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Binding of Drugs in/to Cells
• Receptor = Drug “target”– Membrane protein– Enzyme– Nucleic acid
• Most drugs bind receptors by weak, noncovalent forces (what are these?)– May be reversed by pH change
Molecular Recognition Specificity
• Cellular specificity– Not all receptors in all cells, tissues
• Receptors selectively bind partic ligands– Stereoselectivity
• No drug completely specific
Ligand/Receptor Interactions
• Reversible, bimolecular reaction– D + R DR DR* Response– Where R*=Receptor w/ conform’n change– Each will have rate constant
• What does this remind you of??
Activating Drugs = Agonists
• Drug/receptor binding conform’l change in receptor act’n “downstream” cell biochem
pathway(s) tissue response
• May bind at separate site on receptor– “Allosteric modulators”– Increases response to natural agonist
Some definitions
• Affinity = tendency to bind receptor– Specificity– Association/dissociation constant
• Efficacy = tendency to activate receptor– Full agonists elicit max response– Partial agonists elicit submaximal
response
Antagonists Bind Receptors…
• BUT no activation occurs– No conform’l change in receptor, so no
pathway response• May keep agonists from binding
– Competitive– Book ex: curare blocks ACh from
receptors of neuromuscular junction inhib’n muscle depolarization paralysis
• Allosteric modulators may decrease natural agonist binding
• Best antagonists have efficacy=0
Receptor Superfamilies
• Ligand-gated ion channels• G protein-coupled receptors• Receptor tyrosine kinases• Nuclear hormone receptors
Ligand-Gated Ion Channels
• Brain, periph NS, excitable tissues (heart), neuromuscular junction– Nicotinic cholinergic receptors
(neuromusc)– GABA receptors (brain)– Glutamate receptors (brain)
change membr potential fast synaptic transmission
• Complex prot’s w/ multiple subunits
Book Ex: Nicotinic Receptor
• Number of subunits differs w/ tissue– Antagonists differ– Allows selective blockade
neuromuscular junction• Multiple binding sites for Ach
– Excitatory incr’d Na+/K+ permeability incr’d
depol’n incr’d probability of action potential
• Direct transduction (no biochem intermediates)
• Allosteric modulators may increase/decrease transmitter response in ligand-gated channels
• Ex: benzodiazepines– Antianxiety; sleep disorders– Bind GABA ligand-gated receptors
• GABA inhibitory
– Increases ability of GABA to open channels
G Protein Coupled Receptors
• Single subunit• 7 helices span bilayer• Agonists may bind extracell N-
terminal domain, or between helices– Few allosteric modulators known
• Cytoplasmic loop couples to G protein
G Proteins
• Intermediary mol’s• Bind guanine
nucleotides• Extrinsic (periph)
prot’s at inside bilayer– Anchored to
membr by fa chain– Shuttle between
receptor, target prot’s
• 3 subunits– GTPase activity by
• “Resting state” G prot – trimer w/ GDP occupying site on subunit
• Agonist binding receptor conform’l change w/in cytoplasmic domain
Receptor acquires high affinity for G prot binding G prot to receptor
• GTP replaces GDP duplex dissoc’s from -GTP
– Diffuse along membr– Assoc w/ enzymes, ion channels act’n
or deact’n
• Term’n activity w/ hydrol Pi from GTP w/ GTPase activity of subunit– Trimer reunites
• Single agonist binding can activate sev G-prot mol’s for sev prod’s/act’n results Amplification
• Sev types G prot’s– Interact w/ diff receptors– Control diff effectors
• Gs stim’s enz adenylate cyclase, PLC, others
• Gi inhibits ad cyclase, PLC, others
– Agonist specificity
Cellular Responses
• Amplification of signal through second messengers that activate kinases– cAMP– Phosphatidylinositol
Control regulatory enz’s through covalent mod’n
Large, varied cell responses• GPCRs also control
– PLA eicosanoid release– Ion channels depol’n, transmitter
release, contractility, etc.
Examples of GPCRs
• Receptors for – ACh (muscarinic)– Neuropeptides– Ephinephrine
• Muscle (3 types), liver, fat, epithelium, neurons
Receptor Tyrosine Kinases
• Single transmembr helix• Large extracell domain
– Agonist binding site• Large intracell domain
– Some incorporate tyr kinase activity– Cytokine receptors assoc w/ cytosolic
kinases• Agonist binding act’n
dimerization– Monomeric form inactive
• Dimerized receptors autophosphorylate tyr residues
• Phosphorylated tyr attracts, binds SH-2 domain protein– Src Homology– Conserved seq recognizes
phosphotyrosine on receptor– Various SH2-domain prot’s allow
selectivity for spec receptors– Some are enzymes
• Kinases• Phospholipases
• Some SH-2 Domain prot’s are couplers for other cell prot’s w/ phosphorylated receptors– Phosphorylation cascades– Impt to cell division, diff’n– Ex: Ras/Raf/MAP kinase pathway
• Impt to cancers
• SH-2 Domain prot’s as couplers – cont’d– Ex: Jak-Stat Pathway
• Impt for cytokines, growth hormone, interferons
• Cytosolic kinase phosphorylates receptor dimer
– Various Jak’s specificity
• SH-2 domain prot’s (Stat’s) attracted, phosph’d, dimerize
Nucleus gene expression
Nuclear Hormone Receptors
• Intracellular• Most in nucleus
– Some cytoplasmic• Three domains:
– Agonist binding domain at C-terminal
– Transcriptional control domain– DNA binding domain
• Highly conserved• “Zinc fingers”
• Ligands lipophilic– Traverse lipid bilayer– Examples:
• Steroid hormones• Thyroid hormones• Vitamin D• Retinoic acid
– Impt to embryo dev’t
• Agonist binding to receptor conform’l change
Dimerization of receptors• Dimers recognize specific base
seq’s on DNA near genes– Hormone responsive elements– ~200 bp upstream from genes
• Binding DNA may activate or repress gene transcr’n– So “ligand-activated transcr’n
factors”
Other Targets of Drugs• Ion Channels
– Ligands bind voltage (as well as ligand-gated) channels• Block channel• Affect gating
– Activation GPCRs phosph’n channel prot’s• Affect channel opening• Ex: opioids, -adrenoreceptor agonists
– Modulation intracell Ca+2, GTP, ATP• Channels may bind these mol’s• Ex: sulfonylureas act at ATP-gated K+
channels on pancreatic B-cells
• Enzymes– Drug may be substrate analog
• Competitive or irreversible inhibitor
– False substrate• Appears as substrate, so taken up• Not useful as product
– Ex: 5-FU blocks DNA synth
– Prodrugs• Metabolism active agent
• Carrier molecules– Impt for transport across cell membr’s– Have recognition sites for natural mol– Examples:
• Cocaine, antidepressants inhibit noradrenaline uptake
– Amphetamine acts as false substrate
• Loop diuretics affect Na+/K+/Cl- transporter in renal tubule
• Cardiac glycosides inhibit Na+/K+ pump
Single Agonist May Have Complex Effects
• Families of receptors for agonists– Ex: ACh receptors muscarinic, nicotinic
• Further subtypes
• Some receptors very specific• Some receptors bind similar ligands
– Book ex: dopamine structurally sim to norepi, can stim 1-adrenergic receptors
• Multiple receptor subtypes for one ligand can coexist in single cell
Regulation of Receptors
• Drugs, agonists decrease sensitivity of receptors to ligands– Fast: desensitization, tachyphylaxis– Gradual: tolerance, refractoriness,
drug resistance– Usually w/ continuous exposure
• Sensitivity can be increased• Sensitization, desensitization can
occur by ligand to same ligand or another
• May be due to– Change in receptors
• Phosphorylation – inhibits ability to interact w/ G proteins
• Slow conform’l change
– Exhaustion of mediators• Ex: amphetamines relase amines from
nerve terminals; when endogenous amines depleted, drug doesn’t work
• Loss of receptors– Binding agonist to receptor rapid
migration complex to coated pits• Membr invaginations surrounded by
clathrin
– Form intracell vesicles– Receptor dissociates
• Recycled to cell membr
– Agonist degraded in lysosomes• OR May be released outside cell
• Physiological adaptation– Receptor number not static– Hormones may incr, decr receptor
number• Altered rate receptor synth• Slow• Upreg’n supersensitivity
– W/ antagonist– W/ inhib’n transmitter synth/release
• Downreg’n loss sensitivity– W/ prolonged exposure to agonists
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