Mechanism of Transport across Cell Membrane

1.         Simple Diffusion:

a.         a passive process by which uncharged particles in solution flow down their concentration (chemical) gradient (ie. particles move from areas of high concentration to areas of low concentration.

b.         no external source of energy (or driving force) is required to move particles down a concentration gradient by diffusion.

c.         simple diffusion occurs because the heat content of the solution keeps all of the solvent and solute particles of the solution in constant motion.

d.            process:

i.          each particle moves in an unpredictable (random) fashion; however it is more likely that a particle will move from an area of high concentration to an area of lower concentration.

ii.          net movement ceases when the concentration of the particles equal everywhere within the solution (diffusional equilibrium).

iii.            although random movement of the particles continues after diffusional equilibrium is achieved, the concentration of the particles throughout the solution remains the same.

e.         The selectively permeable plasma membrane regulates the type and rate of molecular traffic into and out of the cell.

f.          It depends on:

i.            membrane solubility characteristics of the phospholipid bilayer.

ii.            presence of specific integral transport proteins.

g.            Permeability:

i.          lipid-soluble particles diffuse through the bilayer of the cell membrane; their permeability is proportional to their lipid solubility.

ii.          water-soluble particles: diffuse through the aqueous channels formed by transmembrane proteins; their permeability is inversely proportional to their molecular weight and charge.

h.         The membrane impedes the entry of ions because:

i.            insolubility of ions in hydrophobic portions.

ii.            electrical charge of ions causes molecules of water to become bonded to ions, forming hydrated ions.

iii.         the membrane surface, being externally negatively charged and internally positively charged, will repel all ions away.

2.            Facilitated Diffusion:

a.         is a carrier-mediated process that enables particles which are previously too large or polar to pass through the cell membrane to enter it through membrane channels by simple diffusion down their concentration gradient.

b.            properties:

i.            specificity: the carrier can move only one molecule or a group of closely related molecues, eg. glucose transporter can move glucose and other 6 carbon sugars.

ii.            competition: a carrier has different affinities for the molecules it transport .eg. increase in glucose concentration will reduce the amount of fructose transported as the carrier has a higher affinity for glucose than for fructose.

iii.            saturation: this occurs when a group of membrane carrier proteins are transporting substrate at their maximum rate; this can be prevented by inserting additional carriers in the membrane.

c.            process:

i.          no external source of energy is required to move particles down a concentration gradient by facilitated diffusion.

ii.          upon binding of the solute, the carrier protein undergoes repetitive spontaneous conformation changes during which the binding site for the solute is alternately exposed to the intracellular fluid and extracellular fluid.

iii.         carrier protein bind solute in one conformation and deposit it on the other side of the membrane in another conformation.

iv.         the solute’s binding and release may trigger the carrier protein’s conformational change.

v.         the solute is more likely to bind to the carrier where it is more highly concentrated and dissociate from the carrier where it is less highly concentrated; therefore, the solute flows down its concentration gradient.

d.         rate:

i.          the rate of facilitated diffusion increases as the concentration gradient increases until all of the binding sites are filled.

ii.          at this point the rate of diffusion can no longer increase with increasing particle concentration and is called saturation.

e.         net rate of diffusion: refers to the difference between the concentration of substance diffusing in both directions and the factors affecting it are:

i.            permeability of membrane.

ii.            difference in concentration of diffusing substance between two sides of membrane.

iii.            pressure difference across membrane.

iv.            electrical potential difference between two sides of membrane.

f.          The selective permeability of protein channels results from certain inherent features:

i.          shape of protein molecule.

ii.            diameter of channel

iii.         nature of electrical charges along its surface.

iv.         in Na channels, the inner surfaces of these channels are strongly negatively charged; these negative charges pull the Na ions more strongly than the other ions.

v.         in K channels, the small hydrated K ions can pass easily through it while the Na ions are rejected.

g.            Transport proteins are integral membrane proteins that transport specific molecules or ions across biological membranes:

i.          may provide a hydrophilic tunnel through the membrane for the transfer of hydrophilic molecules in and out of the cell.

ii.          carrier proteins transport molecules by change of conformation.

iii.         are specific for the substance they translocate; presence of binding sites for the particular substrate molecule which activates the transporter.

h.         Carrier types:

i.            uniporters: carriers that transport a single particle in one direction, such as the facilitated diffusion of glucose.

ii.            symporters transport two particles in the same direction, such as the secondary active transport of glucose.

iii.            antiporters transport molecules in opposite directions, such as the Na-Ca and Na-H exchangers.

i.          some channels are continuously open whereas others are gated; they have gates that open and close.

j.            voltage-gated channels:

i.          gated by alterations in membrane potential.

ii.          sodium pump: when the inside of the cell is negatively charged. the gates remain close; when the inside of the cell loses negative charge, these gates open and allow Na ions to pass inward through the Na pores.

k.         ligand-gated channels:

i.          some protein channel gates are opened by the binding of another molecule with the protein, thus causing a conformation change in the protein molecule that opens or closes the gate.

ii.          the binding molecule is the ligand.

iii.         the ligand is often external as in the neurotransmitter acetylcholine, which upon binding onto acetycholine channels on the postsynaptic membrane, causes the opening of Na channels which continues the transmission of impulses along the neuron.

iv.         it can also be internal: intracellular Ca ions, cyclic AMP, or one of the G proteins produced in cells can bind directly to channels and activate them.

l.          gating of protein channels:

i.            provides means for controlling the permeability of the channels.

ii.          the gates are actually gate-like extensions of the transport protein molecule, which can close over the opening of the channel or can be lifted away by a conformational change in the shape of the protein molecule itself.

m.            facilitated diffusion accomplishes the transport of glucose into red blood cells and muscle and into adipose tissue when insulin is present:

i.          the glucose transporters that are responsible for facilitated diffusion of glucose across cell membrane are a family of closely related proteins that cross the membrane 12 times.

ii.          they appear to surround channels that glucose can enter; conformation then changes and glucose is released inside the cell.

n.            cysinuria: kidney disease where carriers are missing for cystine.

3.         Active Transport

a.         all cells have a resting membrane potential of –70mV across the cell membrane.

b.         two factors that drive passive transport of ions across membranes:

i.            concentration gradient of the ion.

ii.          effect of membrane potential on the ion.

c.            electrochemical gradient: diffusion gradient resulting from the combined effects of membrane potential and concentration gradient (does not apply to non-ions).

d.         active transport is an energy requiring process:

i.          where a cell membrane moves molecules against an electrochemical or concentration gradient.

ii.          involves integral proteins which transport the molecules by conformational changes.

iii.         energy is derived from ATP: phosphorylation of protein changes conformation; ATP is cleaved to ADP and inorganic phosphate.

4.         Primary Active Transport:

a.         directly use the energy obtained from the hydrolysis of ATP to transport solutes against an energy gradient.

b.         most common is the sodium-potassium pump, which uses the membrane-bound ATPase as a carrier molecule.

5.            Sodium-potassium Pump (Na-K-ATPase):

a.         most important function is to maintain cell volume:

i.          control volume of cells without which they will swell until they burst.

ii.          large number of organic molecules in cell tends to draw in water by osmosis.

iii.            continual net loss of ionic molecules out of the cell, which initiates an opposite osmotic tendency to move water out of the cell.

iv.            responsible for maintaining the high K and low Na concentration in the intracellular fluid.

b.            Structure of pump:

i.          a heterodimer made up of 2 alpha subunits (mw=100,000) and 2 beta subunits (mw=55,000).

ii.          both extend through the cell membrane; separation of the subunits inhibit activity.

c.         alpha subunit:

i.            transport of Na and K occurs through the alpha subunit.

ii.          spans the cell membrane 8 times.

iii.         has ATPase enzymatic activity: the ability to convert ATP to ADP, thereby releasing energy.

iv.         the intracellular face contains binding sites for 3 Na ions, an ATP molecule and for phosphorylation to take place.

v.         the extracellullar face contains binding sites for 2 K ions.

d.         beta subunit:

i.          it is a glycoprotein.

ii.          has a single membrane-spanning domain and 3 extracelluar glycosylation sides, all of which have attached carbohydrate residues.

e.            Process:

i.          when 3 ions of Na and an ATP molecule bind to the carrier on the inside of the cell. a phosphate group is transferred from the ATP molecule to an aspartic acid residue of the alpha subunit.

ii.          the addition of a high-energy phosphate group causes a conformational change in the protein, which results in the transport of the 3 Na ions out of the cell.

iii.         when 2 ions of K bind to the carrier on the outside of the cell, the aspartic acid-phosphate bond is hydrolyzed.

iv.         the energy released by this dephosphorylation step results in a second conformational change during which the 2 K ions are transported into the cell.

f.          Effects of Na & K Transport:

i.          the pump is an electrogenic pump as it creates an electrical potential across the membrane as it pumps, ie. net exodus of positive charge.

ii.            accounts for a large part of basal metabolism: 33% of energy utilized by cells and in neurons it accounts for 70%.

iii.         link between Na-K transport and metabolism: greater the pumping, the more ADP is formed, and the available supply of ADP determines the rate at which ATP is formed by oxidative phosphorylation.

g.         the Na-K pump is found in all parts of the body; in some tissues, the active transport of Na is coupled to the transport of other substances.

h.            Inhibition of pump:

i.          digitalis, a drug used for the treatment of heart failure, produces its therapeutic effect by binding to the extracellular face of the alpha subunit and interfering with the dephosphorylation step the transport process.

ii.          the pump requires binding by Na, K and ATP for its operation; therefore, if the concentration of any of these substances are too low, the pump does not function.

6.         Other primary active transport systems that directly rely on the hydrolysis of ATP to transport ions across membranes are:

a.            Calcium pump:

i.          found on the sarcoplasmic reticulum of muscle cells.

ii.          Ca ions are maintained at an extremely low concentration in intracellular fluid.

iii.         one is in the cell membrane and pump Ca to outside of the cell.

iv.         the other pumps Ca into internal cell organelles like mitochondria, sarcoplasmic reticulum.

b.            Potassium-hydrogen (K-H) pump of gastric mucosa cells, which affects the secretion of H ions into the stomach during the digestive process.

7.            Secondary Active Transport (cotransport):

a.         process where a single ATP-powered pump actively transport one solute and indirectly drives the transport of other solutes against their concentration gradients.

b.         use the energy stored in the Na concentration gradient to transport material against an energy gradient.

c.            Functions:

i.          the transport of many ions and nutrients against their electrochemical energy gradients is accomplished by Na dependent secondary active transport.

ii.          glucose and amino acids are reabsorbed from the proximal tubule and absorbed from the intestinal lumen by Na dependent secondary active transport mechanisms.

iii.         calcium is removed from the cytoplasm of cardiac ventricular cells by a Na dependent secondary active transporter called the Na-Ca exchanger; this mechanism causes muscle relaxation.

d.            Process:

i.          the energy contained in the Na electrochemical gradient causes a conformational change in the carrier molecule.

ii.          when Na binds to the carrier molecule, the carrier molecule increases its affinity for the substance to be transported.

iii.         when both Na and the substance are bound to the carrier molecule, the carrier undergoes a conformational change during which both molecules are transported across the membrane.

iv.         in some cases, both Na and the substance are transported in the same direction, whereas in others they are transported in opposite directions.

8.         Na-glucose cotransport in intestinal lumen:

a.         two types of carrier proteins are present in the mucosa cells:

i.          apical region: Na-glucose symporter

ii.            basolateral region: Na-K antiporter and glucose transporter

iii.         the transporting epithelial cells are polarized because their apical and basolateral membranes have different proteins due to the uneven distribution of membrane proteins.

b.         Na-K antiporter maintains low Na concentration in the intracellular fluid, by pumping Na out of the basolateral membrane by means of a Na-K-ATPase, hence generating an electrochemical gradient.

c.         in the apical membrane Na binds to Na-glucose symporter  together with glucose.

d.         this causes a conformational change in the carrier protein which releases both the glucose and Na into the interior of the cell.

e.         energy for transport of glucose against its concentration gradient is derived from the concentration gradient of Na which is maintained by the Na-K antiporter.

f.          once glucose is in the cell, it leaves by moving down its concentration gradient through the facilitated diffusion carrier in the basolateral membrane.

9.         Other antiporters:

a.         Na-H exchanger regulates cytosolic pH via influx of Na and efflux of H.

b.         Na-HCO₃ exchanger is an electrogenic symporter that brings in two or more HCO₃ molecules with each Na.

c.         Na-driven CI-HCO₃ exchanger that couples an influx of Na and HCO₃ to an efflux of CI and H.

d.         Na-independent CI-HCO₃ exchanger.

10.            Transcytosis and vesciular transport:

a.         all molecules leaving and entering an epithelium must cross two cell membranes.

b.            molecules cross the first membrane when they move into an epithelial cell from the external environment, and the second when they leave the epithelial cell to enter the extracellular fluid.

c.         some molecules that are too large to cross membranes on protein carriers are brought into the body by transcytosis.

d.         in this process, the particle is absorbed on one side of the cell using pinocytosis, receptor-mediated endocytosis.

e.         the vesicle that results attaches to microtubules in the cytoskeleton and is transported across the cell into the extracellular fluid by exocytosis.

f.          this movement of vesicles across the cell is known as vesicular transport.

g.            transcytosis makes it possible for large proteins to move across an epithelium and remain intact and is the means by which infants absorb maternal antibodies in breast milk.