membranes pt. 2
DESCRIPTION
Unit 4 pt. 2TRANSCRIPT
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Proteins & Signaling
Membranes ~ Part II
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Maintaining Homeostasis• Cells must communicate with their external
environment• Monitoring external conditions determines
cellular responses• Example – E. coli:
– If the bacteria detects a high concentration of lactose, it synthesizes proteins to import and metabolize lactose
– If it detects a higher concentration of glucose, it it synthesizes proteins to import and metabolize gluose
• Membrane proteins help gather information about the environment
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Cellular Communication• In multi-cellular organisms,
communication is more complex• Each cell communicates with
dozens of other cells• Determines:
– When it should grow– When it should differentiate or die– When it should release protein
products needed by other cells
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Mechanisms of Cell Communication
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Communication Through Contact
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Two Types of Membrane Proteins
• The membrane is a barrier • Prevents interchange of materials
– Special channels are needed to transport some materials into & out of the cell
• It also prevents free exchange of information– Special receptors are needed to gather
information• Therefore the cell membrane has 2 major
types of proteins:– Transporters – Receptors
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Intrinsic & Extrinsic Proteins
• Intrinsic membrane proteins– Embedded in the lipid bilayer– Some extend through it– Transmembrane proteins
• Extrinsic membrane proteins– Absorbed to the surface of the lipid
bilayer– Can be separated from the lipid bilayer
without destroying the membrane
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Transmembrane Proteins
• Intrinsic proteins that extend from one side of the membrane to the other are transmembrane proteins.
• Cells pump ions in and out through their plasma membranes.
• More than half the energy that we consume is used by cells to drive the protein pumps in the brain that transport ions across plasma membranes of nerve cells.
• How can ions be transported across membranes that are effectively impermeable to them?
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Ligand Gated Ion Channel
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Domains• Many transmembrane proteins have three
different domains:• A hydrophilic domain at the N-terminus
– consists of hydrophilic amino acids – pokes out in the extracellular medium
• A hydrophobic domain in the middle of the amino acid chain– often only 20-30 amino acids long– threaded through the plasma membrane– made of amino acids having hydrophobic side
chains
• A hydrophilic domain at the C-terminus – protrudes into the cytoplasm.
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Glycoproteins• Many transmembrane proteins are glycoproteins • Sugar side chains are covalently attached to the
hydrophilic domains that protrude into the extracellular membrane.
• A typical mammalian cell may have several hundred distinct types of glycoproteins studding its plasma membrane.
• Each glycoprotein has its extracellular domain glycosylated with a complex branching bush of sugar residues covalently linked to the asparagine side chains.– Some glycoproteins may have 2 or 3 asparagine-
linked sugar side chains, others may have dozens.
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Multi-membrane Spanning Proteins
• Some transmembrane proteins have multiple transmembrane domains.
• Hydrophilic domains alternate with hydrophobic domains.
• The protein chain weaves back and forth between opposite sides of the plasma membrane.
• Called serpentine membrane proteins b/c they are snake-like– A common structure in many serpentine
transmembrane proteins involves 7 hydrophobic domains inserted into the plasma membrane, separated by hydrophilic regions that are looped out alternatively into either the cytoplasm or the extracellular space = 7 membrane spanning proteins
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Receptors• Specialized transmembrane proteins that
acquire information from the extracellular space
• Relay this information into the cell through the plasma membrane
• Cell surface receptors act as the antennae of the cell.
• Mammalian cells have wide variety of transmembrane receptors
• Two important types: – Growth Factor Receptors – G Protein Receptors
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Growth Factor Receptors• Help the cell determine whether or not it
should grow by binding growth factors• Growth factors may be present in the
medium around the cell – Sometimes called mitogens because they
induce the cell to grow and pass through mitosis
– They are polypeptides, often 50-100 amino acids long.
• When present in sufficient quantity, a growth factor (GF) will stimulate a cell to enter into a round of growth and division.
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Specificity of Binding
• GFs bind to cell surface GF receptors. • Each type of GF binds to the extracellular
domain of its own specific receptor – will not bind to receptors for other growth
factors.
• Each type of receptor binds specifically to its own ligand– accommodates the appropriate growth factor
in a lock-and-key fashion
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Variety of Ligand: Receptor Pairs
• Other ligands besides growth factors convey signals from cell to cell through intercellular space.
• There are at least several hundred distinct receptor: ligand pairs in our body
• Each devoted to the binding of a distinct extracellular ligand such as a growth factor to its cognate receptor.
• Each ligand originates elsewhere and is secreted by a cell or cells specialized for its release.
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Transmembrane Signal Transduction
• The binding of a ligand to its receptor is the beginning of the signalling process.
• How does the interior of the cell learn that the ligand has bound?
• How is this translated into information the cell can use?
• Transmission of information by a protein is a form of signal transduction.
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An Overview of Cell Signaling
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Structure GF Receptor Proteins
• Outside the cell, they have a ligand-binding N- terminal ectodomain
• Inside is a single membrane-spanning transmembrane domain.
• At their C-termini in the cytoplasm, they have a specialized enzyme domain– This becomes activated whenever the
extracellular domain of the receptor binds a GF ligand.
– In the case of many GF receptors, the cytoplasmic enzyme domain contains protein kinase activity.
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Kinases & Signal Transduction
• Kinases are enzymes that attach phosphate groups to their substrates.
• Protein kinases take the gamma-phosphates from ATP and transfer them to protein substrates, resulting in the phosphorylation of the substrate proteins.
• The phosphate groups are attached to the tyrosine side chains of substrate proteins that communicate with or lie near the cytoplasmic domains of the GF receptors.
• These receptors are considered to have protein tyrosine kinase activity to distinguish them from many other protein kinases that are devoted to other signalling functions.
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Sequence of GF Signal Transduction
• The GF ligand binds to the extracellular domain of its receptor.
• This activates the tyrosine kinase domain at the other end of the receptor in the cytoplasm.
• The tyrosine kinase becomes active and phosphorylates a series of cytoplasmic substrate proteins.
• These are activated or altered functionally as a consequence of being phosphorylated.
• They then send signals further into the cell that result in the cell growing and dividing.
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An External Event
• GF ligand does not need to enter the cell in order for transmembrane signalling to occur.
• All active transmembrane signal transduction occurs while the ligand is still in the extracellular space.
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Mechanism of Kinase Activation
• How does the association of GF ligand outside the cell cause tyrosine kinase activation inside the cell?
• Some considerations:– There are many copies of each type of GF
receptor molecule that are displayed on the surface of a given cell.
– These receptor molecules, while tethered in the plasma membrane via their hydrophobic transmembrane domains, diffuse laterally through the plane of the plasma membrane.
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Dimerization
• When a GF ligand binds to a single receptor molecule, it encourages the dimerization of the receptor with another receptor molecule floating in the plasma membrane.
• Often the GF ligand itself has two receptor-binding ends– enables it to serve as a bridge between the
two receptors – attracts two receptors– encourages their dimerization– stabilizes the resulting receptor/ dimer pair.
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Passing the Message
• Dimerization pulls the cytoplasmic domains of the two receptor molecules closer.
• The tyrosine kinase (TK) of one receptor molecule then phosphorylates the kinase domain of the second receptor molecule
• This phosphorylation results in a steric shift in the 3-dimensional structure of the phosphorylated kinase domain
• This causes its functional activation.
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Tyrosine Kinase Receptor Dimers
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The Final Steps
• The two kinase domains phosphorylate and thereby activate each other.
• Once they are activated, they phosphorylate nearby cytoplasmic substrate proteins that then pass signals further into the cell.
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Phosphorylation Cascade
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7 Membrane-spanning Serpentine Receptors
Varied Functions• Receptors on cells of the tongue convey taste. • Hundreds of receptor types in our nose convey
information about odors.• A carotenoid molecule related to vitamin A binds
rhodopsin in the rods and cones of our eyes. – It picks up photons which alters its conformation, and
causes the receptor to which it is bound to release signals into the rod/cone cytoplasm that result in our perception of light.
• Baker's yeast cells communicate their sexual identity to each other by release of polypeptide mating factors that bind this type of receptor
• Epinephrine controls the “flight or fight response”
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Exchange of Yeast Mating Factors
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A G-Protein Receptor
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The Role of Epinephrine• Also known as adrenaline• Released by the adrenal glands above the
kidneys in response to stressful stimuli. • Epinephrine travels through the blood stream and
binds to specific receptors on cells in various tissues throughout the body.
• This results in the mammalian fight / flight reaction.
• This includes:– increased heart rate, – decreased blood flow to gut– increased blood flow to skeletal muscles– increased blood glucose
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Tracing One Action• Epinephrine acts at many sites to
produce a wide array of physiologic changes
• One of these is increased blood glucose
• Epinephrine causes liver and muscle cells to break down glycogen and release the resulting glucose into the circulation
• We will trace this one action of epinephrine
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How Epinephrine Acts
• Epinephrine binds to its receptor on the surface of a variety of cell types throughout the body.
• This beta adrenergic receptor is a 7 membrane-spanning, serpentine receptor embedded in the plasma membranes of these cells.
• As is the case with the growth factor receptors, the epinephrine ligand is not internalized into the cell.
• While bound for a short period of time to its receptor, epinephrine causes the latter to release biochemical signals into the cell cytoplasm.
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The Epinephrine Receptor
• These receptors do not depend upon receptor dimerization to transduce signals across the plasma membrane.
• Instead, single receptor molecules will change their 3 dimensional steric configuration in response to ligand binding.
• This steric shift affects the configuration of the cytoplasmic domains of the receptor (the loops of receptor protein that protrude into the cytoplasm).
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Cytoplasmic Signal Transduction
• The receptor communicates with the cytoplasm by stimulating a second protein
• This is known as a G protein (G = guanine)• The G protein normally lies near the
receptor in an inactive, quiet state. • When the receptor is activated by ligand
binding, it pokes the G protein. • The G protein responds by switching itself
on, into an active state. • Once in the active state, the G protein
sends signals further into the cell.
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The G Protein is Binary• The G protein remains in the active
state for only a brief period, after which it shuts itself off.
• The G protein's two states (ON or OFF) are determined by guanine nucleotide which it binds– thus the term G protein
• When it is inactive, it binds GDP• When active, it binds GTP.
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GTP Binding Activates the Protein
• The resting, OFF form of the G protein sits around with its bound GDP.
• When a ligand-activated receptor pokes it, the G protein releases its bound GDP
• It then allows a GTP molecule to jump aboard.
• The GTP-bound form of the G protein is the active ON state.
• While in the ON state, it releases downstream signals.
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Feedback Regulation• After a short period of time
(seconds or less), the G protein hydrolyzes its own GTP back to GDP . . .
• Thus shutting itself off. • This hydrolysis represents a
negative feedback mechanism • Ensures that the G protein is only in
the active, signal- emitting ON mode for a short period of time.
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Structure of the G Protein• Composed of 3 subunits: alpha, beta &
gamma• In its inactive OFF state, the 3 subunits
are bound together• The alpha subunit binds the guanine
nucleotide, in this case GDP. • When the beta adrenergic receptor
activates the G protein, the alpha subunit releases GDP,
• then binds GTP, • and falls away from the beta and
gamma subunits.
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The Signaling Cascade
• Once GTP is bound, the GTP-bound alpha subunit also loses affinity for the receptor.
• It dissociates from receptor, • moves over and pokes another nearby
protein• the enzyme adenylate cyclase, • which is activated by being poked, • and cyclizes ATP into 3'5' cyclic AMP.
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The Second Messenger
• cAMP is a second messenger• After G protein encounters adenyl cyclase enzyme,
the alpha subunit of the G protein hydrolyzes its bound GTP and releases the adenyl cyclase– Thus, the G protein reverts to an inactive OFF signalling
state.– The alpha subunit rejoins the beta and gamma subunits
• Adenyl cyclase, no longer poked by the activated a subunit of the G protein, shuts down – stops making cAMP from ATP
• The whole cycle results in only a brief signaling pulse– the production of several hundred cAMP molecules
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Cyclic AMP
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Second Messenger Action
• Once made, cAMP molecules act as intracellular glycogen
• The high cAMP concentrations enable A kinase to• phosphorylate and thereby activate an enzyme, that
– activates glycogen phosphorylase, which in turn – breaks down glycogen into glucose-l-phosphate
molecules; and
• phosphorylate glycogen synthase, which – turns it off, – preventing the reconversion of the released glucose to
glycogen.
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cAMP Second Messenger System
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Effect of cAMP on Blood Glucose
• These two changes together ensure the mobilization of glucose through the breakdown of glycogen stored in the liver.
• A number of other reactions are triggered as well that together contribute to the fight/flight response.
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Signal Amplification• There is enormous signal amplification in this
cascade. • A single epinephrine molecule (present at 1O-
10M) may cause the activation of dozens of alpha subunits of proteins.
• Each of these in turn will activate the synthesis of a single adenylate cyclase, and
• each of these in turn will synthesize hundreds of cAMP molecules.
• Each of these in turn can activate a cAMP-dependent kinase that will
• modify hundreds of target molecules in the cell.