Bioenergetics and BIological Oxidation - Menorca.pdf

Bioenergetics and Biological Oxidation Daniel D. Menorca, MD

July 14, 2015 BIOENERGETICS INTRODUCTION Bioenergetics- the biology of energy transformations and energy exchanges within and between living things and their environment. I. LAWS OF THERMODYNAMICS Thermodynamics- study of thermal processes which transform heat energy into useful mechanical work and vice versa (1) Zeroth Law of Thermodynamics: The Thermal Equilibrium The law states that: If two systems are each in thermal equilibrium with a third, they are also in thermal equilibrium with each other. (2) First Law of Thermodynamics: Law of Conservation of Energy The law states that: Energy is neither created nor destroyed, although it can be changed from one form to another The total ENERGY of a system, including its surroundings, REMAINS CONSTANT. a) G = free energy; ΔG = change in free energy Gibbs Free energy (G)- energy associated with a chemical reaction that can be used to do work. b) Neither lost or gained c) May be transferred or transformed into another form of energy 1) Heat enegy/radiant energy →chemical energy 2) Chemical energy→heat energy/radiant energy

1ST SHIFTING

As a system approaches absolute zero, all processes cease and the entropy of the system approaches a minimum value.

ΔG=ΔH-TΔS where: ΔH= change in heat (enthalpy) T=absolute temperature Enthalpy- the sum of the internal energy of a body or system and the product of its volume multiplied by the pressure In biochemical reactions, since ΔH is approximately equal to the total change in internal energy of the reaction (ΔE), the above relationship may be expressed in the following way: ΔG=ΔE-TΔS II. EXERGONIC/ENDERGONIC/IRREVERSIBLE/ EQUILIBRIUM 1. If ΔG is negative, the reaction proceeds spontaneously with loss of free energy = EXERGONIC REACTION 2. If the negative ΔG is of great magnitude = IRREVERSIBLE 3. If ΔG is positive, the reaction cannot proceed spontaneously, unless G is supplied = ENDERGONIC REACTION 4. If the positivity of ΔG is very great = little or no reaction 5. If ΔG is zero = the reaction is at EQULIBRIUM III. EXOTHERMIC/ENDOTHERMIC/ ISOTHERMIC 1. Exothermic reaction = one form of exergonic reaction 2. Endothermic reaction = one form of endergonic reaction 3. Isothermic reaction = typical reactions in biological systems that may be exergonic or endergonic, reversible or irreversible, equilibrium or non-equilibrium IV. STANDARD FREE ENERGY CHANGE (ΔG⁰) Free energy change when the reactants are present in concentrations of 1.0 mol/L occurring at pH 7.0

(3)Second Law of Thermodynamics: Entropy Entropy- extent of disorder or randomness of a system The law states that: Whenever a spontaneous or irreversible process takes place, it is accompanied by an increase in the total entropy of the universe. The total ENTROPY of a system MUST INCREASE if a process is to occur SPONTANEOUSLY. a. S (entropy) b. ΔS= the change in entropy of a system (4) Third Law of Thermodynamics: Law of Enthalpy -Combination of 1st & 2nd law The law states that:

Santos,J., Sarte, Sijas

ΔG°’=-RT ln K’eq where: R = gas constant

RESPIRATORY CONTROL IN METABOLISM A. Catabolism = breakdown or oxidation of fuel molecules. An exergonic reaction. B. Anabolism = synthetic reactions that build up substances. An endergonic reaction. C. Metabolism = combined catabolic and anabolic processes. D. Coupling (exergonic and endergonic reactions)

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LOW AND HIGH-ENERGY PHOSPHATES ~(P), OTHER HIGH ENERGY COMPOUNDS IN INTERMEDIARY METABOLISM A. Low-energy Phosphates Ester phosphates -Intermediates of glycolysis -Their G°’ is smaller than ATP B. High-energy Phosphates Anhydrides (e.g. 1,3 BPG); Enol phosphates (e.g. PEP); Phosphoguanidines (e.g., Creatine-PO4) -Their G⁰’ is higher than ATP

Fig 1. Coupling of an exergonic to an endergonic reaction.

C. Others 1. Thiol Esters 2. Acyl carrier protein 3. Amino acid esters 4. S-adenosyl methionine 5. UDP-glucose 6. Phosphoribosylpyrophosphate

So that there will be no loss of energy as heat, we can have:

A+C→I→B+D Note: This type of system has a built-in mechanism for biologic control at the rate of oxidative process since the common obligatory intermediate (I) allows the rate of utilization of the product of the synthetic path (D) to determine by mass action the rate at which A is oxidized.

E. Compound with High Energy Potential, [~(E)]

Therefore, we can say that ATP has a G0 that is in the intermediate position

HIGH-ENERGY PHOSPHATES ~(P) ACT AS THE “ENERGY CURRENCY” OF THE CELL Table 1. The standard free energy of hydrolysis of some organophosphates (Low and High Energy) of Biochemical importance

Phosphoenolpyruvate

kJ/mol −61.9

ΔG°' kcal/mol −14.8

Carbamoyl phosphate

−51.4

−12.3

1,3-Bisphosphoglycerate

−49.3

−11.8

Creatine phosphate

−43.1

−10.3

ATP → AMP + PPi

−32.2

−7.7

ATP → ADP + Pi

−30.5

−7.3

Glucose 1-phosphate

−20.9

−5.0

PPi

−19.2

−4.6

Fructose 6-phosphate

-15.9

-3.8

Glucose 6-phosphate

-13.8

-3.3

Glycerol 3-phosphate

-9.2

-2.2

COMPOUND

Fig 2. Transfer of free energy from an exergonic to an endergonic reaction via a high-energy intermediate compound (~E)

Note: The biological advantage of the compound with high energy potential ~(E), unlike I, is that there is no need for ~(E) to be structurally related to A, B, C, D. It therefore can transduce energy from a wide range of exergonic and endergonic reactions. -energy potential is adenosine triphosphate (ATP). Both the obligatory intermediate I and the high-energy compound ~(E) supply a basis for the concept of RESPIRATORY CONTROL.

Santos,J., Sarte, Sijas

Note: ATP is able to act as a donor of high-energy phosphate to form those compounds below it in the table above. Likewise, ADP can accept high-energy phosphate to form ATP from those compounds above ATP. In effect, an ATP/ADP CYCLE connect those processes that generate ~(P) to those processes that utilize ~(P). This phenomenon is called ENERGY CONSERVATION OR ENERGY CAPTURE (following the 1st law of Thermodynamics)

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OTHER NUCLEOSIDE TRIPHOSPHATES PARTICIPATE IN THE TRANSFER OF HIGH-ENERGY PHOSPHATES

All of these triphosphates take part in phosphorylation in the cell

BIOLOGICAL OXIDATION TERMINOLOGIES

Fig 3. Role of ATP/ADP cycle in transfer of high-energy phosphate

Note: This occurs at a very rapid rate, since the total ATP/ADP pool is extremely small and sufficient to maintain an active tissue for only a few seconds. Sources of ~(P) taking part in energy conservation or capture: 1. OXIDATIVE PHOSPHORYLATION – Greatest quantitative source of ~(P) in aerobic organisms 2. GLYCOLYSIS – a net formation of two ~(P) from the formation of lactate from one molecule of glucose 3. CITRIC ACID CYCLE – one ~(P) is generated directly from CAC (at the succinate thiokinase step)

ATP ALLOWS THE COUPLING OF THERMODYNAMICALLY UNFAVORABLE REACTIONS TO FAVORABLE REACTIONS A. Thermodynamically unfavorable (endergonic) reaction:

Glucose + Pi → Glucose 6-PO4 + H2O

A Biological Oxidation 1. Oxidation-reduction reaction occurring in biochemical systems. 2. Whereas oxidation is lost of electrons, reduction is gain of electrons. 3. Therefore, oxidation of an electron donor is always coupled with reduction of an electron acceptor. 4. This principle applies equally to biochemical systems. B. Redox Reaction 1. Shortened term for oxidation-reduction reaction. 2. In the laboratory, reaction of hydrogen with oxygen will produce this reaction:

Where: H2 is oxidized (electron donor) O2 is reduced (electron acceptor) 3. In biochemical systems, the reaction of H2 with O2 is controlled by enzymes and instead of dissipating heat, the cells stores energy [~(P)] in the form of ATP. C. Respiration (in biochemical systems) 1. The process by which cells derive energy in the form of ATP from the controlled reaction of hydrogen with oxygen to form water. Also known as cellular respiration

(ΔG⁰’ = +13.8 kJ/mol) B. Thermodynamically favorable (exergonic) reaction:

ATP → ADP + Pi (ΔG°’ = -30.5 kJ/mol) C. Coupling of the two reactions through ATP

The enzymes* involved in this respiratory process are the so called Enzymes of Biologic Oxidation.

(ΔG°’ = -16.7 kJ/mol)

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D. Redox Potential (E0) 1. The tendency of reactions in redox reaction to donate or accept electrons. 2. Proportionate to ΔG° (1st Law of Thermodynamics) Where ΔG°’ = standard free energy change E’0 = standard redox potential. E’0 is normally expressed at pH 7.0 where the electrode potential of hydrogen electrode is –0.42 volts [heat energy (kJ) is transformed to electrical energy (volts), consistent with the 1st Law of Thermodynamics].

SOME REDOX POTENTIALS OF SPECIAL INTEREST IN MAMMALIAN OXIDATION SYSTEM System

E’0 Volts

H+/H2

−0.42

NAD+/NADH

−0.32

Lipoate; ox/red

−0.29

Acetoacetate/3-hydroxybutyrate

−0.27

Pyruvate/lactate

−0.19

Oxaloacetate/malate

−0.17

Fumarate/succinate

0.03

Cytochrome b; Fe3+/Fe2+

0.08

Ubiquinone; ox/red

0.1

Cytochrome c1; Fe3+/Fe2+

0.22

Cytochrome a; Fe3+/Fe2+

0.29

Oxygen/water

0.82

The relative position of redox systems in the table allow prediction of the direction of flow of electrons from one redox couple to another.

Enzymes of Biologic Oxidation A. Oxidases 1. Catalyze the removal of hydrogen from a substrate using oxygen as hydrogen acceptor. 2. They form H2O or H2O2 as reaction product. 3. Some are hemoproteins (e.g., cytochrome oxidase) 4. Some are flavoproteins a. FMN-linked (e.g. L-amino acid oxidase) b. FAD-linked (e.g. aldehyde dehydrogenase)

B. Dehydrogenase 1. Cannot use oxygen as hydrogen acceptor. Perform two functions: a. Transfer hydrogen from one substrate to another in a coupled oxidation-reduction reaction. b. Component of respiratory chain for electron transport from substrate to oxygen. 2. Many depend on Nicotinamide Coenzymes a. NAD-linked – catalyze redox reaction in the oxidative pathways of metabolism i. Glycolysis ii. CAC

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iii. Respiratory Chain b. NADP-linked – found characteristically in reductive syntheses i. Extramitochondrial fatty acid synthesis ii. Pentose Phosphate Pathway iii. Steroid synthesis 3. Others depend on Riboflavin a. Electron transport in the respiratory chain i. NADH dehydrogenase ii. Succinate DH, Acyl CoA DH, Glycerol-3PO4 DH b. Dehydrogenation of reduced lipoate i. Diphydrolipoyl dehydrogenase ii. Oxidative decarboxylation of pyruvate and alpha-ketoglutarate c. Intermediate carrier of electrons between AcylCoA DH and the respiratory chain i. Electron Transferring Flavoprotein (ETF) 4. Cytochromes may also be regarded as dehydrogenases a. Iron-containing hemoproteins in which the iron atoms oscillate between Fe3+ and Fe2+ during redox reaction b. In the respiratory chain, they are involved as carriers of electrons from flavoproteins to cytochrome oxidase c. Aside from the respiratory chain (Cyt b, c, c, c1 and Cytochrome oxidase), some are located in the endoplasmic reticulum (Cyt P450 and Cyt b5), plant cells, bacteria and yeasts. (note: Cyt oxidase is an oxidase while Cyt P450 is a monooxygenase)

C. Hydroperoxidases 1. Protect the body against harmful peroxidase 2. Two enzymes fall into this category: a. Peroxidases b. Catalase 3. In regard to peroxidase: a. Found in milk, leukocytes, platelets and other tissues involved in eicosanoid metabolism

(eicosanoids are signaling molecules made from metabolism of fatty acids, they serve their functions in inflammatory and other immune responses.) b. Hydrogen peroxidase is reduced at the expense of several substances that will act as electron acceptors such as ascorbate, quinones, and cytochrome c.

peroxidase

H2O2 + AH2

2H2O + A

c. In Red Blood Cells, there is the enzyme glutathione peroxidase that utilizes reduced glutathione. Glutathione Peroxidase

H2O2 + 2GSH

2H2O + GS-SG

4. In regard to catalases: a. It is a hemoprotein containing four heme groups b. Uses H2O2 as both electron donor and electron acceptor

Catalase 2H2O2

2H2O + O2 4

c. Found in blood, bone marrow, mucuos membrane, kidney and liver

4. Reoxidized by substrates in a series of reaction known as HYDROXYLASE CYCLE

D. Oxygenases 1. Catalyze the direct transfer and incorporation of oxygen into a substrate molecule 2. Divide into two subgroups: a. Dioxygenases b. Monooxygenases DIOOXYGENASES a. They incorporate both atoms of molecular oxygen into the substrate. [A + O2 AO2] b. Examples: i. Homogentisate dioxygenase ii. 3-OH anthranilate dioxygenase iii. L-Tryptophan dioxygenase MONOOXYGENASES: a. They incorporate only one atom of molecular oxygen into the substrate. [A-H + O2 + ZH2  A-OH + H2O + Z] b. Examples include: i. Cytochrome P450 1. Heme-containing monooxygenase 2. Location and function LOCATION Endoplasmic reticulum of hepatocytes Mitochondria of the adrenal cortex, testis, ovary and placenta Renal cells

FUNCTION Detoxification of substrates and bile synthesis Biosynthesis of steroid hormones from cholesterol Vit. D metabolism

3. Both NADH and NADPH donate reducing equivalents for its reduction

Fig 5. Cytochrome P450 hydroxylase cycle.

E. Superoxide Dismutase - an enzyme responsible for the removal of O2 in all aerobic organisms (although not in obligate anaerobes) indicate that the potential toxicity of oxygen is due to its conversion to superoxide. A. Transfer of a single electron to O2 generates the potentially damaging superoxide anion free radical (O2-). B. The destructive effect of this free radical can be amplified by a side chain reaction 1. Initiation

ROOH + Metal(n)+ X- + RH R- + XH

ROO+ Metal(n-1)+ + H+

2. Propagation

R- + O2 ROOROO- + RH ROOH + R, etc. 3. Termination Fig 4. Electron transport chain in the endoplasmic reticulum. Cyanide (CN ) inhibits the indicated step. −

ROO- + ROOROOR + O2 ROO- + RROOR R - + RRR C. In vivo, the principal chain-breaking antioxidants are superoxide dismutase, which acts in the aqueous phase to trap superoxide free radicals (O2-) urate and vitamin C, which acts in the lipid phase to trap ROO radicals. D. Superoxide can reduce oxidized cytochrome c or be removed by superoxide dismutase. O2- + Cyt c(Fe3+)

O2+ Cyt c (Fe2+)

In this reaction, superoxide acts as both oxidant and reductant. Thus, superoxide dismutase protects aerobic organisms against the potential deleterious effects of superoxide.

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E. This enzyme occurs in all major aerobic tissues in the mitochondria and cytosol. F. Although exposure of animals to an atmosphere of 100% oxygen causes an adaptive increase in superoxide dismutase, prolonged exposure to 100% oxygen leads to lung damage and death.

*Antioxidants, eg, α-tocopherol (vitamin E), act as scavengers of free radicals and reduce the toxicity of oxygen

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