Chapter 11 Biomembranes and Transport Across Biom

embranes

A cell body

A celium

mitochondrion

A digestive vacuole

The endoplasmic reticulum

A secretory vesicle

1. Biomembranes are organized, sheetlike assemblies consisting mainly of proteins and lipids

1.1 The functions carried out by membranes are diverse and indispensable for life.

1.1.1 Biomembranes give cells their individuality by separating them from the environment.

1.1.2 Biomembranes are highly selective permeability barriers with specific protein channels and pumps regulating the molecular and ionic compositions of the intracellular medium (transport across membranes).

1.1.3 Biomembranes control the flow of information between cells and their environment through specific receptors on the plasma membrane (signal transduction).

1.1.4 Eukaryotic cells have an extensive internal membrane system dividing cells into various compartments (forming different organelles).

1.1.5 The two most important energy conversion processes, photosynthesis (occurring in the inner membranes of chloroplasts) and oxidative phosphorylation (ocurring in the inner membranes of mitochondria) are carried out by membrane systems.

1.1.6 Certain biosynthesis (e.g., synthesis of lipids and some proteins, smooth ER?) occur on biomembranes.

1.2 Many common features underlie the diversity of biological membranes.

1.2.1 All membranes share a “three-layer”, sheet-like appearance (of 6 to 10 nm thick) under electron microscopic examination, with two electron-dense layers separated by a less dense central region.

1.2.2 All membranes are closed entities.

1.2.3 Membranes consist mainly of lipids and proteins with mass ratio ranges between 1:4 to 4:1 (membranes with different functions have different proteins, some lipids and proteins have covalently linked carbohydrates).

1.2.4 Membranes contain specific proteins (e.g., pumps, channels, receptors, energy transducers, and enzymes) to mediate their distinctive functions.

1.2.5 All membrane lipids are amphipathic that form bilayer structures spontaneously in aqueous media.

1.2.6 Membranes are cooperative noncovalent assemblies.

1.2.7 Membranes are always asymmetric with two different faces.

1.2.8 Membranes are fluid two-dimensional structures (the fluid mosaic model) with oriented proteins and lipids (which form bilayer structures) that can diffuse laterally but not vertically. (fig and video).

1.2.9 Most membranes are electrically polarized with inside negative (~-70 mV) (the membrane potential plays a key role in transport, energy conversion, and excitability).1.3 Proteins attach to membranes in different ways.

1.3.1 Some proteins, called integral proteins, span the lipid bilayer (e.g., glycophorin in erythrocyte, bacteriorhodopsin in halobacterium halobacterium).

Fluid mosaic model

1.3.2 Release of integral proteins from membranes requires detergents to interfere the hydrophobic interactions between proteins and surrounding lipids.

1.3.3 Released integral proteins are usually water-insoluble (easily precipitate to form insoluble aggregates) when the detergents are removed.

1.3.4 Integral proteins usually have one or more domains rich in hydrophobic amino acid residues.

1.3.5 Transmembrane domains of a protein can be predicted with reasonable accuracy through hydropathy plotting.

1.3.6 The three-dimensional structure of the photosynthetic reaction center of a purple bacterium was the first integral membrane protein to have its atomic structure determined by X-ray crystallography, where hydrophobic residues are on the exterior interacting with the lipid bilayer.

1.3.7 Some proteins, called peripheral proteins, are bound to membranes loosely and reversibly.

1.3.8 Peripheral proteins can be released from membranes by relatively mild treatments and once released are generally water soluble.

1.3.9 Membrane proteins covalently attached to lipids of several kinds are anchored to the lipid bilayer (e.g., fatty acyl chain, farnesyl chain and others).

4. Membrane lipids are amphipathic and form ordered structures spontaneously in water4.1 All membrane lipids contain a polar (hydrophilic) head and a nonpolar (hydrophobic) tail.

4.1.1 Membrane lipids are usually represented by a circle head and one or two attached wavy or straight lines as the tails.4.2 The amphipathic membrane lipids form ordered structures in water.

4.2.1 Amphipathic lipids form oriented monolayers at air-water interfaces. (experiment?).

4.2.2 Fatty acids, lysophospholipids (glycerophospholipids lacking one fatty acyl group) forms the globular micelles in water.

4.2.3 Phospholipids and glycolipids in aqueous media favorably form bimolecular sheets (lipid bilayers) rather than micelles. This is because the two fatty acyl tails are too bulky to fit into the interior of a micelle. (cylindrical-shaped versus wedge-shaped).

4.2.4 The formation of lipid bilayers from phospholipids and glycolipids is rapid and spontaneous, stabilized by the full array of weak interactions.

4.2.5 Lipid bilayers have an inherent tendency to be extensive (due to diffusion? function?).

4.2.6 Lipid bilayers tend to close themselves (to limit the amount exposed hydrocarbon chains), generating artificial structures called liposomes.

4.2.7 Lipid bilayers are self-sealing because a hole in a bilayer is energetically unfavorable (driven by hydrophobic interaction and diffusion).4.3 Liposomes can be used to carry membrane impermeable substances into cells.

4.3.1 Water-soluble substances (e.g., proteins, nucleic acids, drugs) can be encapsulated into liposomes.

4.3.2 Liposomes can fuse with cell plasma membranes (a lipid bilayer), releasing substances into cells (can be used as drug delivery tools).

4.3.3 Liposomes are used as model systems to study membrane permeability (or membrane protein reconstitution).

2. Proteins facilitate ions and solutes to move across the hydrophobic membranes in various ways.

2.1 Simple diffusion of ions and polar molecules in living organisms is impeded by selectively permeable biomembranes.

2.1.1 Only relatively nonpolar molecules (like O2, N2) cross biomembranes by simple diffusion (i.e., move from higher concentration area to lower one until they become evenly distributed).

2.1.2 Water, though polar, diffuses rapidly across biomembranes by mechanisms not fully understood. High concentration (55M) may be the reason.

2.2 Most ions and polar solutes move across biomembranes by carrier-mediated transport.

2.2.1 Carriers are usually proteins (also called pumps (active), transporters or permeases （透 ( 性 ) 酶） (passive)).

2.2.2 Carrier proteins are similar to enzymes, lowering the activation energy of simple diffusion process, by providing an alternative hydrophilic transmembrane pathway.

2.2.3 Initial rate kinetics of protein-mediated transport is very similar to Michaelis-Menton kinetics of enzymes (showing a hyperbolic curve with similar saturation effect).

2.2.4 An equation similar to the Michaelis-Menton equation describes carrier-mediated solute transport across biomembranes. (fig.12-16, p.412)

2.2.5 Carrier proteins are often stereospecific, like enzymes (e.g., the widely existing glucose transporter is highly specific for D-glucose 1.5mM, D-mannose/D-galactose 20-30mM, L-glucose 3000mM, Ktransport reflects the affinity to substrate).

2.3 In facilitated diffusion (also called passive transport) solutes move downhill its concentration gradient.

2.3.1 Facilitated diffusion usually results in equilibrium of a solute across the biomembrane leading to no net accumulation of the solute (unless the solute has a significant different solubility on the two sides or the solutes has a net charge). (electrochemical potential).

2.3.2 Glucose transporter in some animal cells mediate facilitated diffusion of glucose across the plasma membranes (GluT1 from the blood plasma to erythrocytes, GluT2 from hepatocytes to the blood plasma).

2.3.3 Chloride-bicarbonate exchanger (also called anion exchange protein or band 3 protein) on the erythrocyte membrane mediates bidirectional exchange of HCO3- and Cl- by facilitated diffusion (not active transport). It is also called cotransporter (may be active).

2.3.4 Facilitated diffusion process is independent of metabolic energy.

2.4 In active transport, solutes move against the concentration gradient (uphill) resulting in the accumulation of a solute on one side of a membrane.

2.4.1 Active transport is thermodynamically unfavorable (endergonic) and occurs only when an energy source (exergonic process) is coupled.

2.4.2 In primary active transport, energy is provided directly by the hydrolysis of ATP (as with the Na+-K+ ATPase on vertebrate plasma membranes), by electrons flowing down an electron transport system (as with H+ pumping out of the mitochondria inner membranes), or by absorption of sunlight (as with the light-driven H+ pumping of bacteriorhodopsin in halobacterium).

Directly coupled to an energy source or a chemical reaction.

2.4.3 In secondary active transport, ion gradients across the membrane, themselves created by active transport systems (initially by the primary active transports), are used to drive the concentration uptake of other ions or metabolites.

2.4.4 Many cells contain secondary transport systems that couple the spontaneous, downhill flow of H+ or Na+ to the simultaneous uphill pumping of another ion, sugars, or amino acids.

2.4.5 Lactose is transported into E.coli cells through a secondary active transport involving symport of H+ and lactose by the galactoside permease.

2.4.6 Glucose and certain amino acids are transported into intestinal and kidney cells by symport with Na+ (while the Na+ gradient is established by the Na+-K+ ATPase).

2.4.7 Some antibiotics act by shuttling ions across membranes: valinomycin (a cyclic peptide of 12 residues) binds K+ and moves across the membrane, while gramicidin A (a linear small peptide of 15 residues) form a static pore in the membrane through which cations can pass.

2.4.8 These ion shuttling antibiotics (also called ionophores （离子载体） , hydrophobic on the outside) kill bacterial cells by disrupting secondary transport processes and energy-conserving reactions. (destroys the ion gradients).

2.5 Na+-K+ ATPase is responsible to maintain the Na+ and K+ gradients across animal cell plasma membranes.

2.5.1 Most animal cells have a high concentration of K+ (145 mM) and a low one of Na+ (5 mM) relative the external medium (K+, 5mM; Na+, 150 mM?). (fig.)

2.5.2 The Na+-K+ gradient in animal cells controls cell volume, renders nerve and muscle cells electrically excitable, and drives the active transport of sugars and amino acids.

2.5.3 Jens Skou identified (1950s) an enzyme from crab nerves that hydrolyzes ATP to ADP only in the presence of both Na+ and K+. 2.5.4 This Na+-K+ ATPase was subsequently shown to be responsible to pumping Na+ and K+ across membranes against their concentration gradients (Skou shared the 1997 Nobel Prize in Chemistry for this discovery).

2.5.5 For each molecule of ATP hydrolyzed (to ADP and Pi), two K+ ions are pumped inward, and three Na+ pumped outward across the plasma membrane.

2.5.6 More than a third of the ATP consumed by a resting animal is used by this ATPase to pump Na+ and K+.

2.5.7 Ouabain (a cardiotonic steroid isolated from a plant called foxglove) was found specifically bound to the ATPase from the extracellular face, blocking both the ion pumping and ATP hydrolysis, indicating the obligatory coupling of these three processes as catalyzed by a single enzyme (elaborate mechanism?)

2.5.8 It has been demonstrated that K+ has a much higher affinity for the outside surface of the enzyme, while Na+ binds much more tightly to the cytoplasmic side.

2.5.9 It has also been an established fact that Na+ triggers phosphorylation, whereas K+ triggers dephosphorylation of the enzyme (ATPase).

2.5.10 A model has been proposed on the mechanism of Na+-K+ pumping of the enzyme, mainly supposing that the enzyme cycles between two conformations, a phosphorylated one binding to K+ with high affinity (from the outside face of the cell, and to Na+ with low affinity, thus releasing Na+ ions to the outside face), and a dephosphorylated one binding to Na+ with high affinity (from the inside face of the cell, and to K+ with low affinity, thus releasing K+ ions to the inside face).

2.6 Ion-selective channels enable specific ions to flow through biomembranes rapidly downhill its concentration gradient.

2.6.1 Ion channels form pores that open and close quickly.

2.6.2 The opening and closing of the ion channel pores can be regulated by ligands or voltage change (called ligand-gated and voltage-gated channels, respectively).

2.6.3 Acetylcholine （乙酰胆碱） receptor, sodium channel and potassium channel are among the best studied ion channels.

2.6.4 The sodium and potassium channels mediate the transimission of nerve impulse (or atction potential) down the motor neuron (triggering a wave of depolarization).

2.6.5 Acetylcholine receptor mediates the transmission of nerve signals across synapses: binding of acetylcholine (released from the presynaptic neuron) to the receptor opens the ion channels, allowing Na+ and K+ (Ca2+) to diffuse through quickly down their concentration gradients (thus depolarizing the postsynaptic neurons or muscle cells).

2.6.6 Rate of ion movement across the opened ion channel is not saturable in the way that carrier-mediated translocation is saturable in the transporting of ions or solutes (this kinetic property distinguishes ion channels from carrier mediated transport across membranes). (different molecular mechanims).

2.6.7 The flow of ions through a single channel and the transition between different states were monitored with a time resolution of microseconds using the revolutionary patch-clamp （膜片钳） techniques by Erwin Neher and Bert Sakmann in the 1970s (they won the 1991 Nobel Prize in medicine or physiology for such high resolution studies on ion channels).