Membrane Transport of
Small Molecules and the
Electrical Properties of
Membranes
Chapter 11, Molecular Biology of the Cell
Jin-Chung Chen
Lipid bilayer: to separate the inner and outer
environment of the cell; require special design of
carrier protein to transport the water-soluble nutrient
into the cell
15-30% membrane proteins exert function of
transport (special case: 2/3 of the metabolic
energy consumed on transport in special cell)
The higher the hydrophobicity or non-polar
molecule make ease to pass through the
membrane structure
Carrier protein disorder: cystinuria-cystine
accumulates in the uria formed kidney stone
Two classes of transfer protein:
(1) Carrier protein (permease, transporter,
pump) : for specific molecule; usually
coupled with energy source
(2) Channel protein: inorganic ions; down to its
concentration gradient; fast
overall, transfer proteins create electrical
(because of membrane potential) and
concentration gradient, in turn, used as a
driving force (electrochemical gradient) to
facilitate the transport process
Electrochemical gradient combines with
membrane potential as a driving force
Involved conformation change
Both carrier and channel
contain specialized
transmembrane domain
Transport through channel: fast transport
Passive transport: facilitated diffusion for all channel protein
and part of carrier (concentration or electrochemical gradient)
Active transport: against electrochemical gradient; mediated
by carrier (pump); use ATP or ion gradient as energy source
Ionophores: small hydrophobic molecules (originally
formed by microorganism) in membrane to transport
specific ions; not coupled to energy source (down to
concentration gradient)
1.
2.
3.
4.
Valinomycin: potassium ion (mobile)
FCCP: hydrogen ion (mobile)
A23187: calcium and magnesium ion (mobile)
Gramicidin A: monovalent cation (channel former)
Model of Carrier protein: passive transport
Kinetics of simple diffusion and carrier-mediated
diffusion (expressed as Vmax/Km or Bmax/Kd)
Three ways of driving active transport:
(a) coupled carrier: downhill solute drive uphill solute
(b) ATP-driven: ATP hydrolysis to drive uphill solute
(c) light-driven: light energy trigger uphill solute
bacteriorhodopsin
Three types of carrier-mediated transport: uniport, symport
( kidney/GI epitghelial cells ) and antiport (determined by its
path direction)
Application of Na+-driven carrier proteins (use
ion gradient as energy source to drive solute):
a. Na+ driven glucose into the cell (stomach brush border)
(symporter)
Na+ pump that regulates the cellular pH :
a. Na+-H+ exchanger: influx of Na+ while pump out H+
(maintain the inner pH of the cell)
b. Na+-driven Cl-HCO3- exchanger: influx Na+ and
HCO3- while pump out H+ and Cl- (maintain pH of the
cell)
c. Na+-HCO3- symporter of the glial cell: electrogenic,
pump in one Na+ and two HCO3- help to regulate the
extracellular pH in near-by neurons during electrical
activity
Transcellular transporter: apical to basolateral transport
nutrients (intestinal epithelial cells: microvilli)
P-type transporter: 1. Na+-K+ pump
1. [K+]i: [K+]o = 20 : 1 whiles [Na+]i: [Na+]o = 1: 15
across the cell membrane;
2. The concentration difference is maintained by Na+-K+
pump (sodium pump; all the animal cells)
3. The pump is an antiporter, executes active transport
and also is an ATPase; its consists of a catalytic subunit
(1000 a.a.) and a small glycoprotein (critical for
membrane docking)
4. Most important and energy consumption protein on
the membrane
5. Pump 3 Na+ out in exchange of 2 K+ in (electrogenic)
Immunocytochemical
localization of the Na,K pump
in choroid plexus. Choroid
plexus contains epithelial
cells with intensely stained
microvillar and intermicrovillar
plasma membranes. The
basal and lateral plasma
membrane surfaces are not
stained. Bar = 2 µm.
Schematic diagram of Na-K pump (P-type transport
ATPase: can be reversed experimentally to produce
ATP
Aspartic acid
Na+-dependent phosphorylation;
K+-dependent dephosphorylation
Down-regulation of Na,K pumps can be initiated by dopamine
via GPCR activation of endocytosis
Na+-K+ pump regulates cell volume
1. Water move along with the solutes determines the
cell volume (osmolarity; tonicity)
2. Hypotonic vs. hypertonic solution
3. The fixed anions (nucleic acid and proteins) inside
the cell try to pull the water molecule; while
excellular Na+ (driven by pump) and Cl- (expelled by
membrane potential) ions balance the force
4. Ouabain: Na-K pump inhibitor; cell swell and burst
due to break up the net Na+ outflow
Calcium homeostasis:
Cytoplasmic Ca2+ is regulated
coordinately by a Na+/Ca2+ antiporter in plasma membranes and by Ptype Ca-ATPases in plasma membranes and endoplasmic reticulum.
P-type transporter: 2. Calcium pump (Ca2+-ATPase)
1. Eucaryotic cells: [Ca2+]i: [Ca2+]o = 10-7 M: 10-3M
2. Ca2+ pump located in the endoplasmic reticulum
(SR in muscle cell consist of 90% of membrane
protein)
3. Ca2+ pump of the SR brings the calcium ion into the
SR for storage, upon activation release the calcium
into the cytosol for Ca2+-dependent cell signaling
4. Contains 10 transmembrane -helices; two calcium
ion bind to calcium binding domain cause
phosphorylation and lead Ca2+ release into the SR
ABC Transporter: transport ATPase
1. Largest transport ATPase family (> 50 members)
2. Contains four domains: two hydrophobic domains each with
six transmembrane span (function to translocate) and two
ATP-binding cassettes (ABC)
3. In procaryotes, the transporter locates in the inner
membrane to carry nutrients into the cell
4. The counterpart of the eucaryotic: multidrug resistance
(MDR) protein, which produce resistance to drug (i.e.
chloroquine and anticancer drug resistance)
5. Cyctic fibrosis: a mutation on one ABC transporter (cyctic
fibrosis transmembrane regulator [CFTR] protein) that
function as a Cl- channel in the epithelial
Gram-negative bacteria
(cassettes): ABC
Ion channels: channels mediate inorganic ion transport
1. Narrow, highly selective pores that can open and close
2. Approximately 100 million ions can pass through / second
(105 times greater as compared to any carrier)
3. Can not couple to energy source to perform active
transport (always passive – downhill)
4. They are gated (open and close status); prolong stimulation
would desensitized or inactivated ( closed; usually through
phorphorylation)
5. All animal cells contain ion channels, not limited to neuron;
each neuron might have more than 10 types of channel
Ion channels open in response to special stimulus
Establish the Membrane Potential
1. Active electrogenic pump (such as Na-K pump) and
passive ion diffusion create the difference of ionic
strength across the membrane; hence the difference in
electrical charge on two side of the membrane
2. Major force: Na+-K+ pump and K+ leaking channels
(more positive excellularly as compared to negative
intracellularly)
a. High concentration of potassium ion inside the cells
(due to Na-K pump) keep the electric balance of the
negative charged macromolecule
b. K+ leak out down to its concentration gradient
3. Resting membrane potential: no net flow of ions across
the membrane
Number of ions that move across the membrane to set up the
membrane potential actually is quite minute
Nernst Equation (and Goldmann equation)
RT
Co
V = zF ln Ci
V= the equillibrium potential in volts
Co/Ci = outside and inside concentration of
the ion, respectively
R= the gas constant (2 cal mol-1K-1)
T= the absolute temperature (K)
F= Faraday’s constant (2.3 x 104 cal V-1
mol-1)
z= the valence (charge) of the ion
ln= logarithm to the base e
The more permeable the membrane for a given
ion, the more strongly the membrane potential
tends to be driven toward the equilibrum value
for that ion:
changes in a membrane’s permeability to ions
can cause significant changes in the membrane
potential ( resting membrane potential will be
determined by most permeable ion(s) )
Diagram of the functional units of a voltage-gated Na+ channel,
the hypothesized binding sites for several drugs and toxins are
illustrated to affect the opening and closing of the channels
Structural model of the Na+ channel.
Transmembrane organization of voltage-gated
Ca2+ channels, K+ channels and relatives.
3-D Structure of Potassium Channel
(exhibited only 2 out of 4 transmembrane
subunits)
Separated
about 8 A
Selection of ion by selectivity filter
Sodium ion is too
small for carbonyl
oxygen interaction
and water expel
The ion selectivity filter and pore of Na+ and K+ channel
illustrated with the extracellular side upwards.
The four ion-coordination sites are
thought to work in pairs. In one
cycle of outward K+ conductance,
K+ ions occupy sites 1 and 3
(orange), shift to sites 2 and 4 (gray),
and then the K+ ion in site 4 moves
into the extracellular space while a
new K+ ion occupies site 1 and the
K+ ion in site 2 moves to site 3, reestablishing the initial state.
Typical nerve cell: somatodendrites and axon
Generation of an Action Potential
1. An electrical stimulus that exceeds a certain threshold
triggers an electrical activity that is (a) self-propagated and
(b) sustained by automatic amplification
2. Voltage-gated cation channels are responsible for
generating the action potential
3. Consequence of an action potential:
a. stimulus triggers voltage-gated Na+ channel to open
b. more Na+ channel open in neighboring area, cause
depolarization (reach Na+ equilibrum potential [+50mV])
c. Na+ channel inactivated
d. voltage-gated K+ channel open (bring membrane potential
to resting status; reach K+ equilibrium potential [-70mv])
N-terminal 20 a.a. function as a tethered ball to
inactivate the potassium channel after activation
A hinged-lid model for Na+ channel inactivation,
illustrating the inactivation gate formed by the
intracellular segment connecting domains III and IV
and the critical cluster of hydrophobic residues
that forms a latch holding the inactivation gate
closed.
Function of Myelination:
insulated neurons to
increase the rate of
propagation; formed by
Schwann cells (peripheral)
or oligodendrocytes (central)
Node of Ranvier: myelin
sheath space; most Na+
channels concentrated here
Saltatory conduction:
action potential propogate
node to node (travel faster
and metabolic energy is
conserved)
Patch clamp technique : allow current recording
from single channel
Individual volgate-gated Na+
channel open in all-or-none
fashion; the aggregate curent
(approached by intracellular
recording) across the
membrane of entire cell
represent the total number
of channels ( not the degree)
that open at a given time
Voltage-gated cation
channels (Na+, K+ or Ca2+ )
are evolutionarily and
structurally related
Neurotransmission from presynaptic to postsynaptic
Re-uptake
Transmitter-gated ion
channel is not sensitive to
membrane potential (can
not produce selfexcitation). They produce
local permeability changes
(depend on numbers of
released transmitter) until
can open nearby voltagegated cation channels to
initiate an action potential
of the next neuron
Cl--dependent
transporter
Neurotransmitter can determine post-synaptic
excitation or inhibition (channel receptor)
A. Excitatory neurotransmitter: open cation
channel (Ca2+, Na+) (i.e. NMDA glutamate
receptor)
B. Inhibitory neurotransmitter: open Cl- or K+
channel (i.e. GABAA receptor)
Model of nicotinic acetylcholine receptor
Consequence of neuromuscular transmission
1. Nerve impulse reach nerve terminal (depolarization) to
open voltage-gated Ca2+ channels; calcium ion flow
into the nerve terminal triggers ACh release
2. ACh binding to acetylcholine receptor (nACh) in
muscle membrane; opens the cation channels and Na+
influx initiate local depolarization
3. Local depolarization opens more voltage-gated Na+
channels and result in action potential
4. AP activates voltage-gated Ca2+ channels in transverse
[T] tubules
5. In turn, causes Ca2+ release channels in SR to open and
trigger actin-myosin sliding / muscle contraction
Actual look of the
synapses in a cell body
Signal summation at PSP
1. Cell body and dendrites of a single neuron are covered by
numerous synapse (presynaptic terminals)
2. Excitatory postsynaptic potential (EPSP): presynaptic
transmitter release evoke excitatory response
3. Inhibitory postsynaptic potential (IPSP): presynaptic
transmitter release evoke inhibitory response
4. All the PSPs from the dendritic tree converge at the cell
body: spatial summation
5. The incoming signals summate at a given time (translate
frequency into magnitude of PSP): temporal summation
6. If net excitatory input predominate: combined PSP is a
depolarization; otherwise a hyperpolarization
Temporal summation:
Each presynpatic action
potential arriving at a synapse
produces a small postsynaptic
potential (PSP). When
successive action potentials
arrive at the same synapse, each
PSP produced adds to the tail of
the preceding one to produce a
larger combined PSP.
Firing frequency of an axon determines the magnitude of
combined PSP
Characters of Neuronal Computation
1. Action potentials are initiated at the axon hillock:
concentrated voltage-gated Na+ channels
2. Axon hillock also contain delayed K+ channels
(repel K+ after AP-induced Na+ inflow), early K+
channels (reduce rate of firing) and Ca2+activated K+ channel (neuronal adaptation after
prolonged and constant stimulation)
Learning and Memory
(molecular mechanism of Long-term Potentiation)
1. LTP: a short burst of repetitive firing leads to
subsequent single AP in the presynaptic cells evoke a
greatly enhanced response in the postsynaptic neurons
2. Major brain area: Hippocampus
3. Presynaptic glutamate release evokes postsynaptic nonNMDA receptor (AMPA) activation EPSPs and
action potential (Na+ inflow) remove Mg2+ ion of the
NMDA receptor NMDA activation (Ca2+ inflow)
trigger cascade response responsible for LTP
a. insert more AMPA receptor in postsynaptic cell
b. enhanced presynaptic glutamate strength
Essential membrane components during normal
synaptic transmission (excitatory synapse)
Cellular alteration during the formation of LTP
Most critical event in LTP formation in postsynaptic
neurons: New AMPA receptor insertion (exocytosis)