Organisms Use the First Law Big-Time

Biochemistry II OUTLINE Review of 421 Goals of 422 Review of chemical principles Thermodynamics & ENERGY C/O cycles Overview of Metabolism ATP cycle Energy Coupling Chemical Reactivity Bioenergetics Organisms Use the First Law Big-Time (perform energy transformations) to Stay Alive Favorable and Unfavorable Reactions • DS > 0 The breakdown of some metabolites releases a significant amount of energy (exergonic). – Their cellular concentration is far higher than their equilibrium concentration. – Metabolites, such as ATP, NADH, NADPH, can be synthesized using the energy from sunlight and fuels….. • DG°< 0 Synthesis of complex molecules and many other metabolic reactions requires energy (endergonic). – A reaction might be thermodynamically unfavorable (DG°> 0). catabolism • Creating order requires work and energy. DG°> 0 anabolism • Biochemistry couples exergonic with endergonic reactions to insure organisms continue to grow and divide. 1 Energetics of Life • Living organisms cannot create energy from nothing. • Living organisms cannot destroy energy into nothing. • Living organism may transform energy from one form to another. • In the process of transforming energy, living organisms must increase the entropy of the universe. • In order to maintain organization within themselves, living systems must be able to extract useable energy from their surroundings and release useless energy (heat) back to their surroundings. Overview of Metabolism 2 Metabolism Issues: • Thermodynamics and biochemistry; carbon/oxygen cycle & nitrogen cycle • Common organic-chemical principles in biochemistry • Some biomolecules are “high energy” with respect to their hydrolysis and group transfers. • Energy stored in reduced organic compounds can be used to reduce cofactors such as NAD+ and FAD, which serve as universal electron carriers and lead to ATP formation. Metabolism • Carbon/oxygen cycle + • Recall that living organisms are built of complex structures. • Building complex structures means paying a huge entropic price; need a constant input of energy. • The ultimate source of this energy on Earth is sunlight 3 Metabolism – In biochemistry, the oxidation reduced Metabolism is the Sum of All Chemical Reactions in of the Cell fuels with O2 is stepwise and controlled. – Recall that being thermodynamically favorable is not the same as being kinetically rapid. Catabolism: cofactor interm ediates Metabolism – In biochemistry, the oxidation reduced Metabolism is the Sum of All Chemical Reactions in of the Cell fuels with O2 is stepwise and controlled. – Recall that being thermodynamically favorable is not the same as being kinetically rapid. Catabolism: convergent 4 Metabolism NAD+ and NADP+ Are Common Redox Cofactors • These are commonly called pyridine nucleotides. • They can dissociate from the enzyme after the reaction. • In a typical biological oxidation reaction, hydride from an alcohol is transferred to NAD+, giving NADH. FAD and FADH2 are another Common Redox Cofactors • These are commonly called flavins. • They are usually covalently bound at the active site of enzymes. • They can undergo both 1-electron and 2-electron redox reactions. Metabolism THE ATP CYCLE ATP-4 + H2O –2 + H+ – + HPO ADP-3 + H2PO 4 4 (at pH 7) Adenosine diphosphate (ADP) This “buffering” of energy in the cell keeps the [ATP] high Energy enough to keep fighting the second law of thermodynamics. Charge = + ½ [ADP-3] [ATP-4] + [ADP-3] + [AMP-2] [ATP-4] 5 Metabolism –22– – + + HPO HPO + H 4 44 + H 2PO (at pH 7) + H+ + H2O Adenosine diphosphate (ADP) ADP-3 ATP-4 (at pH 7) If this reaction is allowed to come to equilibrium, what is the DG? the actual free-energy change of a reaction depends on TWO things: 1) A constant (the standard change in free energy, DG°) 2) The actual concentrations of products and reactants – For the reaction aA + bB ⇌ cC + dD: c d [C ] [ D] DG′= DG'°′ ° + RT ln [ A]a [ B]b [In biochemistry, we add a prime (′) because we pull [H+] out of the expression and set it to 10-7, not 1 M] Metabolism + HPO4–2 + H+ + H2O (at pH 7) Adenosine diphosphate (ADP) c d [C ] [ D] DG′= DG'°′ ° + RT ln [ A]a [ B]b [C ]c [ D]d DG′= DG°′ '° + RT ln K′eqa b d ] [ B] [C ]c [ D[]A DG = DG°′ '°=+–RT ln K′eqa b [ A] [ B] TABLE 13-2 Relationship between Equilibrium Constants and Standard FreeEnergy Changes of Chemical Reactions DG' DG°′= DG°ʹ= –7.3 kcal/mole kcal/mole (kJ/mol) 103 –17.1 102 –11.4 –2.7 101 –5.7 –1.4 0.0 0.0 1 TABLE 13-3 Relationships among K'eq, DG'˚, and the Direction of Chemical Reactions When K'eq is . . . >1.0 1.0 <1.0 DG'˚ is . . . Starting with all components at 1 M, the reaction . . . negative proceeds forward zero is at equilibrium positive proceeds in reverse 10–1 But, what is the actual DG′ of ATP Hydrolysis IN THE CELL? (kcal/mol)a K'eq –4.1 5.7 1.4 10–2 11.4 2.7 10–3 17.1 4.1 10–4 22.8 5.5 10–5 28.5 6.8 10–6 34.2 8.2 Although joules and kilojoules are the standard units of energy and are used throughout this text, biochemists and nutritionists sometimes express DG'˚ values in kilocalories per mole. We have therefore included values in both kilojoules and kilocalories in this table and in Tables 13-4 and 13-6. To convert kilojoules to kilocalories, divide the number of kilojoules by 4.184. a 6 Metabolism + HPO4–2 + H+ + H2O (at pH 7) Adenosine diphosphate (ADP) c Go back the original equation: At 25 °C: DG°′ of ATP Hydrolysis is Mg++ Dependent DG′ = DG°′ [ADP-3] -2] [HPO4 [ATP-4] [ADP-3] [HPO4-2] + 0.59ln [ATP-4] DG′ = DG°′ + RT ln [MgADP- ] × [Pi ] DG = DG °'+ RT ln [MgATP2- ] d [C ] [ D] DG′= DG'°′ ° + RT ln [ A]a [ B]b In RBCs = 1.5 x 10-4 M DG′ = DG°′ + 1.36log (1.5 x 10-4) DG′ = (-7.3) + (–5.2) DG′ = -12.5 kcal/mole WOW! Other than concentrations, what makes the DG′ of ATP so high, i.e., DG°′ ? Metabolism • Contributions of the incredible favorability of the ATP hydrolysis reaction: – Better charge separation in products (relief of charge repulsion) – More favorable resonance stabilization of products – Ionization of products (ADP or Pi) ❸ OR H2PO4– ⇌ HPO 42– + H+ (at pH 7) Are there other compounds in the cell with high negative DG°′ of hydrolysis? 7 Metabolism DG°′= –12 kcal/mole TABLE 13-4 Standard Free-Energy Changes of Some Chemical Reactions D G'˚ Reaction type 1,3-bisphosphoglycerate DG°′= –11.8 kcal/mole DG°′= –10.3 kcal/mole (PCr) (kJ/mol) (kcal/mol) Hydrolysis reactions Acid anhydrides Acetic anhydride + H2O à 2 acetate ATP + H2O à ADP + Pi ATP + H2O à AMP + PPi PPi + H2O à 2Pi UDP-glucose + H2O à UMP + glucose 1-phosphate –91.1 –30.5 –45.6 –19.2 –43.0 –21.8 –7.3 –10.9 –4.6 –10.3 Esters Ethyl acetate + H2O à ethanol + acetate Glucose 6-phosphate + H2O à glucose + Pi –19.6 –13.8 –4.7 –3.3 Amides and peptides Glutamine + H2O à glutamate 1 NH4+ Glycylglycine + H2O à 2 glycine –14.2 –9.2 –3.4 –2.2 Glycosides Maltose + H2O Lactose + H2O –15.5 –15.9 –3.7 –3.8 –7.3 –1.7 –1.7 –0.4 à à 2 glucose glucose + galactose Rearrangements Glucose 1-phosphate à glucose 6-phosphate Fructose 6-phosphate à glucose 6-phosphate Elimination of water Malate à fumarate + H2O Oxidations with molecular oxygen Glucose + 6O2 à 6CO2 + 6H2O Palmitate + 23O2 à 16CO2 + 16H2O 3.1 0.8 –2,840 –9,770 –686 –2,338 DG°′= –14.8 kcal/mole = –61.9 kJ/mole (PEP) Metabolism Phosphates: Ranking by the Standard Free Energy of Hydrolysis Phosphate can be transferred from compounds with higher ΔG°′ to those with lower ΔG°′. 8 Metabolism TABLE 13-5 Total Concentrations of Adenine Nucleotides, Inorganic Phosphate, and Phosphocreatine in Some Cells Concentration (mM)a ATP ADPb AMP Energy Charge Pi Rat hepatocyte 3.38 1.32 0.29 4.8 0 Rat myocyte 8.05 0.93 0.04 0.81 0.94 0.87 0.94 0.86 8.05 28 2.72 4.7 Rat neuron 2.59 0.73 0.06 Human erythrocyte 2.25 0.25 0.02 E. coli cell 7.90 1.04 0.82 PCr 1.65 0 7.9 0 a For erythrocytes the concentrations are those of the cytosol (human erythrocytes lack a nucleus and mitochondria). In the other types of cells the data are for the entire cell contents, although the cytosol and the mitochondria have very different concentrations of ADP. PCr is phosphocreatine, discussed on p. 516. b This value reflects total concentration; the true value for free ADP may be much lower (p. 509). Cellular ATP concentration is usually far above the equilibrium concentration, making ATP a very potent source of chemical energy. Energy Coupling 9 Energy Coupling • Question is how do cells use these “high-energy compounds” to ensure that thermodynamically unfavorable reactions can become favorable? • Chemical coupling of exergonic and endergonic reactions allows otherwise unfavorable reactions. • TWO basic mechanisms: – e.g., The hydrolysis of the “high-energy” molecule (ATP), which is exergonic, is coupled with an otherwise endergonic reaction during an enzyme mechanism. – e.g., a series of otherwise endergonic reactions during a metabolic pathway are linked in the reaction series to an overwhelming exergonic reaction (all catalyzed by enzymes). Energy Coupling enzyme mechanism Enzymatic reaction These two reactions take place at the same time and place (active site) Replace the H–O–H with H–O–R (glucose) + Glucose Kinase (aka Hexokinase) Glucose 6-phosphate 10 Energy Coupling enzyme mechanism Two-Step Group Transfers Facilitate ATP-Dependent Reactions Energy Coupling metabolic pathway Standard free-energy changes are additive: (1) A (2) B Sum: A B ΔG1 C ΔG2 C ΔG1 + ΔG2 Coupling exergonic and endergonic reactions: SgPgQgRgZ Net: S P Q R S P Q R Z Z DG = +12 kcal/mole DG = +1 kcal/mole DG = -4 kcal/mole DG = -14 kcal/mole DG = -5 kcal/mole 11 Chemical Reactivity First lets quickly review bond cleavage and nucleophilicity in terms of biochemical instances Chemical Reactivity Chemistry at Carbon: Homolytic versus Heterolytic Cleavage • Covalent bonds can be broken in two ways. • Homolytic cleavage is rare in biochemistry, but makes up some of the most interesting reactions. • Heterolytic cleavage is the most common, but the products are highly unstable, dictating the chemistry that occurs: mostly carbanion chemistry 12 Chemical Reactivity Nucleophiles and Electrophiles in Biochemistry Definition: -phile (suffix, Latin): to love, or demonstrate fondness towards • Nucleophiles love nuclei. The nuclei of an atom contains positively charged particles (protons), so nucleophiles love to react with positive charges and are electron rich. • Electrophiles love electrons. Electrons are negatively charged, so electrophiles must have at least a partial positive charged. 13