
- –> conformational change after binding
- Takes a longer response time compared to LGIC’s
Receptor Tyrosine Kinases
Insulin Receptor
IR
- Agonist (insulin) –> IR –> Autophosphorylation
- receptor aquires kinase activity
- Phosphorolated receptor RECRUITS substrate (IRS1)
-
IRS1 is phosphorolated
- –> binds to effector molecules = PI-3-kinase
-
PIP3
- –> transduce information to nucleus
-
IRS1 is phosphorolated
- longer response time needed to complete
Ligand-Activated Transcription Factors
ex. SERMs
- Ligand (estrogen) –> ERbeta
- –> binding to Major groove of DNA promoter region
- DNA transcribed –> mRNA
- translated –> variety of protein products
- DNA transcribed –> mRNA
- –> binding to Major groove of DNA promoter region
- ex. myc / VEGF / Bcl2 / IGFR1 / IRS1/ TGFa / CD1
Allosteric Activator
Makes agonist effective at LOWER concentrations
Binding to allosteric site changes the affinity of the agonist site
- less agonist is needed to exhibit response
- bind to different (allo) sites
Allosteric Inhibitor
Makes agonist effective at HIGHER concentrations
Binding to allosteric site changes the affinity of the agonist site
- MORE agonist is needed to exhibit response
- bind to different (allo) sites
Lock & Key Model
Linus Pauling
- Ligand = key
- binds to specific receptor = Lock
- –> unlocks cell response
- binds to specific receptor = Lock
- First proposed by Emil Fishsher / JBS Haldane
Agonist
- Drug = Agonist
- Many drugs work by MIMICKING a natural ligand:
- drug causes receptor to respond in the same way as the natural substance
Antagonist
- Drug = Antagonist = Competitive inhibitor for enzymes
- Bind to receptor @ Orthosteric binding site
- and DO NOT produce a response
-
Prevent receptor from binding the natural ligand
- # > inhibit function
- this also counts for causing structure DEFORMATION that inhibits fxn
- Bind to receptor @ Orthosteric binding site
Dose-Response Curve
Fraction of Receptors Bound (B) Vs Drug Concentration (D)
- Pysiological responses are NOT simple fxn of the amount of dose given
- May be INITIALLY LINEAR
- but level off @ higher doses
- May be INITIALLY LINEAR
- Bmax = 100% of receptors bound
- B IS NOT the same as effect or efficacy (vs EC50)
KD
Drug-Receptor Dissociation Constant
- When drug cocentration (D) shows HALF SATURATION
-
[D] when [B = 50%]
- half of receptors are occupied
-
[D] when [B = 50%]
-
Smaller the KD = Greater the affinity
- less drug is needed to saturate the receptor
- KD is related to the free energy of ligand-receptor interactions
- not the same as EC50**
- inverse of Ka
Ka
Association Constant = AFFINITY
inverse of dissociation constant KD
Ka = k1 / k-1
- Greater the Ka = greater the affinity
Semi-Logarithmic
Receptor Dose-Response Curves
- Scale @ low concentration (where binding changes rapidly)
- compresses [D] @ HIGH concentration (where binding changes slowly
- Does not change value of Bmax & KD
Advantages of Semi-Log
Dose response curve
- Better defined Plateau
- Plotting of wide range of [D]
- Enables comparison of drugs w/ different affinites due to compression of x axis
Graded Dose Response Curve
Emax
ED50
Drug Efficacy [E] vs Drug Concentration [D]
-
Emax = maximum response (effect) achieved by agonist
- = drug efficacy
-
ED50= druc conc. (dose) at which50% of Emax is achieved
- = drug potency
- different units of E vs B
Drug Response Curves of Agonist
- Agonist w/ High Affinity
-
shit curve to the LEFT
- binding & drug action start @ LOWER concentrations of agonist
-
shit curve to the LEFT
- Agonist w/ LOW AFFINITY
-
causes curve to move to the RIGHT
- MORE agonist is needed to make the drug-receptor complex
-
causes curve to move to the RIGHT
Quantal Dose-Response Curve
Individuals Responsing (%) Vs Drug Dose [D]
- Describe POPULATION rather than single individual responses
- based on plotting comulative frequency of distribution of responsers
- vs log of drug dose
- based on plotting comulative frequency of distribution of responsers
-
Emax & ED50
- like GRADED dose response curve
Graded Response
INFINITE Number of intermediate stages
- Blood vessel dialation
- Bood Pressure Change
- Heart Rate Change
Quantal Response
BINARY all-or-none
- Death
- Pregnancy
- Cure
- Pain relief
- Effect of a given magnitude
Neutral Agonist
Have an efficacy of 0
Similar to Antagonist
bind to receptor without exerting an intracellular effect
Types of Antagonism:
Pharmacological
-
blockage of the action of a drug-recepter interaction
- by another compound
- ex. cimetidine –> blocks interaction of:
- histamine –> H2 Receptors
- –> lower gastric acid secretion
- histamine –> H2 Receptors
Types of Antagonism:
Physico-chemical
- Interaction of two drugs in solution
- such that the effect of the active drug is lost
- Ex. metal chelators + toxic metals
Types of Antagonism:
Biochemical
Agonist is DESTROYED by biochemical reaction
- Ex. Sodium bicarb neutralizes Gastric acid secretion
- Ex. Protamine antagonizes heparin
- __protamine = alkaline w/ +e charge
- heparin = acid w -e charge
Types of Antagonism:
Physiological
- Interaction of two drugs with opposing physiological actions
-
Ex. Histamine vs Epinephrine
- __Histamine LOWERS arterial pressure thru VASODILATION (h1 receptor)
- Epinephrine RAISES arterial pressure thru VASOCONSTRICTION
- B-adrenergic receptors
- Ex. Insulin antagonizes gluticocorticoids
- __hyperglycemic effects counteracted
Pharmacological Antagonist:
Competative (surmountable) Antagonist
- prevent biological actions of agonist
- Bind to SAME SITE on receptor as agonist
- inhibition CAN be overcome by increasing agonist concentration
- = reversible
-
primarily affect agonist POTENCY
- DO NOT alter receptor function
- May also bind to another site on receptor that BLOCKS the action of an agonist
- SHIFT RIGHT
Pharmacological Antagonist:
Non-Competative (insurmountable) Antagonist
- Bind Covalently to SAME SITE as agonist (IRREVERSIBLE)
- or to a site distinct from that agonist (irreversible or reversible)
- Inhibition CAN NOT be overcome by increasing agonist concentration
- primarily affect EFFICACY
- SHIFT DOWN
Naloxone
u-opioid receptor ligand
COMPETATIVELY ANTAGONIZES
action of morphine / heroin
opiate overdose treament
Perampanel
Fycompa, 2012
NONcompetative antagonist of AMPA
ionotropic glutamate receptor
Treatment of epilepsy (antiepileptic)
Effect on Dose-Response Curves
Competative Antagonist
Agonist + Competative Agonist (B)
Agonist has REDUCED POTENCY
but still MAXIMUM EFFICACY
- Produce a PARALLEL RIGHT-SHIFT
-
Magnitide of the shift is dependent on:
- [B] , concentration of antagonist
- KB , potency of antagonist
Extent of Antagonism
- Depends on the plasma concentration of the Antagonist:
- dose of the antagonist
- rate of clearance of the antagonist
-
concentration of the agonist
- –> can OVERCOME competative antagonist
-
prescription must take into account possible changes in the endogenous agonist concentration
- ex. norepinephrine vs Propranolol
Effect on Dose Response Curves:
Non-Competitive Antagonist
aka Allosteric / Allotopic
Agonist + Non-Competitive Antagonist
Agonist has Maximum Potency
but REDUCED EFFICACY
-
curve moves DOWN (depressed response)
- no horizontal shift
Glutamate
Allosteric Antagonist (modulator) Effect
modulator REDUCES mGlu1R signaling
DECREASES effect (shift down)
without effecting binding or potency (no lateral shift)
GABA
Allosteric Antagonist (modulator) Effect
INCREASES Gaba POTENCY (<shift left<)
&
Maximal Effect (^shift up^)
CP55940
full cannainoid receptor agonist
Allosteric Antagonist (modulator) Effect
DECREASES EFFICACY (moves down)
Increases Potency (moves left)
Positive Allosteric Agonist of
GABA-AR
More Agonist Sites can be occupied
INCREASE maximal GABA response
- Examples:
- Etomidate / Neurosteroids
- Barbiturates / Benzodiazapenes
- Alcohols /Isoflurane / Propofol
Propofol on GABA
Positive Allosteric Agent (propofol) on Gaba-ar
-
Receptor binds GABA more TIGHTLY
- dose response curve shifts to the LEFT
-
@ particular GABA concentration, more agonist sites are occupied in the presence of propofol
- response increases – shift ^^UP^^
Occupancy Model
=
Two-State Model
Agonist occupies receptor –> Trigers Response
Strength of response : directly related to fraction of receptors occupied
does not explain all observations
Two-Way Model
Occupency Model
-
DR*- Agonist binding –> conformational change
- R->R* active
-
R* - Conformational change occurs SPONTANEOUSLY
- in absense of agonist
- –> is active
- small fraction of R* is in EQ without athe agonist
- agonist drives the conformational change, stabilizes R*
- in absense of agonist
Negatives/Shortfalls of
Occupency Model / Two-way model
- Occupency model is INSUFFICIENT to explain these facts:
- Experimentally shown:
-
Maximal response can be found when only a fraction of total receptor population is occupied
- spare receptors are present
- Diff. Drugs have different capacities to initiate response when bound
- = different efficacies
- Certain receptors are active even in absence of agonist
- Certain drugs act as “inverse” agonist
- shut down intrinsically active receptors
-
Maximal response can be found when only a fraction of total receptor population is occupied
Relationship between
Occupancy & Physiological effect
- assume that response (E) is :proportional: to receptor bound drug, where Emax = maximum effect
-
Magnitude of effect (E) is dependent on:
- Drug Concentration [D]
- Affinity KD (receptor occupancy)
-
Ability to activate the receptor “e”
- aka intrinsic activity
Potency Vs Efficacy
Curve
Potency INCREASE as response shift to the LEFT >>>
Efficacy INCREASES as maximal effects of receptor shift ^^UP^^
Efficacy
Maximal Response produced by a drug
analogous to Maximal Velocity for Enzyme
- Depends on 3 factors:
- Intrinsic Activity (e)
- Number of drug receptor complexes formed
- Efficiency of signal transduction in producing physiological response
- Possible to have High Affinity w/ low efficacy
- or vice verse
A?
FULL Agonist
Max Potency = lowest drug conc to reach max effect
Max Efficacy = reaches max effect
B?
Partial Agonist
Maximum Potency = reaches it max potential
Reduced Efficacy = Is not as Effective as a full agonist
C?
FULL Agonist
Reduced Potency = requires more drug to reach max efficacy
Maximum Efficacy = Reaches its full effect
D?
Partial Agonist
- Reduced Potency = requires more drug to be effective*
- Reduced Efficacy = Is not as Effective as a full agonist*
Partial Agonist
Agonist with efficacy <1 (e<1)
can serve as competitive agonist (also lower efficacy)
Shifts RIGHT! but can also shift down
- May completely occupy receptor sites
-
BUT the response reaches a plateau at a value less than the maximal 100%
- reduced effect SHIFT DOWN
-
BUT the response reaches a plateau at a value less than the maximal 100%
-
Agonist binding may favor new conformation of receptor
- –> but has LESS intrinsic activity (lower efficacy)
-
= poor contact w/ “downstream” partners in signal path
- or more receptor states may need to be considered
Partial Agonist Vs Antagonist
-
Heroin = MOST efficacious agonist of opiod receptor
- FULL AGONIST
- Codeine** = most potent **partial agonist of opiod receptor
-
Nalaxone = antagonist
-
**effect of antagonist needs agonist to be present to be observed
- otherwise silent
-
**effect of antagonist needs agonist to be present to be observed
Inverse Agonist
Has Independent impact upon receptor activity
produces Opposite effect to agonist
-
Unlike antagonist, they are active in the absence of agonist
- intrinsically / constitutively active
-
Ex.
- cimetidine –> H2 receptors
- Haloperidol –> D2 receptors
Quantitative Theory
Cheng-Prusoff Equation
- Applicable to:
- Simple Occupency model (esp. for enzyme INHIBITION)
- Binding to Noncatalytic proteins (receptors)
- Only consider competitive antagonism
-
KL = dissociation constant of AGONIST (ligand) used in displacement assay
- obtained from literature / determined beforehand
- {1 + [L]/KL} = Dose Ratio
KD & EC50
EC50 is ALWAYS HIGHER than KD
-
KD = thermodynamic paramater
- interaction between receptor / antagonist
- (absolute value / constant, like melting point)
-
EC50 = functional strength of a ligand
-
depends on experimental conditions
- ex. conc of L / presence of spare receptors
-
depends on experimental conditions
- EC50 would INCREASE if L was at a higher conc.
KD = EC50 when?
if NO LIGAND IS PRESENT
when, [L] = 0
ED50 Vs EC50
ED50 = DOSE for 50% of population to obtain therapeutic effect
Vs
EC50 = DOSE that reduces response to 50% of maximal value (invidual)
Drug Potency
- Expressed as the Dose or Concentration
- that produces 50% of maximal response ED50 or EC50
- under experimental conditions
- that produces 50% of maximal response ED50 or EC50
-
LOWER EC50 = GREATER Potency
- less drug needed to produce same effect
What effects EC50 ?
- Affinity of drug for receptor KD
-
But also:
- metabolism
- distribution
- Presence of other ligands
KL
Dissociaition constant of AGONIST
used in displacement assay
Obtained from literature or determined beforehand
-
KD = dissociation constant of the receptor-antagonist complex
- = affinity
Fraction of Occupied Receptor
f
Receptor occupency “protein saturation”
Molar Fraction of occupied receptor
- When [L] = KD, 50% protein is bound
- When [L] = 0.1 KD only 9% of the protein is bound
- When [L] = 10 KD, the protein is 91% saturated
