Transmembrane proteins span the bilayer. α-helix transmembrane domain. Multiple transmembrane helices in one polypeptide

Transcription

1 Transmembrane proteins span the bilayer α-helix transmembrane domain Hydrophobic R groups of a.a. interact with fatty acid chains Multiple transmembrane helices in one polypeptide Polar a.a. Hydrophilic pore Nonpolar a.a. Membrane transporter for polar or charged molecules ECB Fig Mobility of transmembrane proteins Bleach with laser beam If protein is mobile then fluorescent signal moves back into bleached area Recovery rate measures mobility 2

2 Peripheral membrane proteins (associated with membrane, but not in bilayer) Lecture 5 (cont d) Membrane Proteins Proteins as enzymes Binding sites Free energy Activation energy, enzyme function Enzyme mechanisms Kinetic parameters of enzymes Proteins as membrane transporters Enzymes bind substrates Substrate (ligand) Non-covalent interactions Binding site ECB Fig Enzyme (protein) 3

3 How do enzymes work? Start by considering free energy Free energy is amount of useful energy available to do work G (Delta G) = free energy change (Reactants - Products) In a chemical reaction G = H T S H = heat; heat released is negative S = entropy (randomness); increased randomness is positive Reactions occur spontaneously if G is negative Enzymes lower activation energy but have NO effect on G Energy of reactants Activation energy G Energy of products ECB Fig Uncatalyzed reaction Catalyzed reaction Enzymes accelerate reaction rates X Y Uncatalyzed reaction X Y Enzyme catalyzed reaction ECB Fig

4 How do enzymes accelerate reactions? Enzymes can hold substrates in positions that encourage reactions to occur Enzymes can change the ionic environment of substrates, accelerating the reaction Lower activation energy Enzymes can put physical stress on substrates Adapted from ECB Fig Thermodynamically Unfavorable Reactions ( G+) Many reactions in cells have positive G: e.g. condensation reactions (forming polymers reduces randomness so S -, G +) G = H T S Y G + Solution: couple to reaction where G - (Often hydrolysis of ATP) X Y G + ATP ADP + P i G - X + ATP Y + ADP + Pi + G - Example of coupled reaction: synthesis of sucrose ECB Panel 3-1 G values are additive 5

5 ATP (Nucleotide) G of hydrolysis = -7.3 kcal/mole ADP + P i + energy Enzymes can be regulated Inhibitors can bind to active site Binding in the active site can prevent substrate interaction Enzymes can be regulated at sites other than the active site Example: phosphorylation Fig ECB

6 Lecture 5 Outline Protein Secondary Structure Membrane Proteins Proteins as enzymes Proteins as membrane transporters (Ch 12 ECB) Channel Carrier proteins Facilitated diffusion Active transport Lipid Bilayer Permeability Small hydrophobic Molecules O 2, CO 2, N 2, benzene Properties of a pure synthetic lipid bilayer Small Uncharged polar molecules H 2 O, glycerol, ethanol Large, uncharged Polar molecules Amino acids, glucose, nucleotides IONS H +, Na +, HCO 3 -, K +, Ca 2+, Cl -, Mg 2+ ECB 12-2 Transmembrane proteins allow movement of molecules that cannot move through bilayer ECB 12-1 But it is not that simple 7

7 Membrane impermeability results in electrical and chemical gradients across membrane Charged molecules - transport influenced by concentration gradient and membrane potential (electrochemical (EC) gradient ) out Electrochemical gradient in ECB 12-8 Concentration gradient only Conc.. Gradient with membrane potential (-) inside Ion gradients across the plasma membrane ph 7.2* ph 7.4* Different electrochemical gradient for each ion Electrical and concentration gradient can be opposite (e.g. K + ) Transport problems faced by cells: - Need to get an impermeable molecule across the membrane - going WITH its electrochemical gradient - Need to get a molecule (permeable or impermeable) across the membrane going AGAINST its electrochemical gradient Solution -- specialized membrane proteins for transport functions. 8

8 Two broad classes of transmembrane proteins A. channel protein B. carrier proteins ECB 12-3 Conformational change Transport can be passive or active electrochemical ECB 12-4 Channels - Passive transport down elecrochemical gradient Impermeable Channel protein ECB 12-4 Channel-mediated diffusion (facilitated diffusion) 9

9 Channel structure Aqueous pore due to polar and charged R groups ECB Always passive transport Mechanism of K + channel selectivity ECB 12-7 Carrier mediated Diffusion (facilitated diffusion down EC gradient) Active transport (energy-driven) Carrier Proteins: Transport against EC gradient Transfer across membrane driven by conformational change in transporter Slower than channels Binds transported ligand - highly specific 10

10 Active transport - three types -uses energy to drive transport against EC gradient through carrier protein ECB 12-9 Coupled transport Down EC gradient Cotransported Molecule (against EC gradient) ECB Symport- - move same direction Antiport- - move opposite directions Na-Glucose symporter Move glucose against its EC gradient, using the energy stored in the Na + gradient. ECB

11 ATP-driven pumps Move against EC gradient ATP ADP + Pi Typically move ions generating EC gradient EC gradient can then be used in coupled transport Na + /K + pump in animal cells ECB Cyclic transport by Na + /K + pump Phosphoryation regulates the enzyme conformation 3 Conf. change 1 Low affinity Na binding sites High affinity K binding sites 3 High affinity Na binding sites Low affinity K+ binding sites 3 2 NaKATPase.avi 2 Conf. change

12 Chemiosmotic coupling of pumps and cotransport H + transporters in vacuole and lysosome are similar Osmosis Osmosis: movement of water from region of low solute concentration to region of high solute concentration (or high water potential to low water potential) How do cells prevent osmotic swelling? ECB