Membrane Transport Chapter 20 January 10 Lecture 2 1

Important Thermodynamic Concepts 2 Standard State Convention in Biochemistry Biochemical reactions typically occur under different conditions than what are assigned as ‘Standard’ by thermodynamic convention. ~neutral pH dilute aqueous conditions A different Standard State Convention has been adopted for Biochemistry Water’s standard state  defined as pure water so NOT incorporated into the equilibrium term H + ion activity defined as 1 at pH 7 (instead of pH 0) Standard Free Energies are only valid at pH 7 Symbolized by  G 0’ instead of  G 0 If a biochemical reaction does NOT include H 2 O or H +,  G 0 =  G 0’

Important Thermodynamic Concepts 3 Standard State Convention in Biochemistry If a biochemical reaction includes H 2 O If a biochemical reaction includes H +

Important Thermodynamic Concepts 4 Basic Equations

Important Thermodynamic Concepts 5 Coupled Chemical Reactions Hess’ Law

Important Thermodynamic Concepts 6 LactatePyruvate How much energy does the reduction of NADH need to provide to make this a spontaneous reaction?

Thermodynamics of Phosphate Compounds 7 Note that ATP hydrolysis lies in the middle of the table – this enables the central role that ATP plays in metabolic pathways  G is highly dependent on metabolite concentrations. An estimation of -50 kJ mol -1 has been made for the hydrolysis of ATP.

Thermodynamics of Phosphate Compounds 8 High Energy Phosphoanhydride Bonds (~) AR-P~P~P Do not confuse High Energy Bond with High Bond Energy High Energy Bond - The energy required to hydrolyze a bond High Bond Energy – The energy required to break a bond

Thermodynamics of Phosphate Compounds 9 What makes Phosphoanhydride bonds high energy? Stability of reactants vs. products Electrostatic repulsion of phosphoanhydride Solvation Energy? Challenging to estimate

Role of ATP 10 “Energy Conduit” – ATP is a general intermediate in energy transfer from really high energy compounds to lower energy phosphate compounds Biological systems are able to evolve such that multiple enzymes utilize this intermediate Enzymes can easily adopt an ATP-binding fold and then evolve to bind another substrate One very common ATP-binding motif is the Walker-A Motif

ATP Consumption 11 Early Stages of Catabolic Pathways Formation of CTP, GTP, UTP and TTP Physiological Processes Muscle Contraction

ATP Formation 12 Substrate Level Phosphorylation by Kinases Oxidative Phosphorylation and Photophosphorylation Large amounts of ATP are generated in these electron transport chain reactions Adenylate kinase reaction ATP is then generated through substrate level phosphorylation

Phosphocreatine as an Energy Reservior 13 Under physiological conditions, [ATP] >> [ADP] – how would this influence equilibrium and  G? The creatine/phosphocreatine system generates an ATP “Buffer” that can store ATP energy for times of need. ATP can be generated from phosphocreatine within 5 seconds of a muscle burst!

Oxidation-Reduction Reactions 14 Why do we care? Aerobic Respiration C 6 H 12 O 6 + O 2  CO 2 + H 2 O Anaerobic Respiration – Fermentation (the cool one!) C 6 H 12 O 6  CO 2 + CH 3 CH 2 OH

Oxidation-Reduction Reactions 15 Assuming no PV work is done Difference in electric potential Number of electrons flowing in balanced equation Faraday’s Constant J V -1 mol -1

Oxidation-Reduction Reactions 16

Oxidation-Reduction Reactions 17 Is this reaction spontaneous under standard conditions? CytC(Fe 3+ ) + CytA 3 (Fe 2+ )  CytC(Fe 2+ ) + CytA 3 (Fe 3+ ) If the concentration of all other reaction components is buffered at 50  M, how much CytC(Fe 3+ ) needs to be added to make the reaction spontaneous?

Oxidation-Reduction Concentration Gradients 18 Energy Needed Energy Produced Examples Proton gradients drive ATP synthesis Na+ and K+ gradients drive electrical nerve impulses

Thermodynamics of Membrane Transport 19 A (out)  A (in) Chemical Potential (for neutral species) Membrane Potential created by a difference in charge across a membrane Electrochemical Potential (for charged species)

Kinetics of Membrane Diffusion 20 A (out)  A (in) Nonmediated – Occurs independent of protein based pores Flux – Rate of passage per unit area Permeability coefficient – Ability of a solute to transfer from solvent to membrane core

Kinetics of Membrane Diffusion 21 Nonmediated – Occurs independent of protein based pores

Kinetics of Membrane Diffusion 22 A (out)  A (in) Nonmediated – Occurs independent of protein based pores Mediated – Requires a protein based channel to facilitate membrane transport Passive – Flow to minimize concentration gradient

Kinetics of Membrane Diffusion 23 A (out)  A (in) Nonmediated – Occurs independent of protein based pores Mediated – Requires a protein based channel to facilitate membrane transport Passive – Flow to minimize concentration gradient

Kinetics of Membrane Diffusion 24

Kinetics of Membrane Diffusion 25 A (out)  A (in) Nonmediated – Occurs independent of protein based pores Mediated – Requires a protein based channel to facilitate membrane transport Passive – Flow to minimize concentration gradient Active – Transport opposes concentration gradients and requires energy

Mediated Transport 26 Uniport – movement of a single solute Symport – movement of two different solutes simultaneously in the same direction Antiport – movement of two different solutes simultaneously in opposing directions Example: Glucose Transporters Example: (Na + -K + )-ATPase 3Na + (in) + 2K + (out) + ATP + H 2 O  3Na + (out) + 2K + (in) + ADP + P i Example: Lactose Permease Mediated by ATP-hydrolysis Active Transport (Energy Dependent)

Structure Example: Potassium Channel 27 Important for: Osmotic Balance Signal Transduction Nerve Impulses ~10 8 ions per second! VERY selective for K + over Na + Na + K+K+ Ionic Radius (Å) Hydration Energy (kJ mol -1 ) Selectivity Filter TVGYG

Structure Example: Potassium Channel 28 Electrostatic repulsion of 4 K+ ions in such close proximity suggests that this structure is the superposition of 2 structures

Structure Example: Potassium Channel 29 Closed formOpen form Twisting of 4 subunits More sophisticate mechanisms such as a ‘swinging protein ball’