Biological Membranes and Transport

Biological membranes and transport Objectives: - Recognize the basic structural elements, physical properties and molecular constituents of biological membranes. - Understand different types of membrane movements - Know different types of proteins present at biological membranes - Understand the mechanisms of solute transport across the membrane Physical properties of Biological membranes self sealing, flexibility, selectivity The molecular constituents of Biological membranes -different compositions of lipid membranes - micelle, lipid bilyar, liposome - Membrane proteins Integral proteins, peripheral proteins Solute transport across the membrane - Passive transport - Facilitated diffusion - Active transport Different Examples for solute transport across the biological membranes. - Aquaporin - Glucose transport - Chloride-Bicarbonate transport - Sod-Pot transport Ion selective channels Ionophores Biological membranes and transport Membrane: - Define the external boundaries and regulate the molecular traffic across that these boundaries. - In eukaryotic cells, they can divide the internal spaces into compartments to segregate processes and components - They organize complex reaction sequences, They are important in cell- cell communication -within the cell, membranes organize cellular processes as energy transduction in mitochondria and chloroplast Physical properties of membranes 1- flexibility: allows the shape changes in the cell growth and movements (as the amoeboid movement) 2- self sealing: ability to break and to reseal without the leak of the cellular components, as in the case of exocytosis (two membranes can fuse) or endocytosis (one single compartment divide to give two sealed compartments) 3- selectively permeable, they can retain certain compounds and ions within cells and within specific cellular compartments while excluding others *Membranes are not absolutely passive barriers They include different proteins specialized for catalyzing various cellular processes A lipid bilayer is the basic structural element of membranes Structural lipids are insoluble in water ➔ when they are mixed with water, they will form microscopic lipids aggregates in a phase separate from the aqueous surrounding. Depending on the precise conditions and the nature of lipids three types of lipid aggregate are formed Micelle, Bilayer, Liposome Micelle: spherical structure, the molecules are arranged with hydrophobic regions aggregated in the interior, excluding the water and the hydrophilic head groups at the surface in contact with water Micelle formation is favored when the cross-sectional area of the head group is greater than that of the acyl side chain, as free fatty acids, lysophospholipids, detergents Single tailed lipids tend to form micelles Bilayer Two lipid monolayers form a two-dimensional sheet, bilayer formation occurs readily when the cross-sectional areas of the head group and the acyl side chain are similar, as in glycerphosphlipids and sphingolipids, the hydrophobic portions in each monolayer, excluded from water, interact with each other. The hydrophilic head groups interact with water at each surface of the bilayer. The two hydrocarbon tails of glycerophospholipids and spingolipids tend to form bilayers or liposomes Liposome Because the hydrophobic regions at the edges of the bilayer are in contact with water, the bilayer sheet is relatively unstable and spontaneously fold back on itself to form a hollow sphere called liposome. These vesicles enclose water creating a a separate water compartment *membrane lipids are asymmetrically distributed between the monolayers of the bilayer Mobility of Lipid Molecules • The lipid bilayer itself is stable but individual molecules have a great freedom of motion • Rarely will a lipid molecule transfer across a bilayer in a process termed transverse diffusion or flip – flop • However, lipids are very mobile in the plane of the bilayer and move in a process called lateral diffusion this randomizes the positions of the lipid molecules in few seconds • The interior of a bilayer is in constant motion Temperature Dependence of the Fluidity of a Bilayer • The interior of the bilayer is fluid because the hydrocarbon chain of fatty acid are in constant motion produced by rotation about C-C bonds of the acyl chains. The degree of fluidity depend on the lipid composition and temperature: • At low temp.➔ bilayer becomes a gel -like solid Paracrystalline state • at high temp. ➔ lipids undergo rapid motion and the bilayer becomes more fluid • The transition temperature : the temp above which the paracrstalline changes into fluid and it is characteristic to the membrane and depend on the lipid composition • The transition temperature is dependant on chain length and the degree of saturation of its component fatty residues • Cholesterol broadens the temperature range of the phase transition:The rigid structure of the sterol nucleus has two effect on the fluidity of the membrane -Below the Trans. Temp -Above the Trans. Tem Sterols moderate the extremes of solidity and fluidity of membranes The Molecular constituents of membranes • Lipids and proteins and some carbohydrates as part of glycoprotein and glycolipids. • The relative proportions of protein and lipids vary with the type membrane➔ reflecting the diversity of biological role. e.g. Plasma membrane of mitochondria contains more protein than lipid; many enzyme-catalyzed processes take place there. while Mylelin sheath = plasma membrane of neurons➔ act as electrical insulator ➔ contains high % of lipids. * The lipid and protein composition of membranes from different sources vary reflecting the biological activity. The supramolecular architecture of membranes Common properties • Biological membranes are impermeable to most polar or charged solutes but permeable to non-polar compounds • 5-8 nm thickness -fluid mosaic model for the structure of biological membranes • Phospholipids and sterols form a lipid bilayer in which the non-polar regions face each other at the core of the bilayer and the polar head groups face outward. • The proteins are embedded at irregular intervals fixed with hydrophobic interactions between lipids and hydrophobic domains in the proteins • lipids and protein form a fluid mosaic, because most of their interactions are not covalent this leave the lipid and protein molecules move freely laterally in the membrane • The orientation of the protein is asymmetric; the proteins exposed at one side are different from those at the other side • Some proteins present at on side, others have domains exposed at the two sides Membrane Structure & Assembly Fluid Mosaic Model • Lipids and proteins can undergo rotational and lateral movement • 30 - 90% of membrane proteins are freely mobile • Membranes are asymmetric * Lateral Asymmetry – Proteins can associate and cluster in the plane of the membrane they are not uniformly distributed in many cases – Lipids can cluster in the plane of the membrane - they are not uniformly distributed * Transverse asymmetry – N-terminus is often extracellular whereas C-terminus is intracellular – In most cell membranes, the composition of the outer monolayer is quite different from that of the inner monolayer Membranes are full of proteins - Can be 20-80% protein (dry wt.) - In eukaryotes, different membranes differ in their protein composition - Proteins can be associated with membranes with different ways Association of proteins with membranes - Integral (intrinsic): = proteins that pass through membrane, largely buried within the membrane and can be exposed on both membrane faces, they are tightly bound to membranes by hydrophobic interactions, removable only by agents that interfere with hydrophobic interactions, as detergents, organic solvents or denaturants. Detergents can form a micelle like structure around the protein molecule Integral membrane proteins • Attachment of an integral protein to a membrane requires protein has one or more hydrophobic domains that can span the lipid bilayer. Most often, these are a-helical domains. • Integral proteins are amphiphlies • Oligosaccharides can be linked covalently to the integral proteins at the outer face; out side the membrane • The protein segment inserted into the membrane can vary in structure. • Serve as transporters, ion channels, receptors and play important role in cell fusion , cell-cell recognition Stretch of the protein most likely to be EXTRACYTOPLASMIC Association of proteins with membranes - Peripheral (extrinsic) associated with membrane through electrostatic interactions and hydrogen bonding with the polar head groups of membrane lipid or with hydrophilic domains in the integral proteins, they can be released by relatively mild treatments that interfere with electrostatic interactions or break H-bonds (eg. changes in ionic strength or pH, removal of Ca2+ by chelator, urea) They serve as regulators of membrane bound enzyme or may limit the mobility of integral proteins -linked-Lipid: some proteins released by treatment with PIspecific phospho-lipase C: cleaves a “glycosyl phosphatidyl-inositol” membrane anchor attached at COOH terminus of some proteins PROTEIN I C=O NH CH2 CH2 P sugars P inositol PL-C Lipid-anchored proteins Thioester-linked 16:0 palmitate - Note sidedness Amide-linked 14:0 myristate (at N-terminus) - The lipid anchors are added to a protein after a protein has been made - Always attached to a C-terminal residue - Examples: surface antigens, adhesion molecules Thioether-linked 15 or 20 carbon isoprenoid on C-terminal Cys GlcN cytoplasm Extra cytoplasmic Amide link between ethanolamine N and C-terminal aa of protein Membrane protein movements -Many proteins can are free to diffuse laterally in the plane of the bilayer and are in constant motion *Fusion of a mouse cell with a human cell result in the randomization of the membrane proteins from the two cells -Some membrane proteins associate together to form large aggregate➔ can't move -Other protein are linked to to internal structures that prevent their free diffusion Membrane fusion Biological membrane is able to undergo fusion with another membrane without losing its integrity. Exocytosis, endocytosis, cell division, fusion of the egg and sperm cells and the entry of many viruses into the host cells, all involve the fusion of two membranes without loss of continuity. Specific fusion requires that 1- the two membranes recognize each other 2- their surfaces become closely opposed, removal of the water associated with the polar head groups. 3- Their bilayer structures become locally disrupted and the two bilayers fuse to form a single continuous bilayer Solute transport across Membranes - Cells must i) import raw materials for biosynthesis and energy production ii) export by-products of metabolism - Non-polar gases such as O2, hydrophobic molecules such as steroid hormones, and small, non-polar molecules can enter and leave a cell by diffusing through the membrane. -The hydrophobic core of a lipid bilayer is a barrier to polar and charged molecules -Import/export occurs across permeable membranes and mediated by proteins as transmembrane channels, carriers, pumps • The diffusion of a substance between two sides of a membrane thermodynamically resembles a chemical equilibration. • The difference in the concentration of a substance on two sides of a membrane generates a chemical potential difference • The movement of ions across a membrane also results in a charge difference generating an electrical potential difference Transport may be mediated or non-mediated Non-mediated transport occurs through simple diffusion where the rate of diffusion depends on the concentration gradient and the substances solubility in the membrane’s non-polar core – O2, N2 , CO2 and methane and steroids diffuse readily Mediated transport is classified into two categories depending on the thermodynamics of the system 1. Passive –mediated transport or facilitated diffusion. Molecules flow from high concentrations to low concentrations 2. Active transport. A molecule is transported against its concentration gradient. This endergonic process must be coupled with an exergonic process to make it thermodynamically favorable. *Passive transport Simple diffusion A solute moves from a region of high concentration to one of low concentration by simple diffusion -The difference in the solute concentration across the membrane is called chemical gradient - When ions of opposite charge are separated by a permeable membrane, there is a transmembrane electrical gradient, = membrane potential (in volts or millivolts) -Membrane potential produces a force that drives ion movements to make the potential equal zero -The direction in which a charged solute tends to move spontaneously across a membrane depends on both the chemical gradient (= difference in [solute]) and the electrical gradient across the membrane -Together, these two factors are referred to as the electrochemical gradient or electrochemical potential. Membranes as permeability barriers Simple Diffusion - Non-polar cores of bilayers make them very impermeable to most ionic and polar substances and proteins (water molecules traverse membranes slowly). - A small molecule/ion must first shed its hydration shell of water, then become dissolved in the hydrocarbon core of the membrane, then diffuse through the core, and become rehydrated on the other side. - Although the energy used to strip the solute of its hydration shell is regained as the compound is rehydrated on the other side of the membrane i.e. the consumed energy in the passive diffusion is is zero, the need to remove a coordination shell of water molecules creates a very high activation energy (DG‡ ) Facilitated diffusion - Transmembrane passage of polar compounds is made possible by proteins that lower the activation energy (by providing an alternative path for specific solutes through the bilayer) - Membrane proteins that speed the movement of a solute across the membrane by facilitating diffusion is called transporters or permeases . - Transporters bind substrates specifically through many weak, non-covalent interactions, replacing water. - The -ve free energy change associated with these interactions (DGbinding) counterbalances the +ve free energy change for dehydration of the solute (DGdehydration), and lowers DG‡ for transmembrane passage. Non-mediated and mediated/facilitated transport 1. Passive diffusion 2. Carrier-mediated transport Not saturable Rate increases w. concentration difference across membrane Saturable (like enzyme) i) No energy required: “facilitated diffusion” or “passive transport” ii) Energy required: “active transport” Three General classes of transport system - Transporters differ in the number of solutes transported and in the direction in which each is transported - This classification, however, tells us nothing about whether these are energyrequiring or energy-independent processes. Facilitated Diffusion Examples 1)Aquaporins, 2) Glucose transport, 3) chloride and bicarbonate Aquaporins form hydrophilic transmembrane channels for the passage of water Aquaporins (AQPs): a family of integral proteins that provide channels for rapid movement of water molecule across plasma membranes in a variety of specialized tissue.membranes of erythrocytes and proximal renal tubule cells are rich in aquaporins Aquaporins are type III integral proteins with six transmembrane helical segments, four monomers form tetrameric transmembrane channel lined with hydrophilic side chains and having a diameter of 3 A to allow the passage of water molecules AQPs reduce the activation energy for the passage of water ➔ water moves in the channels in a continuous stream Example: Glucose transporter of Erythrocyte mediates passive transport of glucose (GluT1) Glucose enters the erythrocyte by facilitated diffusion via specific glucose transporter. Glucose transporter is integral protein and exists in two conformations T1, with the glucosebinding site exposed on the outer surface of the plasma membrane and T2 with the binding site at the inner surface T in conformation 1: Glc-binding site exposed on outer surface of membrane 1. Glc binds to specific site, lowering activation energy for…. 2. Conformational change from Sout.T1 to Sin.T2, which “brings about” passage of Glc through the membrane 3. Glc released from T2 into cytosol 4. T2 returns to T1 conformation Note: transport process is fully reversible: Glucose can’t be accumulated in cells at Concentrations higher than in the medium. Transporter achieves equilibration of Glucose at a much higher rate Example: Chloride and bicarbonate are co-transported across the erythrocyte membrane The chloride-bicarbonate exchanger(anion exchange protein) is an integral protein that mediate mediates the simultaneous movement of two anions: for each HCO3 ion that moves in one direction, one Cl ion moves in the opposite direction. The coupling of Cl and HCO3 movement is obligatory, the absence of Cl, the transport of bicarbonate stops, Active transport - Results in accumulation of a solute above the equilibrium point because of the movement of the molecules against an electrochemical gradient -Thermodynamically unfavorable (endergonic process): occurs only when coupled (directly or indirectly) to an energy-yielding (exergonic) process. Primary active transport: solute accumulation is coupled directly to the exergonic chemical reaction, like the breakdown of ATP, oxidation reaction, absorption of sunlight. Secondary active transport: solute accumulation is coupled indirectly to exergonic reaction as concomitant flow of another chemical species down its electrochemicalgradient - Ion gradient X (often Na+) is established by 1° active transport - Movement of ion X down its concentration gradient provides the energy to drive co-transport of a second solute(s) against its electrochemical gradient Secondary active transport Secondary active transport: solute accumulation is coupled indirectly to exergonic reaction as associated with flow of another chemical species down its electrochemical gradient x-inside X-outside ATP ADP +Pi X-outside x-inside s-outside S-inside ============================= s-outside + ATP S-inside + ADP +Pi Active transport: examples Primary active transport example: Na+ K+ ATPase The energy released by ATP hydrolysis drives solute movement against its electrochemical gradient. There are several types of ATP-dependent active transporters, that called transport ATPases which are differing in the structure, mechanism, and localization in specific tissues and intracellular compartments - Responsible for setting and maintaining intracellular [Na+] and [K+] in animal cells - Internal [Na+] < external [Na+] and internal [K+] > external [K+] = 2-subunit integral membrane protein in plasma membrane - Antiport - three positive charges exit for every two that enter Na+ K+ ATPase - For each ATP converted to ADP + Pi, transporter moves 2 K+ into cell and 3 Na+ out - Process creates a net separation of charge across membrane: = transmembrane potential (-50 to -70 mV, inside -ve relative to outside) - Current model: ATP-dependent phosphorylation of the transporter changes its conformation: transporter cycles between: a phosphorylated form with high affinity for for K+ and low affinity for Na+, and a dephosphorylated form with high affinity for Na+ and low affinity for K+. Secondary active transport example: Accumulation of glucose in the intestinal cell Glucose and some amino acids can be accumulated in the cell by the symport with Na +, using the Na + gradient established by the Na + -K+ ATPase of the plasma membrane Na + -glucose symporters take up the glucose from the intestine in a process driven by the down gradient flow of the Na+ *glucose is co-transported with Na+ ions across the plasma membrane of the intestinal cells. It passes into the blood via (GluT2)passive glucose transporter. The Na + -K + ATPase continues to pump Na + outward to maintain the gradient of Na + that drives the glucose uptake Passive Transport: Ion selective channels Passive transport 1- Simple diffusion 2- Facilitated Diffusion 3- Ion selective channels 4- Ionophores Ion selective channels Ion selective channels are another mechanism for moving inorganic ions across membranes, together with Na+-K+ pumps they regulate the cystolic concentration of ions and the membrane potential Ion channels are distinguished from transporters in the following 1- Rate of flux through ion channels is much greater than that of transporter 2- Ion channels are not saturable 3- They are “ gated” can be opened or closed in response to some cellular event. Ligandgated channels ; the binding of some molecules forces changes in the protein lead to open or close the channel. Voltage-gated ion channels ; change in the transmembrane electrical potential causing the ion channel to open or close The most popular channels K +, and Na+ channels that present in neurons, heart cells, skeletal muscles * Role of Na+-K+ channels and Na+-K+ pumps in the signal transmission of the neuron ??? Passive Transport: Ionophores Ionophores • An ionophore is an organic molecule that binds an ion facilitating the ions diffusion through the membrane • Ionophores are often antibiotics of bacterial origin. • Ionophores can act as carriers, valinonycin, or as channel formers, gramicidin A • Ionophores can only dissipate an ion gradient