Chapter 10. Membrane Structures Importance of membranes to cells: o Plasma membrane separates cellular components from the environment o Allows organelles to execute specialized functions by keeping the contents of the organelle separate from the rest of the cell o Movement of “cargo” from one location in a cell to another o Provides boundaries that establish electrochemical gradients. These are used to make ATP and to generate nerve impulses o Has proteins embedded in them to adhere to surfaces and other cells o Biological membranes are thin films composed mainly of amphiphilic lipids and proteins. Most of the molecules are held together by noncovalent interactions. This model of the membrane is known as the Fluid Mosaic Model Important Properties of Lipids: o Lipid molecules organized into a bilayer approximately 5 nm thick o Each layer of the bilayer is called a leaflet o Lipids diffuse rapidly in the plane of the bilayer, but do not flip from one leaflet to the other without assistance from specific proteins (ATP-dependent flippases) o Self-sealing – when one microinjects something into a cell, the membrane seals automatically when the needle is withdrawn o Purified phospholipids spontaneously from bilayers in water Membrane Proteins: o Proteins embedded in the membrane span from one side to the other o Proteins diffuse in the plane of the bilayer unless they are anchored to something o Proteins do not flip-flop o Lipids provide the basis for the spontaneous formation of membranes o Lipids are amphiphilic meaning they have a hydrophobic part (nonpolar tail) and a hydrophilic part (polar head) o Lipids pack to minimize the interaction between water and the nonpolar part of the molecule o Because the phospholipids used to make membranes typically have 2 tails…they pack together to form a phospholipid bilayer to minimize contact between water and their hydrocarbon chains What affects how lipids move...? o Movement of lipids should: o Maintain favorable interactions: water and the polar head groups o Avoid unfavorable interactions: water and the nonpolar tails o Therefore… biological membranes behave like two-dimensional (2D) fluids o For example: a typical lipid can diffuse the whole length of a bacteria in 1 sec o Sweet Spot: the fluidity of a biological membrane must be maintained to sustain transport and enzymatic activities. The fluidity is influenced by temperature and lipid composition o Cis-double bonds (unsaturated tails) and short hydrocarbon chains increase fluidity because the tails can’t pack as closely (due to the kinks in the chains) o Absence of double bonds (saturated tails) and long hydrocarbon chains decrease fluidity, because the tails pack more closely Membranes of animal cells are composed primarily of three types of lipids: o Phospholipids – most abundant o Cholesterol o Glycolipids Properties of cholesterol: o o o o Hydroxyl group (OH) is the polar head group Hydroxyl is attached to rigid steroid ring One hydrocarbon tail Found in eukaryotes, not in bacteria What is the effect of cholesterol on membranes? o Cholesterol tightens the packing near the polar head groups and decreases the permeability (leakiness) of the membrane Some lipids are asymmetrically distributed o Glycolipids are restricted to the extracellular leaflet o Phosphatidylserine is restricted to the cytoplasmic leaflet in a healthy cell Why do cells create asymmetry? o Dead cells are distinguished from live cells because phosphatidylserine (PS) becomes exposed on the outer leaflet in dead cells. Macrophages (immune cells) detect the PS and destroy the dead cell Movement of proteins in proteins o Many are free to move in the plane of the bilayer but none will “flip-flop” (on their own) How can proteins become membrane proteins, or membrane-associated proteins? o o o o o o o Single pass transmembrane ™ domain (alpha helix) Multi-pass TM domain (alpha helices) Multi-pass TM domain (beta barrel) Amphiphilic alpha helix Lipid modification (covalently bonded) Sugar/lipid modification (GPI anchor) Noncovalent association with a protein that is a membrane protein Alpha helical Trans-Membrane (TM) domains o Asymmetrical arrangement forms H-bonds with itself, not fatty acid o We can predict TM domains bioinformatically with some skill Beta Barrel Trans-Membrane (TM) domains o Some can act as transporters (regulated pore) o Beta sheets can provide the necessary hydrogen bonds via backbone interactions, too Proteins can receive post-translational modifications (PTMs) o Carbohydrates, lipids, etc. Modifications can provide additional functions to the proteins o Carbohydrates/ oligosaccharides can serve as recognition signals for extracellular events o Lipids can allow proteins to be anchored into membranes without needing a TM domain o Detergents can be used to lyse cells, extract membrane-associated proteins Detergents can be used to extract membrane-associated proteins: o Purify membrane proteins o Stabilize in a defined vesicle o Biochemical/functional assays Potential methods of restricting the motion of membrane proteins o o o o Cluster Anchor Boundary Contact Not all proteins undergo lateral diffusion. For example, attachment to the cytoskeleton immobilizes some proteins o Spectrin: cytoskeleton also givens shape to the plasma membrane o Spectrin is anchored to the membrane by associating with other proteins o Glycophorin: 1 TM domain membrane protein, binds with band 4.1, spectrin o Ankyrin: associated with spectrin o Band 3: associates with ankyrin in order to associate with spectrin Boundaries: o Even though fluidity of membrane proteins is important, in some cases localization can be restricted to defined regions of the cell membrane by other factors o Tight junctions between cells of epithelia lining surrounding many organs restrict proteins to one side of a cell o These would act as boundaries to membrane associated proteins Membrane proteins can affect the structure of the lipid bilayer o Proteins can “wedge” into one layer, pry head groups apart o Proteins can interact with the head groups without inserting o Two outcomes: induce deformation, stabilize existing curvature o Proteins can bind to specific head groups to cause them the cluster Chapter 11: Membrane Transport of Small Molecules and the Electrical Properties of Membranes Cells need to maintain a specific intracellular concentration of ions to function correctly o Cells have a preference for the balance of ions inside and outside The phospholipid bilayer is a barrier that controls the transport of molecules in and out of the cell o o o o o Gases diffuse freely, no proteins require (O2, CO2, N2, etc) Small uncharged particles diffuse slowly. Aquaporins facilitate H2O transport Large uncharged particles diffuse very slowly, involves a transporter (glucose & sucrose) Charge molecules are virtually impermeable. Involves transporters and channels Studies of synthetic lipid bilayers help identify which types of molecules are likely to involve a protein for transport Transport of molecules that are impermeable to the lipid bilayer by two classes of membrane proteins: Transporters, Channels o Transporter (“carriers”) o Conformation change “carries” the solute across the membrane o Channel protein o Aqueous pore provides a passageway so that solute molecules can diffuse through the membrane (“facilitated diffusion”) Two types of transport are defined by whether metabolic energy is expended to move a solute across the membrane o Passive transport (facilitate diffusion): no metabolic energy require, solute is moving down its concentration gradient or, if charged, down the electrochemical gradient o Active transport: metabolic energy required, solute is moving up (against) the concentration gradient, or if charged, against the electrochemical gradient o Uncharged solute: only consider the concentration gradient o Charged solute: movement across the membrane is affected by both the chemical gradient and the electrical gradient, often called the membrane potential o The combination of the two is called the electrochemical gradient A transporter binds solute on one side of a membrane, undergoes a conformational change, and releases the solute on the other side of the membrane A transporter is like an enzyme in many ways Enzyme Binding site for substrates Transporter Binding sites for solute Chemically transforms substrate Transports solute Reaction rate saturates at high substrate concentration Transport rate saturates at high solute concentration Three types of transporters are defined by the direction and number of solutes each transports o Uniport, symport, antiport A transporter can couple the conformational change to a source of energy to perform active transport o Three classes of pumps that work against gradients o Coupled transporter, ATP-driven pump, light-driven pump Energetically unfavorable reactions can be made to occur by coupling them to energetically favorable reactions The electrochemical gradient allows Na+ to power a glucose transporter to transport glucose up the glucose gradient into the cell o Note that there are specific binding sites for Na+ and glucose on this transporter o The energetically favorable movement for Na+ down its electrochemical gradient is coupled to the energetically unfavorable transport of glucose up its concentration gradient. Hence, glucose is being subjected to active transport The Na+--K+ pump (aka Na+--K+ ATPase) established concentration gradients with Na+ low inside the cell and high outside, and K+ high inside the cell and low outside The Na+--K+ pump is a “P-type” pump. “P” indicates that it phosphorylates itself. A cycle of phosphorylation and dephosphorylation is coupled to conformation changes and to the binding and release of ions resulting in active transport of ions across the membrane The phospholipid bilayer is a barrier that controls the transport of molecules in and out of the cell o Charged molecules are virtually impermeable o Their movement across a membrane involves transporters and channels Ion Channels o Multipass transmembrane proteins with alpha-helical transmembrane domains organized into a hydrophilic channel o Selectivity filter specifies which ion pass through the pore. The specificity arises because the channel narrows and forces the ions to come in contact with amino acid side-chains lining the channel o Movement of solutes by channels is MUCH faster than by transporters o Limited to passive transport Ion channels have a narrow region called the “selectivity filter” Ion channels fluctuate between open and closed states to regulate ion flow o Voltage-gated channels respond to the membrane potential o Ligand-gated channels respond to association of small molecules called ligands o Mechanically gated channels respond to movement Ions move through the channel making transient contacts with amino acids in the center of the channel o The carbonyl oxygens that line the selectivity filter are “presented” as parts of loops Ions are also selected for/ against based on charge o Ions must pass through surfaces of charge amino acid residues Understanding the electrical properties of membranes o Establishing a resting membrane potential in animal cells results from the coordinated action of a transporter and ion channels o The resting potential is defined as the membrane potential occurring when there is no net flow of ions. Cells typically have resting potentials between -20 and -200mV o The changes in membrane potential are achieved by the movement of very few ions close to the lipid bilayer. Hence, the overall concentrations of potassium and sodium inside and outside the cell do not change when various channels open and close. Sodium stays high outside and potassium stays high inside. Small fluxes of ions are sufficient to change potentials o The major contributor is the K+ leak channel: these channels allow small numbers of K+ ions to exit the cell leaving behind a negative charge across the membrane Nerve Cell o o o o Dendrites—cell protrusions that receive signals from axons Cell body – location of nucleus Axon – single long protrusion that send signals away from the cell body A nerve impulse results from electrical disturbances in the plasma membrane that spread from one part of the cell to another. The electrical disturbance is called an action potential and it consists of a wave of membrane depolarization that moves down the axon A threshold depolarization initiates the action potential o When a nerve receives a signal, a modest depolarization of the membrane occurs o If this depolarization reaches the threshold, all of the voltage-gated Na+ channels in that part of the cell will open simultaneously o Na+ rushes into the cell causing a rapid and large depolarization of the membrane o Red trace shows the change in membrane potential at a specific site on the axon. This “electrical disturbance” is the action potential. The green trace shows the hypothetical change that would occur if there were no voltage gated ion channels Voltage-gated channels are they key to an action potential o Depolarization to a specific membrane potential causes the channels to open o This specific membrane potential can be thought of as the threshold that must be reached to get the channel to open o The voltage-gated Na+ channel opens briefly but then automatically becomes inactive. It will remain inactive (and closed) until it is “reset” by the return to the original resting potential o Only after resetting, can the channel respond again to another threshold potential (refractory period) The return to the resting potential relies, in part, on inactivation of the voltage-gated Na+ channel o Repolarization: the cell’s efforts to return the charge across the membrane back towards the normal resting potential o Repolarization also depends on opening of voltage-gated K+ channels. These open more slowly than the voltage-gated Na+ channels so there is time for the influx of Na+ to depolarize the membrane before the efflux of K+ begins to repolarize the membrane The voltage-gated Na+ channel will not respond to another action potential until it has been “reset” by the re-information of a negative membrane potential Nerve Cell o Unidirectional—from dendrites to axon branches Transmission of a nervous impulse involves the propagation of an action potential across a neuron The cycle of (opened, closed, inactivated) channels along the axon results in unidirectional propagation of the action potential Nerve Cell to Nerve Cell communication o Once the signal has been transmitted along the length of the nerve cell, it needs to be sent to the next nerve cell o The plasm membrane of one nerve cell is not directly connected to the plasma membrane of the next nerve cell 0020