Membrane Transport of Small Molecules and Electrical Properties of Membranes (Chapter 11) PDF

Membrane Transport of Small Molecules and the Electrical Properties of Membranes (Chapter 11) Not for sale or distribution; property of York University © Garland Science  What types of molecules are transported within an eukaryotic cell?...

Membrane Transport of Small Molecules and the Electrical Properties of Membranes (Chapter 11) Not for sale or distribution; property of York University © Garland Science  What types of molecules are transported within an eukaryotic cell? Nucleus Mitochondrion Chloroplast Lysosomes Not for sale or distribution; property of York University Transport © 2010 Nature Education All rights reserved.  Permeability of protein-free lipid bilayers Membrane permeability is inversely proportional to size, polarity and charge ex: water permeates the membrane 109x faster than small ions! Figure 11-1 Molecular Biology of the Cell (© Garland Science)  Solutions to Transport Problems Depends on size and hydrophobicity SIZE of MOLECULE TRANSPORT MECHANISM Ions, small molecules (e.g. sugars) Transport proteins (chapter 11) Macromolecules (e.g. protein, Protein translocators, nuclear RNA) pores (chapter 12) Macromolecules, large particles Vesicles, endocytosis, (e.g. foreign particles) phagocytosis (chapter 13)  Ion concentrations inside and outside a typical mammalian cell Besides Cl-, there are many other negative charges: HCO3-, PO43-, metabolites, nucleic acids, etc Shown are concentrations of free cytosolic ions (not bound to other molecules and not inside organelles) Table 11-1 Molecular Biology of the Cell (© Garland Science 2008) Therefore, the cell needs mechanisms to allow specific molecules to cross the membrane barrier: membrane transport proteins Membrane transport proteins are specific for one or a few types of molecules All membrane transport proteins are multi-pass integral membrane proteins There are 2 types: Channels and Carriers/permeases Classes of protein transporters in biological membranes Membrane Transport proteins Carriers/permeases Channels passive active What are we talking about? Principles of membrane transport Different classes of membrane transport proteins Passive vs Active Transport Common types of transporters Channel structure and function Electrical properties of membrane Fun Fact: In some mammalian cells (nerves and kidney), two-thirds of their total metabolic energy is consumed in membrane transport. Diffusion across the lipid bilayer depends on size and hydrophobicity. How exactly does urea or sucrose get across? Membrane Transport Proteins are multipass Transmembrane Proteins Create a hydrophilic pathway through the hydrophobic core Two Main Classes of Membrane Transport Proteins Membrane protein that mediates the passage of ions or molecules across a membrane.  There Are Two Main Classes of Membrane Transport Proteins: Transporters and Channels a) Transporters (also called carriers or permeases) b) Channels Figure 11-3 Transporters move solute by conformational change Transporters have binding sites specific to solute. Conformational changes required for transport (expose solute to other side of membrane) Figure 11-3 Channels form a pore for solute movement without conformational change Pore through bilayer Most open/close (shape matters) Interact minimally with solute Most Specific (selectivity filter) Fast Where does the energy for transport come from? All Channels and some Transporters use diffusion to power transport passive transport - solute movement down the concentration gradient Facilitated passive transport Figure 11-4  Concentration gradients have potential energy. Diffusion is spontaneous, exergonic, increasing entropy proceeds to dynamic equilibrium Diffusion is driven by an Increase in Entropy ΔG = ΔH − TΔS Ion Gradients are critical to cell function What do these things have in common? Lethal injection Thousands of bananas Image Credit: Mika Baumeister Image Credit: Rodrigo dos Reis  Hyperkalemia too much potassium in the bloodstream  Membrane Potential Membrane Potential or Potential Difference refers to the difference between the inside charge and outside charge. OUTSIDE Ca ++ Cl - K+ Na + Difference is a negative value (-20mv to K+ -150mv) Ca++ Na+ Cl- INSIDE Movement of charged solutes (ions) depends on concentration gradient and electrical gradient + + + + + + + + + + Fig 11.4 Movement of charged solutes (ions) depends on the electrochemical gradient Movement of ions is influenced by: a) Concentration gradient b) Electrical potential or voltage across the membrane (membrane potential) The combined net driving force is the electrochemical gradient Electrochemical gradients are used to do work in the cell Cells use electrochemical gradients for a) Transport a) Signalling (action potentials) b) ATP production (bacteria, mitochondria, chloroplasts) c) Other functions (eg opening and closing stomata) Electrical potential across animal cell membrane is -20mv to -120mv (more negative charge inside cell) Transporters can use passive or active transport Conformational change Movement of solute through a transporter involves conformational change Passive transporter transitions Random So how does solute move Reversible from high to low Do not require solute concentration? Transporters that transport 1 solute = uniporter Figure 11-5  Question During passive transport, how can the random flipping of the transport protein between two conformations result in directional transport? Direction in passive transport is determined by the solute concentration gradient If the solute concentration is higher on the outside of the membrane, more solute will bind to the transporter in the outward-open state/conformation. This will result in a net transport down its concentration gradient. Figure 11-5  In what way are passive transporters NOT like enzymes? GLUT Transporters are a family of passive transporters for glucose encoded by the SLC2 genes GLUT 1 – red blood cells and other tissue GLUT 2 – liver, gut epithelium 12 membrane-spanning domains major glucose transporter To move solute against its concentration gradient you need energy active transport moves solute against the concentration or electrochemical gradient and requires energy. Figure 11-4 Energy for active transport can come from different sources. Energy powers conformational change Ion Gradient ATP Light Redox Reax Figure 11-7 Active: Coupled transporters use the energy stored in ion concentration gradients Couple the uphill transport of one solute across the membrane to the downhill transport of another solute (ion) e.g. identified in bacteria, plants, animals Figure 11-7 Coupled Transporters may be symporters or antiporters Uniporters: 1 solute transported Coupled: 2 solutes transported, in same or different directions Also called secondary active transport high low Figure 11-8 Transporters Cotransport: Transport of 2 different ligands in the same direction Transporters Countertransport: Transport of 2 different ligands in the opposite direction Active Transport Can Be Driven by Ion-Concentration Gradients The Na+-glucose cotransporter (symporter) moves glucose against its concentration gradient using the Na+ gradient  The plasma membrane Na+-glucose symporter The Na+-glucose symporter is found in the plasma membrane of epithelial cells in kidney and intestines. Its function is to recover glucose from extracellular medium before excretion However, [Glucose]cytosol >> [Glucose]extracellular To transport glucose against this concentration gradient, cells use the strong electrochemical gradient of Na+: [Na+]extracellular >> [Na+]cytosol An Asymmetric Distribution of Transporters in Epithelial Cells Underlies the Transcellular Transport of Solutes Gut epithelial cells transport glucose from lumen to extracellular space. An Asymmetric Distribution of Transporters in Epithelial Cells Underlies the Transcellular Transport of Solutes Gut epithelial cells transport glucose from lumen to extracellular space. How might the cell create and maintain different transporter distributions in different areas? ATP-Transporters ATP-driven transporters (pumps) couple the transport of the solute ( ) against its conc. gradient to the hydrolysis of ATP Figure 11-7 There Are Three Classes of ATP-Driven Pumps (Transport ATPases, ATP Transporters) 1. P-type 2. ABC 3. V type P-type pump: transporters that self-phosphorylate P-type pumps typically setup and maintain ion gradients across membranes maintain low Ca2+ levels in cytoplasm/cytosol create Na+/K+ gradients (e.g. in animal cells) create H+ gradients (e.g. in plants and fungi) F-type H+ pumps: these are multi-subunit, turbine-like complexes found in bacteria, mitochondria and chloroplasts. use H+ gradients to synthesize ATP. V-type H+ pumps: related to F-type but use ATP hydrolysis to pump H+ against electro-concentration gradient. acidifies organelles. ABC Transporter: Uses ATP hydrolysis to pump small molecules across the membrane. Does not get phosphorylated P-type pumps are phosphorylated by ATP during the pumping cycle. Phosphorylation/dephosphorylation drives conformational change to move ions against their concentration gradient Figure 11-12 P-type pumps maintain important ion gradients across membrane: primary transport There Are Three Classes of ATP-Driven Pumps (Transport ATPases, ATP Transporters) 1. P-type 2. ABC 3. V type ABC (ATP-Binding Cassette) transporters pump small molecules across the cell membranes. ABC transporters constitute the largest family of membrane transporters. Figure 11-12 ABC Transporters Constitute the Largest Family of Membrane Transport Proteins ABC transporters contain two highly conserved ATPase domains or ATP-binding cassettes on the cytoplasmic side e.g. import of nutrients Figure 11-16 F-type ATPase (ATP synthase) is structurally related to V-type proton pumps. use the H+ gradient across the membrane to synthesize ATP. e.g. ATP synthase in chloroplasts. Figure 11-12 V-type pumps are turbine-like protein machines. Transfer H+ into organelles such as lysosomes, vesicles or vacuoles to acidify in their interior. Figure 11-12  Cystic fibrosis gradual deterioration of gastrointestinal + reproductive tract, lungs basis abnormal form of cystic fibrosis transmembrane conductance regulator CFTR protein: Cl- channel protein (encoded on chromosome 7) most prevalent CFTR mutation → deletion of 3 nucleotides (Phe508) sweat duct cells are impermeable to Cl- consequently, Na+ remains in the sweat duct  Cystic fibrosis > 600 mutations in the gene located in the NBD1  Membrane spanning Membrane spanning Cystic fibrosis domain 1 domain 2 transmembrane conductance regulator protein (CFTR) Consists of 5 domains 2 domains that 2 domains form the channel bind/use ATP across the ATP1 cytoplasmic ATP2 membrane Cystic fibrosis transmembrane conductance regulator protein (CFTR) Consists of 5 domains Nucleotide 2 domains binding domain 2 bind/use ATP Nucleotide ATP1 binding domain 1 ATP2 Channel Proteins and the Electrical Properties of Membranes Movement through channels is fast. Water channels pass 10 billion water molecules/s Ion channels pass up to 100 million ions/s. This is 100,000 X greater than the fastest rate of transport by a transporter. Passive Transport: Ions move through channels down their electrochemical gradient. Some channel proteins are non-selective, but most are highly selective Non-Selective Gap Junctions Porins (bacteria, mito, chloro) Some ion channels Selective Aquaporins Most ion channels Some cells are very sensitive to hypotonic environments and some cells aren’t http://www.xenopus.com/products.htm Aquaporins Are Permeable to Water But Impermeable to Ions Water channels - 10 billion water molecules/s Narrow pore lined with carbonyl groups from peptide backbone which H bond to water as it moves through Aquaporins Are Permeable to Water But Impermeable to Ions Carbonyl oxygens in a peptide chain peptide chain backbone Aquaporins Are Permeable to Water But Impermeable to Ions Preventing a H+ relay: two Asn residues in the center of the pore tether the oxygen of the water molecule to prevent a H+ relay through the pore. Preventing other ions: pore size and hydrophobic side Figure 11-20  Ion channels form a narrow aqueous pore Passage is highly selective for a single ion type Selection accomplished by a selectivity filter found in the pore Aqueous pore can be closed or open Movement is from high to low electrochemical gradient never the other way Rate of ion flow can be 105 times faster than any known transporter Molecular Biology of the Cell (© Garland Science) Ion Channels Are Ion-Selective and Most Fluctuate Between Open and Closed States. Properties of Ion channels: 1) Selectivity 2) Gating 3) Desensitization/inactivation Figure 11-21 Ion selectivity filter is the narrowest part of the channel Ions normally pass through the channel in single file. The selectivity filter controls which ions can pass through. Figure 11-21  Summary 1. The membrane is permeability barrier for many molecules but possesses mechanisms to allow exchange across it. Diffusion across the lipid bilayer depends on size and hydrophobicity. 2. Small molecules cross the membrane by using membrane transporters and ion channels. 3. Membrane transporters permit passive or active transport. Active transport can be coupled to concentration gradient, ATP hydrolysis and photon capture 4. ATP transporters consist of: P-type, F-type and ABC families. 5. Ion channels only allow passive transport of ions by forming an aqueous pore selective for one or several types of ions