CHAPTER 3 Bioenergetics Enzymes and Metabolism 3 1
CHAPTER 3 Bioenergetics, Enzymes, and Metabolism
3. 1 Bioenergetics • The study of the various types of energy transformations that occur in living organisms.
The Laws of Thermodynamics and the Concept of Entropy • Energy – capacity to do work, or the capacity to change or move something. • Thermodynamics – the study of the changes in energy that accompany events in the universe.
The First Law of Thermodynamics (1) • The first law of thermodynamics – the law of conservation of energy. • Energy can neither be created nor destroyed. • Transduction – conversion of energy from one form to another. • Electric energy can be transduced to mechanical energy when we plug in a clock.
The First Law of Thermodynamics (2) • Cells are capable of energy transduction. • Chemical energy is stored in certain biological molecules, such as ATP. • Chemical energy is converted to mechanical energy when heat is released during muscle contraction.
The First Law of Thermodynamics (3) • Energy transduction in the biological world: conversion is the conversion of sunlight into chemical energy – photosynthesis. • Animals, such as fireflies and luminous fish, are able to convert chemical energy back to light.
The First Law of Thermodynamics (4) • The universe can be divided into system and surroundings. – The system is a subset of the universe under study. – The surroundings are everything that is not part of the system. – The energy of the system is called the internal energy (E), and its change during a transformation is called ΔE.
The First Law of Thermodynamics (5) • The first law of thermodynamics: ΔE = Q – W, where Q is the heat energy and W is the work energy. • When there is energy transduction (ΔE) in a system, heat content may increase or decrease. – Reactions that lose heat are exothermic. – Reactions that gain heat are endothermic. – The first law does not predict whether an energy change will be positive or negative.
The First Law of Thermodynamics (6)
The Second Law of Thermodynamics (1) • The second law of thermodynamics: events in the universe tend to proceed from a state of higher energy to a state of lower energy. – Such events are called spontaneous, they can occur without the input of external energy. – Loss of available energy during a process is the result of a tendency for randomness to increase whenever there is a transfer of energy.
The Second Law of Thermodynamics (2) • Entropy is a measure of randomness or disorder. – It is energy not available to do additional work. – Loss of available energy equal TΔS, where ΔS is the change in entropy.
The Second Law of Thermodynamics (3) • Every event is accompanied by an increase in the entropy of the universe. – Entropy associated with random movements of particles or matter. – Living systems maintain a state of order, or low entropy
Free Energy (1) • The first and second laws of thermodynamics can be combined and expressed mathematically. – Equation: ΔH = ΔG + TΔS – Free energy, ΔG, is the energy available to do work. – Spontaneity of the reaction is ΔG, if <0 the reaction is exergonic, if >0 it is endergonic. – Spontaneity depends on both enthalpy and entropy.
Free Energy (2)
Free-Energy Changes in Chemical Reactions (1) • All chemical reactions are theoretically reversible. • All chemical reactions spontaneously proceed toward equilibrium (Keq = [C][D]/[A][B]). • The rates of chemical reactions are proportional to the concentration of reactants. • At equilibrium, the free energies of the products and reactants are equal (ΔG = 0).
Free-Energy Changes in Chemical Reactions (2) • Free energy changes of reactions are compared under standard conditions. – The standard free energy changes, ΔG°’, are described for each reaction under specific conditions. – Standard conditions are not representative of cellular conditions, but are useful to make comparisons. – Standard free energy changes are related to equilibrium: ΔG°’ = -RT ln K’eq
Calculation of free energy changes (1) • Non-standard conditions are corrected for prevailing conditions. – Equation: ΔG = ΔG°’ + RT ln Keq. – Prevailing conditions may cause ΔG to be negative, even when G°’ is positive. – Making ΔG negative may involve coupling endergonic and exergonic reactions in a sequence. – Simultaneously coupled reactions have a common intermediate. – ATP hydrolysis is often coupled to endergonic reactions in cells.
Calculation of free energy changes (1)
Coupling Endergonic and Exergonic Reactions
Coupling Endergonic and Exergonic Reactions
Equilibrium versus Steady-State Metabolism • Cellular metabolism is nonequilibrium metabolism. • Cells are open thermodynamic systems. • Cellular metabolism exists in a steady state. – Concentrations of reactants and products remain constant, but not at equilibrium. – New substrates enter and products are removed. – Maintaining a steady state requires a constant input of energy, whereas maintaining equilibrium does not.
Steady State versus Equilibrium
3. 2 Enzymes as Biological Catalysts • Enzymes are catalysts that speed up chemical reactions. • Enzymes are almost always proteins. • Enzymes may be conjugated with nonprotein components. – Cofactors are inorganic enzyme conjugates. – Coenzymes are organic enzyme conjugates.
Properties of Enzymes (1) • Are present in cells in small amounts. • Are not permanently altered during the course of a reaction. • Cannot affect thermodynamics of reactions, only the rates. • Are highly specific for their particular reactants called substrates. • Produce only appropriate metabolic products. • Can be regulated to meet the needs of a cell.
Properties of Enzymes (2)
Overcoming the Activation Energy Barrier • A small energy input, the activation energy (EA) is required for any chemical transformation. – The EA barrier slows the progress of thermodynamically unstable reactants. – Reactant molecules that reach the peak of the EA barrier are in the transition state.
Enzymes lower the activation energy • Without an enzyme, only a few substrate molecules reach the transition state. • With a catalyst, a large proportion of substrate molecules can reach the transition state.
The Active Site • An enzyme interacts with its substrate to form an enzyme-substrate (ES) complex. • The substrate binds to a portion of the enzyme called the active site. • The active site and the substrate have complementary shapes that allow substrate specificity.
The Active Site
Mechanisms of Enzyme Catalysis (1) • Substrate orientation means enzymes hold substrates in the optimal position of the reaction.
Mechanisms of Enzyme Catalysis (2) • Changes in the reactivity of the substrate temporarily stabilize the transition state. – Acidic or basic R groups on the enzyme may change the charge of the substrate. – Charged R groups may attract the substrate. – Cofactors of the enzyme increase the reactivity of the substrate by removing or donating electrons.
Mechanisms of Enzyme Catalysis (3) • Inducing strain in the substrate. – Shifts in the conformation after binding cause an induced fit between enzyme and the substrate. – Covalent bonds of the substrate are strained.
Mechanisms of Enzyme Catalysis (4) • Conformational changes and catalytic intermediates. – Various changes in atomic and electronic structure occur in both the enzyme and substrate during a reaction. – Using time-resolved crystallography, researchers have determined the threedimensional structure of an enzyme at successive stages during a reaction
Mechanisms of Enzyme Catalysis (4) • Conformational changes and catalytic intermediates. – Various changes in atomic and electronic structure occur in both the enzyme and substrate during a reaction. – Using time-resolved crystallography, researchers have determined the threedimensional structure of an enzyme at successive stages during a reaction
Enzyme Kinetics (1) • Kinetics is the study of rates of enzymatic reactions under various experimental conditions. • Rates of enzymatic reactions increase with increasing substrate concentrations until the enzyme is saturated. – At saturation every enzyme s working at maximum capacity. – The velocity at saturation is called maximal velocity, Vmax. – The turnover number is the number of substrate molecules converted to product per minute per enzyme molecule at Vmax.
Enzyme Kinetics (2) • The Michaelis constant (KM) is the substrate concentration at onehalf of Vmax. – Units of KM are concentration units. – The KM may reflect the affinity of the enzyme for the substrate.
Enzyme Kinetics (3) • Plots of the inverses of velocity versus substrate concentrations, such as the Lineweaver-Burk plot, facilitate estimating Vmax and KM. • Temperature and p. H can affect enzymatic reaction rates.
Enzyme Kinetics (4)
Enzyme Kinetics (4)
Enzyme Inhibitors (1) • Enzyme inhibitors slow the rates for enzymatic reactions. – Irreversible inhibitors bind tightly to the enzyme. – Reversible inhibitors bind loosely to the enzyme. • Competitive inhibitors compete with the enzyme for active sites – Usually resemble the substrate in structure. – Can be overcome with high substrate/inhibitor ratios.
Enzyme Inhibitors (2)
Enzyme Inhibitors (3) • Noncompetitive inhibitors – Bind to sites other than active sites and inactivate the enzyme. – The maximum velocity of enzyme molecules cannot be reached. – Cannot be overcome with high substrate/inhibitor ratios.
The Human Perspective: The Growing Problem of Antibiotic Resistance (1) • Antibiotics target human metabolism without harming the human host. – Enzymes involved in the synthesis of the bacterial cell wall. – Components of the system by which bacteria duplicate, transcribe, and translate their genetic information. – Enzymes that catalyze metabolic reactions specific to bacteria.
The Human Perspective: The Growing Problem of Antibiotic Resistance (2) • Antibiotics have been misused with dire consequences. – Susceptible cells are destroyed, leaving rare and resistant cells to survive and replicate. – Bacteria become resistant to antibiotics by acquiring genes from other bacteria by various mechanisms.
3. 3 Metabolism • Metabolism is the collection of bio-chemical reactions that occur within a cell. • Metabolic pathways are sequences of chemical reactions. – Each reaction in the sequence is catalyzed by a specific enzyme. – Pathways are usually confined to specific locations. – Pathways convert substrates into end products via a series of metabolic intermediates.
An Overview of Metabolism (1) • Catabolic pathways break down complex substrates into simple end products. – Provide raw materials for the cell. – Provide chemical energy for the cell. • Anabolic pathways synthesize complex end products from simple substrates. – Require energy. – Use ATP and NADPH from catabolic pathways.
An Overview of Metabolism (2) • Anabolic and catabolic pathways are interconnected. – In stage I, macromolecules are hydrolyzed into their building blocks. – In stage II, building blocks are further degraded into a few common metabolites. – In stage III, small molecular weight metabolites like acetyl-Co. A are degraded yielding ATP.
Oxidation and Reduction: A Matter of Electrons (1) • Oxidation-reduction (redox) reactions involve a change in the electronic state of reactants. – When a substrate gains electrons, it is reduced. – When a substrate loses electrons, it is oxidized. – When one substrate gains or loses electrons, another substance must donate or accept those electrons. • In a redox pair, the substrate that donates electrons is a reducing agent. • The substrate that gains electrons is an oxidizing agent.
Oxidation and Reduction: A Matter of Electrons (1)
The Capture and Utilization of Energy • Reduced atoms can be oxidized, releasing energy to do work. • The more a substance is reduced, the more energy that can be released. • Glycolysis is the first stage in the catabolism of glucose, and occurs in the soluble portion of the cytoplasm. • The tricarboxylic (TCA) cycle is the second stage and it occurs in the mitochondria of eukaryotic cells.
Glycolysis and ATP Formation (1) • Of the reactions of glycolysis, all but three are near equilibrium (ΔG ~ 0) under cellular conditions. • The driving forces of glycolysis are these three reactions.
Glycolysis and ATP Formation (2)
Glycolysis and ATP Formation (3) • Glucose is phosphorylated to glucose 6 phosphate by using ATP. • Glucose 6 -phosphate is isomerized to fructose 6 -phosphate. • Fructose 6 -phosphate is phosphorylated to fructose 1, 6 -bisphophate using another ATP. • Fructose 1, 6 -bisphosphate is split into two three-carbon phosphorylated compounds.
Glycolysis and ATP Formation (4) • NAD+ is reduced to NADH when glyceraldehyde 3 -phosphate is converted to 1, 3 bisphoglycerate. – Dehydrogenase enzymes oxidize and reduce cofactors. – NAD+ is a nonprotein cofactor associated with gluceraldehyde phosphate dehydrogenase. – NAD+ can undergo oxidation and reduction at different places in the cell. – NADH donates electrons to the electron transport chain in the mitochondria.
Glycolysis and ATP Formation (5)
Glycolysis and ATP Formation (5)
Glycolysis and ATP Formation (6) • ATP is formed when 1, 3 bisphoglycerate is converted to 3 phosphoglycerate by 3 phosphoglycerate kinase. – Kinase enzymes transfer phosphate groups. – Substrate-level phosphorylation occurs when ATP is formed by a kinase enzyme.
Glycolysis and ATP Formation (7) • ATP formation is only moderately endergonic compared with other phosphate transfer in cells. – Transfer potential shown when molecules higher on the scale have less affinity for the group being transferred than are the ones lower on the scale. – The less the affinity, the better the donor.
Glycolysis and ATP Formation (8) • 3 -phosphoglycerate is converted to pyruvate via three sequential reactions, in one of them a kinase phosphorylates ADP. • Glycolysis can generate a net of 2 ATPs for each glucose. • Glycolysis occurs in the absence of oxygen, it is an anaerobic pathway. • The end product, pyruvate, can enter aerobic or anaerobic catabolic pathways.
Anaerobic Oxidation of Pyruvate: The Process of Fermentation (1) • Fermentation restores NAD+ from NADH. – Under anaerobic conditions, glycolysis depletes the supply of NAD+ by reducing it to NADH. – In fermentation, NADH is oxidized to NAD+ by reducing pyruvate. – In muscle and tumor cells pyruvate is reduced to lactate. – In yeast and other microbes, pyruvate is reduced and converted to ethanol. – Fermentation is inefficient with only about 8% of the energy of glucose captured as ATP.
Anaerobic Oxidation of Pyruvate: The Process of Fermentation (2)
Reducing Power (1) • Anabolic pathways require a source of electrons to form larger molecules. • NADPH donates electrons to form large biomolecules. – NADPH is a nonprotein cofactor similar to NADH. – The supply of NADPH represents the cell’s reducing power. – NADP+ is formed by phosphate transfer from ATP to NAD+.
Reducing Power (2) • NADPH and NADH are interconvertible, but have different metabolic roles. • NADPH is oxidized in anabolic pathways. • NAD+ is reduced in catabolic pathways. • The enzyme transhydrogenase catalyzes the transfer of hydrogen atoms from one cofactor to the other. – NADPH is favored when energy is abundant. – NADH is used to make ATP when energy is scarce.
Metabolic Regulation (1) • Cellular activity is regulated as needed. • Regulation may involve controlling key enzymes of metabolic pathways. • Enzymes are controlled by alteration in active sites. – Covalent modification of enzymes regulated by phosphorylation such as protein kinases. – Allosteric modulation by enzymes regulated by compounds binding to allosteric sites. • In feedback inhibition, the product of the pathway allosterically inhibits one of the first enzymes of the pathway.
Metabolic Regulation (2)
Separating Catabolic and Anabolic Pathways (1) • Glycolysis and gluconeogenesis are the catabolic and anabolic pathways of glucose metabolism. – Synthesis of fructose 1, 6 -bisphosphate is coupled to hydrolysis of ATP. – Breakdown of fructose 1, 6 -bisphosphate is via hydrolysis by fructose 1, 6 -bisphosphatase in gluconeogenesis. – Phosphofructokinase is regulated by feedback inhibition with ATP as the allosteric inhibitor. – Fructose 1, 6 -bisphosphatase is regulated by covalent modification using phosphate binding. – ATP levels are highly regulated.
Separating Catabolic and Anabolic Pathways (2)
Separating Catabolic and Anabolic Pathways (3) • Anabolic pathways do not proceed via the same reactions as the catabolic pathways even though they may have steps in common. – Some catabolic pathways are essentially irreversible due to large ΔG°’ values. – Irreversible steps in catabolic pathways are catalyzed by different enzymes from those in anabolic pathways.
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