Energetics of carbohydrate

and lipid metabolism

1

Metabolism: The sum of all the chemical transformations taking

place in a cell or organism, occurs through a series of enzyme-

catalyzed reactions that constitute metabolic pathways.

Each of the consecutive

• Steps in a metabolic pathway brings about a specific, small

chemical change, usually the removal, transfer, or addition of a

particular atom or functional group.

• The precursor is converted into a product through a series of

metabolic intermediates called metabolites.

2

• The combined activities of all the metabolic pathways that

interconvert precursors, metabolites, and products of low

molecular weight, Mr <1,000 (intermediary metabolism).

Catabolism (Degradative phase) :

• Organic nutrient molecules (carbohydrates, fats, and

proteins) are converted into smaller, simpler end products

(such as lactic acid, CO2, NH3).

• Catabolic pathways release energy, some of which is

conserved in the formation of ATP and reduced electron

carriers (NADH, NADPH, and FADH2); the rest is lost as heat.

3

Anabolism(biosynthesis): Small, simple precursors are built

up into larger and more complex molecules, including

lipids, polysaccharides, proteins, and nucleic acids.

• Anabolic reactions require an input of energy, generally

in the form of the phosphoryl group transfer potential of

ATP and the reducing power of NADH, NADPH, and

FADH2.

• Some metabolic pathways are linear, and some are

branched.

• Catabolic pathways are convergent and anabolic pathways

divergent.

• Some pathways are cyclic: one starting component of the

pathway is regenerated in a series of reactions that converts

another starting component into a product.

• Most cells have the enzymes to carry out both the

degradation and the synthesis of the important categories

of biomolecules—fatty acids.

• Both anabolic and catabolic pathways to be essentially

irreversible, the reactions unique to each direction must include at

least one that is thermodynamically very favorable, a reaction for

which the reverse reaction is very unfavorable.

• Metabolic pathways are regulated at several levels from within

the cell and from outside.

• The most immediate regulation is by the availability of substrate;

when the intracellular concentration of an enzyme’s substrate is

near or below Km, the rate of the reaction depends strongly upon

substrate concentration.

• The number of metabolic transformations taking place in a

typical cell. Most cells have the capacity to carry out

thousands of specific, enzyme-catalyzed reactions:

Examples:

• Transformation of a simple nutrient such as glucose into

amino acids, nucleotides, or lipids.

• Extraction of energy from fuels by oxidation.

• Polymerization of monomeric subunits into macromolecules.

The reactions in living cells fall into one of five

general categories usually proceed by a limited set

of mechanisms and often employ characteristic

cofactors :

(1) Oxidation-reductions; (2) Reactions that make or

break carbon–carbon bonds; (3) Internal

rearrangements, isomerization's, and eliminations,

(4) Group transfers, and (5) Free radical reactions.

Reactions within each

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• Two phases of glycolysis. For each molecule of glucose that passes

through the preparatory phase (a), two molecules of

glyceraldehyde 3-phosphate are formed; both pass through the

payoff phase (b).

• Pyruvate is the end product of the second phase of glycolysis.

• For each glucose molecule, two ATP are consumed in the

preparatory phase and four ATP are produced in the payoff phase,

giving a net yield of two ATP per molecule of glucose converted to

pyruvate.

•

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12

13

ATP Formation Coupled to Glycolysis

• During glycolysis some of the energy of the glucose molecule is

conserved in ATP, while much remains in the product, pyruvate.

• The overall equation for glycolysis is

• For each molecule of glucose degraded to pyruvate, two

molecules of ATP are generated from ADP and Pi. We can now

resolve the equation of glycolysis into two processes

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The conversion of glucose to pyruvate (exergonic)

The formation of ATP from ADP and Pi, (endergonic)

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The sum of Equations 14–2 and 14–3 gives the overall standard free-energy change of glycolysis, ΔGs´°

Fermentation: The general term for such processes, which

extract energy (as ATP) but do not consume oxygen or change

the concentrations of NAD+ or NADH.

• In case of insufficient oxygen NAD+ is regenerated from

NADH by the reduction of pyruvate to L-lactate by lactate

dehydrogenase at pH 7.

16

The overall equilibrium of this reaction strongly favors lactate formation, as shown by the large negative standard free-energy change.

• An endergonic reaction (also called a heat absorb

nonspontaneous reaction or an unfavorable reaction) is a

chemical reaction in which the standard change in free

energy is positive, and energy is absorbed.

• A reaction that is thermodynamically favored has a

negative ΔG, so the products are at a lower energy than

the reactants so that reaction will spontaneously occur. For

ΔG to be negative, ΔH has to be small in the equation

ΔG=ΔH−TΔS. (exergonic reaction)

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1- Phosphorylation of Glucose in glycolysis

• Use the energy of ATP

• Multiple isoforms of hexokinase exist in organisms (e.g.,

hexokinase I, II, III, and IV (glucokinase)).

• Nucleophilic oxygen at C6 of glucose attacks the last (γ)

phosphate of ATP.

• ATP-bound Mg++ facilitates this process by shielding the

negative charges on ATP.

• Highly thermodynamically favorable/irreversible

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2- Phosphohexose Isomerization

Slightly thermodynamically unfavorable/reversible product

concentration kept low by pairing with favorable next step to drive

reaction forward.

3- 2nd Priming Phosphorylation by Phosphofructokinase-1

This process uses the energy of ATP. Highly thermodynamically

favorable/irreversible

4- Aldol Cleavage of F-1,6-bP by Aldolase

Thermodynamically unfavorable/reversible

5- Triose Phosphate Interconversion by Triose Phosphate

Isomerase

Thermodynamically unfavorable/reversible

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6- Oxidation of GAP by Glyceraldehyde-3-Phosphate

Dehydrogenase

Thermodynamically unfavorable/reversible

7- 1st Production of ATP by Phosphoglycerate Kinase

Highly thermodynamically favorable/reversible

8- Migration of the Phosphate

Thermodynamically unfavorable/reversible

9- Dehydration of 2-PG to PEP

Slightly thermodynamically unfavorable/reversible

10- 2nd Production of ATP by Pyruvate kinase

Highly thermodynamically favorable/irreversible

20

Only a Small Amount of Energy Available in Glucose Is Captured in Glycolysis

2G′ = –146 kJ/mol

Glycolysis

Full oxidation (+ 6 O2)

G′ = –2,840 kJ/mol6 CO2 + 6 H2O

GLUCOSE

Cellular Respiration

• Process in which cells consume O2 and produce CO2

• Provides more energy (ATP) from glucose than glycolysis

• Also captures energy stored in lipids and amino acids • Evolutionary origin: developed about 2.5 billion years

ago• Used by animals, plants, and many microorganisms• Occurs in three major stages:

- acetyl CoA production- acetyl CoA oxidation- electron transfer and oxidative phosphorylation

Respiration: Stage 1Acetyl-CoA Production

Generates some ATP, NADH,

FADH2

Carbohydrates release 1/3 of total potential CO2

during Stage 1.

Respiration: Stage 2Acetyl-CoA Oxidation

Generates more NADH, FADH2,and one GTP

Remaining carbon atoms from carbohydrates, amino acids, and fatty acids are released during Stage 2.

Respiration: Stage 3Oxidative Phosphorylation

Generates the vast

majority of ATP during catabolism

In Eukaryotes, Stages 2 and 3 Are Localized to the Mitochondria

• Glycolysis occurs in the cytoplasm.

• Citric acid cycle occurs in the mitochondrial matrix†.

• Oxidative phosphorylation occurs in the inner membrane.

†Except succinate dehydrogenase, which is located in the inner membrane

The Citric Acid Cycle (CAC)

Sequence of Events in the Citric Acid Cycle

• Step 1: C-C bond formation between acetate (2C) and

oxaloacetate (4C) to make citrate (6C)

• Step 2: Isomerization via dehydration/rehydration

• Steps 3–4: Oxidative decarboxylations to give 2 NADH

• Step 5: Substrate-level phosphorylation to give GTP

• Step 6: Dehydrogenation to give FADH2

• Step 7: Hydration

• Step 8: Dehydrogenation to give NADH

One Turn of the Citric Acid Cycle

Direct and Indirect ATP Yield

TABLE 16-1 Stoichiometry of Coenzyme Reduction and ATP Formation in the Aerobic

Oxidation of Glucose via Glycolysis, the Pyruvate Dehydrogenase Complex

Reaction, the Citric Acid Cycle, and Oxidative Phosphorylation

Reaction Number of ATP or reduced

coenzyme directly formed

Number of ATP

ultimately formeda

Glucose glucose 6-phosphate –1 ATP –1

Fructose 6-phosphate fructose 1,6-bisphosphate –1 ATP –1

2 Glyceraldehyde 3-phosphate 2 1,3-bisphosphoglycerate 2 NADH 3 or 5b

2 1,3-Bisphosphoglycerate 2 3-phosphoglycerate 2 ATP 2

2 Phosphoenolpyruvate 2 pyruvate 2 ATP 2

2 Pyruvate 2 acetyl-CoA 2 NADH 5

2 Isocitrate 2 α-ketoglutarate 2 NADH 5

2 α-Ketoglutarate 2 succinyl-CoA 2 NADH 5

2 Succinyl-CoA 2 succinate 2 ATP (or 2 GTP) 2

2 Succinate 2 fumarate 2 FADH2 3

2 Malate 2 oxaloacetate 2 NADH 5

Total 30-32

aThis is calculated as 2.5 ATP per NADH and 1.5 ATP per FADH2. A negative value indicates consumption.bThis number is either 3 or 5, depending on the mechanism used to shuttle NADH equivalents from the cytosol to the

mitochondrial matrix; see Figures 19-30 and 19-31.

Fats Provide Efficient Fuel Storage

• The advantage of fats over polysaccharides:

– Fatty acids carry more energy per carbon because they are

more reduced.

– Fatty acids complex or carry less water because they are

nonpolar.

• Glucose and glycogen are for short-term energy needs and quick

delivery.

• Fats are for long-term (months) energy needs, good storage,

and slow delivery.

Fatty Acid Oxidation Occurs in the Mitochondria in Three Stages

• Stage 1 consists of oxidative conversion of two-carbon units into acetyl-CoA via oxidation with concomitant generation of NADH and FADH2.– involves oxidation of carbon to thioester of fatty acyl-CoA

• Stage 2 involves oxidation of acetyl-CoA into CO2 via citric acid cycle with concomitant generation NADH and FADH2.

• Stage 3 generates ATP from NADH and FADH2 via the respiratory chain.

Stages of Fatty Acid Oxidation

The -Oxidation Pathway Each pass removes one acetyl moiety in the form of acetyl-CoA.

Fatty Acid Catabolism for Energy

• For palmitic acid (C16)– Repeating the previous four-step process six more

times (seven total) results in eight molecules of acetyl-CoA.

• FADH2 is formed in each cycle (seven total).• NADH is formed in each cycle (seven total).

• Acetyl-CoA enters citric acid cycle and further oxidizes into CO2.– This makes more GTP, NADH, and FADH2.

• Electrons from all FADH2 and NADH enter ETF.

NADH and FADH2 Serve as Sources of ATP

TABLE 17-1 Yield of ATP during Oxidation of One Molecule of Palmitoyl-CoA to CO2

and H2O

Enzyme catalyzing the oxidation step

Number of NADH or

FADH2 formed

Number of ATP

ultimately formeda

β Oxidation

Acyl-CoA dehydrogenase 7 FADH2 10.5

β-Hydroxyacyl-CoA dehydrogenase 7 NADH 17.5

Citric acid cycle

Isocitrate dehydrogenase 8 NADH 20

α-Ketoglutarate dehydrogenase 8 NADH 20

Succinyl-CoA synthetase 8b

Succinate dehydrogenase 8 FADH2 12

Malate dehydrogenase 8 NADH 20

Total 108

aThese calculations assume that mitochondrial oxidative phosphorylation produces 1.5 ATP per FADH2 oxidized and 2.5 ATP

per NADH oxidized.bGTP produced directly in this step yields ATP in the reaction catalyzed by nucleoside diphosphate kinase (p. 516).

Oxidation-Reduction Reactions

Reduced organic compounds serve as fuels from which electrons can be stripped off during oxidation.

Reversible Oxidation of a Secondary Alcohol to a Ketone

• Many biochemical oxidation-reduction reactions involve transfer of two electrons.

• In order to keep charges in balance, proton transfer often accompanies electron transfer.

• In many dehydrogenases, the reaction proceeds by a stepwise transfers of proton (H+) and hydride (:H–).

Reduction Potential Determines Flow of Electrons

• Reduction potential (E)– affinity for electrons; higher E, higher affinity– electrons transferred from lower to higher E

E’ = −(RT/nF)ln (Keq) = G’/nF

∆E’ = E’(e− acceptor) – E’(e− donor)

∆G’ = –nF∆E’For negative G, need positive E

E(acceptor) > E(donor)

TABLE 13-7a Standard Reduction Potentials of Some Biologically Important Half-

Reactions

Half-reaction E'˚ (V)

1/2 O2 + 2H+ + 2e– H2O 0.816

Fe3+ + e– Fe2+ 0.771

NO3– + 2H+ + 2e–

NO2– + H2O 0.421

Cytochrome f (Fe3+) + e– cytochrome f (Fe2+) 0.365

Fe(CN)63– (ferricyanide) + e–

Fe(CN)64– 0.36

Cytochrome a3 (Fe3+) + e– cytochrome a3 (Fe2+) 0.35

O2 + 2H+ + 2e– H2O2 0.295

Cytochrome a (Fe3+) + e– cytochrome a (Fe2+) 0.29

Cytochrome c (Fe3+) + e– cytochrome c (Fe2+) 0.254

Cytochrome c1 (Fe3+) + e– cytochrome c1 (Fe2+) 0.22

Cytochrome b (Fe3+) + e– cytochrome b (Fe2+) 0.077

Ubiquinone + 2H+ + 2e– ubiquinol 0.045

Fumarate2– + 2H+ + 2e– succinate2– 0.031

2H+ + 2e– H2 (at standard conditions, pH 0) 0.000

Crotonyl-CoA + 2H+ + 2e– butyryl-CoA –0.015

Oxaloacetate2– + 2H+ + 2e– malate2– –0.166

Source: Data mostly from R. A. Loach, in Handbook of Biochemistry and Molecular Biology, 3rd edn (G. D. Fasman, ed.), Physical and Chemical Data, Vol. 1, p. 122, CRC Press, 1976.aThis is the value for free FAD; FAD bound to a specific flavoprotein (e.g., succinate dehydrogenase) has a different E'˚ that depends

on its protein environment.

TABLE 13-7b Standard Reduction Potentials of Some Biologically Important Half-

Reactions

Half-reaction E'˚ (V)

Pyruvate– + 2H + 2e– lactate– –0.185

Acetaldehyde + 2H+ + 2e– ethanol –0.197

FAD + 2H+ + 2e– FADH2 –0.219a

Glutathione + 2H+ + 2e– 2 reduced glutathione –0.23

S + 2H+ + 2e– H2S –0.243

Lipoic acid + 2H+ + 2e– dihydrolipoic acid –0.29

NAD+ + H+ + 2e– NADH –0.320

NADP+ + H+ + 2e– NADPH –0.324

Acetoacetate + 2H+ + 2e– β-hydroxybutyrate –0.346

α-Ketoglutarate + CO2 + 2H+ + 2e– isocitrate –0.38

2H+ + 2e– H2 (at pH 7) –0.414

Ferredoxin (Fe3+) + e– ferredoxin (Fe2+) –0.432

Source: Data mostly from R. A. Loach, in Handbook of Biochemistry and Molecular Biology, 3rd edn (G. D. Fasman, ed.), Physical and Chemical Data, Vol. 1, p. 122, CRC Press, 1976.aThis is the value for free FAD; FAD bound to a specific flavoprotein (e.g., succinate dehydrogenase) has a different E'˚ that depends

on its protein environment.

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, hydridefrom an alcohol is transferred to NAD+, giving NADH.

NAD and NADP Are Common Redox Cofactors

Formation of NADH Can Be Monitored

by UV-Spectrophotometry

• Measure the change of absorbance at 340 nm

• Very useful signal when studying the kinetics of

NAD-dependent dehydrogenases

Flavin Cofactors Allow Single Electron Transfers

• Permits the use of molecular oxygen as an ultimate electron acceptor– flavin-dependent oxidases

• Flavin cofactors are tightly bound to proteins.