Energy Production by the Cell Overview of Glucose Metabolism The primary energy source utilized by the cell is glucose

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Energy Production by the Cell

Overview of Glucose Metabolism

The primary energy source utilized by the cell is glucose.

The complete breakdown of glucose results in carbon dioxide, water, and energy.

C6H12O6 + O2 → CO2 + H2O + energy

Cellular respiration is the oxidation of organic fuels by molecular oxygen; oxygen thus serves as the final electron acceptor in respiration (Lehninger, 1975).

Glucose is broken down within the cell by way of two series of reactions:

1) Glycolysis

2) Citric Acid Cycle

As the glucose molecule is broken down, energy is released. This energy is captured and converted to a chemical known as ATP in the electron transport chain.

Glycolysis is the breakdown of glucose to pyruvate with the release of energy.

In glycolysis, the six-carbon glucose molecule is broken down into two 3-carbon molecules of pyruvic acid.

Glycolysis takes place in the cytoplasm of cells (outside of the mitochondrion). Glycolysis involves 10 steps, or chemical reactions, each catalyzed by a specific enzyme.

Glycolysis does not require oxygen. Glycolysis may take place in the presence of oxygen (aerobic conditions) or absence of oxygen (anaerobic conditions). In aerobic organisms the pyruvate produced by glycolysis will continue to be broken down as it enters the mitochondrion. There it will be completely broken down to CO2 and H2O. If oxygen becomes unavailable, pyruvate is converted to lactate. Under anaerobic conditions, yeast carries out fermentation, converting pyruvate into ethanol.

Energy released in the breakdown of glucose is recaptured and stored in the form of ATP (adenosine triphosphate). For each molecule of glucose broken down in glycolysis there is a net gain of two ATP Molecules. Two molecules of ATP are used to get the reaction going, but eventually four ATP molecules are produced, resulting in a net gain of two ATP molecules. In addition, four electrons are harvested as NADH that can be used to form ATP by aerobic respiration.

Products of Fermentation, Glycolysis, or Continuation to Citric Acid Cycle

In many microorganisms and in the cells of most higher animals and plants, the pyruvic acid produced by glycolysis under low oxygen conditions is converted into two molecules of the three-carbon lactic acid as sole end product.

In alcoholic fermentation, characteristic of many yeasts, the glucose molecule is broken down into two molecules of the two carbon compound ethanol (C2H5OH) and two molecules of CO2.

If oxygen is available, the pyruvate is converted into acetyl-Coenzyme A and CO2 and enters the Citric Acid Cycle where it is completely broken down into CO2, H2O, and energy.

Citric Acid Cycle (Overview)

If oxygen is available, the breakdown of glucose can continue via the reactions of the Citric Acid Cycle, a form of Aerobic Respiration. These reactions result in the complete breakdown of glucose into carbon dioxide, water, and energy. These reactions take place in the mitochondrion of the cell.

The Oxidation of Pyruvate to Produce Acetyl-CoA

If oxygen is available, the pyruvate that was produced in the reactions of glycolysis can enter the mitochondrion and be broken down by the Citric Acid Cycle to produce additional amounts of energy. The breakdown of pyruvate begins with a decarboxylation reaction in which one of pyruvate’s three carbon atoms is removed and then released in the form of carbon dioxide. This leaves a two-carbon molecule called acetyl. The acetyl is then combined with coenzyme A, forming acetyl Coenzyme A. Also, during the reaction a pair of electrons and one hydrogen ion is picked up by NAD+, reducing it to NADH, and H+ which is released into the solution.

To enter the Citric Acid Cycle, the coenzyme-A is first removed from Acetyl coenzyme-A. The acetyl group then enters a cyclic series of nine reactions inside the mitochondrion. In the first reaction, the acetyl group combines with oxaloacetate forming citrate. The resulting six-carbon molecule then goes through a sequence of electron-yielding oxidation reactions (Raven and Johnson, 2002, p. 169). As the molecule is broken down, two CO2 molecules are removed. In addition, a pair of hydrogens and electrons is removed in four of the reactions. Three of the four pairs of electrons are picked up by NAD+; FAD picks up the other pair. In addition, one ATP molecule is produced directly in the cycle by way of a substrate-level phosphorylation.

The Electron Transport Chain

The electron transport chain is a series of molecules along which electrons flow as they are removed from the previous molecule, which is oxidized, and passed along to the next, which is reduced. As the glucose molecule is broken down electrons are removed. NAD+ and FAD pick up these electrons and bring them to the electron transport chain, which like the Citric Acid Cycle is also located within the mitochondrion. The electrons are passed along from one molecule to another along the length of the electron transport chain. As they move along the chain, the energy in the electrons is released. When the hydrogens and electrons reach the end of the chain, they combine with oxygen, and water is formed. Thereby oxygen is the final acceptor of the hydrogens and electrons that move along the electron transport chain.

Oxidative Phosphorylation

As electrons move along the Electron Transport Chain, proton pumps transport hydrogen ions (H+) across the inner mitochondrial membrane into the intermembrane space. This creates a concentration gradient of the hydrogen ions across the membrane. The concentration gradient contains energy that can be used to produce ATP. The hydrogen ions pass through an enzyme known as ATP synthase, which uses the energy in the hydrogen ion gradient to combine ADP and a phosphate group to produce ATP.

Glycolysis

Glycolysis is the breakdown of glucose to pyruvate with the release of energy.

The Stages of Glycolysis

Phosphorylation of Glucose to Glucose 6-Phosphate

In this step, glucose is phosphorylated by ATP to form glucose 6-phosphate. In this reaction, a phosphoryl group is transferred from ATP to the hydroxyl group on C-6 of glucose. This reaction is catalyzed by the enzyme hexokinase. Although the ultimate purpose of glycolysis is to produce energy, note that in this reaction a molecule of ATP is used.

Isomerization of glucose 6-phosphate to fructose 6-phosphate

The next step in glycolysis is the isomerization of glucose 6-phosphate to fructose 6-phosphate. The six-membered pyranose ring of glucose 6-phosphate is converted into the five-membered furanose ring of fructose 6-phosphate. This reaction is catalyzed by the enzyme phosphoglucose isomerase.

Phosphorylation of fructose 6-phosphate by ATP to fructose 1,6-bisphosphate

A second phosphorylation reaction follows the isomerization step. Fructose 6-phosphate is phosphorylated by ATP to fructose 1,6 -bisphosphate. This reaction is catalyzed by phosphofructokinase. A second molecule of ATP is used in this step.

Cleavage of Fructose 1,6 –bisphosphate to Dihydroxyacetone phosphate and Glyceraldehye 3-phosphate

In this reaction, fructose 1,6 -bisphosphate is split into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate. This reaction is catalyzed by the enzyme aldolase.

Isomerization of Dihydroxyacetone phosphate and Glyceraldehyde 3-phosphate

The further breakdown of the three-carbon sugars can continue from glyceraldehyde 3-phosphate only, and not from dihydroxyacetone phosphate. However, dihydroxyacetone phosphate can be used by converting it into another molecule of glyceraldehyde 3-phosphate. These compounds are isomers. The isomerization of these three-carbon sugars is catalyzed by triose phosphate isomerase. Thus, two molecules of glyceraldehyde 3-phosphate are formed from one molecule of fructose 1,6 –bisphosphate by the sequential action of aldolase and triose phosphate isomerase.

The preceding steps in glycolysis have transformed one molecule of glucose into two molecules of glyceraldehyde 3-phosphate. No energy has yet been extracted. On the contrary, two molecules of ATP have been invested thus far. We come now to a series of steps that harvest some of the energy contained in glyceraldehyde 3-phosphate.

Conversion of Glyceraldhyde 3-phosphate into 1, 3 Biphosphoglycerate

In this reaction, glyceraldehyde 3-phosphate is converted into 1,3 –bisphosphoglycerate (1,3-BPG). The reaction is catalyzed by the enzyme glyceraldehyde 3-phosphate dehydrogenase.

Note that in this reaction, a phosphate group is added to the molecule without breaking down a molecule of ATP. The phosphate group that is used is an inorganic phosphate.

Formation of ATP by Transferring a Phosphate Group from 1, 3 –Bisphosphoglycerate to ADP

In this reaction, a phosphoryl group is transferred from the acyl phosphate of 1,3 –bisphoglycerate to ADP. This reaction produces ATP and 3-phosphoglycerate. The reaction is catalyzed by the enzyme phosphoglycerate kinase. This is the first ATP-generating reaction in glycolysis.

Thus, the outcomes of the reactions catalyzed by glyceraldehyde 3-phosphate dehydrogenase and phosphoglycerate kinase are: Glyceraldehyde 3-phosphate, an aldehyde, is oxidized to 3-phosphoglycerate, a carboxylic acid.

NAD+ is concomitantly reduced to NADH.

ATP is formed from Pi and ADP.

Conversion of 3-phosphoglycerate to 2-phosphoglycerate

In this reaction, 3-phosphoglycerate is converted into 2-phosphoglycerate. The reaction involves the shifting of the phosphoryl group from carbon 3 of 3-phosphoglycerate to carbon 2, forming 2-phosphoglycerate. The reaction is catalyzed by phosphoglycerate mutase.

Dehydration of 2-phosphoglycerate into phosphoenolpyruvate

In this reaction, a molecule of water is removed from 2-phosphoglycerate, converting it into phosphoenolpyruvate. The reaction is catalyzed by enolase.

Conversion of Phosphoenolpyruvate into pyruvate and formation of a second molecule of ATP

In this reaction, pyruvate is formed, and ATP is generated. The transfer of a phosphoryl group from phosphenolpyruvate to ADP is catalyzed by pyruvate kinase.

Energy Yield in the Conversion of Glucose into Pyruvate

The net reaction in the transformation of glucose into pyruvate is

Glucose + 2 Pi + 2 ADP + 2 NAD+ →

2 pyruvate + 2 ATP + 2 NADH + 2 H+ + 2 H2O

Thus, two molecules of ATP are generated in the conversion of glucose into two molecules of pyruvate. The two molecules of ATP that are formed represent a net gain of 2 ATP molecules. Two molecules of ATP are used initially, but eventually four molecules of ATP are produced, resulting in a net gain of 2 ATP molecules. In addition, two molecules of NADH are produced. If oxygen is available, they can be used to generate additional ATP.

Significance of Glycolysis

Glycolysis is an example of an anaerobic fermentation reaction, in which organisms such as bacteria produce energy from organic fuels in the absence of molecular oxygen.

In higher organisms glycolysis is the initial stage of glucose breakdown. It produces pyruvate. If oxygen is available the pyruvate is converted to Acetyl Coenzyme A. This enters the mitochondrion and enters the Citric Acid Cycle. These reactions complete the breakdown of glucose into carbon dioxide, water, and energy. Thus, in higher organisms, glycolysis has become a preparatory pathway for aerobic glucose catabolism.

If oxygen is not available, the pyruvate acid produced by glycolysis is converted into lactic acid. This reaction yields a small amount of energy for short periods when oxygen is not available. For example, if a runner is running a race, the muscles may begin to work so hard that his or her muscles may begin to use up oxygen faster than it can be supplied. As a result, the muscles begin to work under anaerobic conditions. Since oxygen is not available, the pyruvate produced by glycolysis cannot enter the reactions of the Citric Acid Cycle and be broken down completely. Instead, the pyruvate is converted into lactate. This yields a small amount of energy in comparison to the Citric Acid Cycle. The lactate that is produced is a waste product in the muscles. It is believed to cause the feeling of pain that is associated with fatigue. The runner experiences fatigue and must rest. Over a period of time, the lactate enters the blood stream and is carried to the liver. There it is converted back to glucose. Soon the individual has recovered from the fatigue and is ready for new work.

The Oxidative Decarboxylation of Pyruvate to Acetyl –CoA

The oxidative decarboxylation of pyruvate to form acetyl CoA, which occurs in the mitochondrial matrix, is the link between glycolysis and the citric acid cycle.

Pyruvate + CoA + NAD+ → acetyl CoA + CO2 + NADH

The conversion of pyruvate to acetyl-CoA is catalyzed by the pyruvate dehydrogenase complex, which is a large multienzyme complex consisting of three enzymes.

Citric Acid Cycle

The Citric Acid Cycle is the second and final process in the breakdown of glucose.

The Citric Acid Cycle occurs in the mitochondria.

The chemical change produced by the citric acid cycle is that pyruvate is broken down and oxidized to form carbon dioxide and water.

Six molecules of oxygen are used up in the process. This is the way cells utilize oxygen (to carry out the citric acid cycle) and the reason they must have a continuous supply of oxygen to survive.

Enzymes that catalyze the chemical reactions of the citric acid cycle are protein molecules located in the thousands of particles attached to the outer membrane of each mitochondrion.

As pyruvate acid is metabolized in the Citric Acid Cycle pairs of hydrogen atoms and electrons are removed from the molecule. The hydrogens and electrons are transported to the respiratory chain where ATP is produced. For each glucose molecule metabolized via the Citric Acid Cycle, 18 ATPs are produced.

An Overview of the Citric Acid Cycle

The overall pattern of the citric acid cycle is as follows: A four-carbon compound (oxaloacetate) condenses with a two-carbon acetyl unit to yield a six-carbon tricarboxylic acid (citrate). An isomer of citrate is then oxidatively decarboxylated. The resulting five-carbon compound (α –ketoglutarate) is oxidatively decarboxylated to yield a four-carbon compound (succinate). Oxaloacetate is then regenerated from succinate. Two carbon atoms enter the cycle as an acetyl unit and two leave the cycle in the form of two molecules of CO2. An acetyl group is more reduced than CO2, and so oxidation-reduction reactions must take place in the citric acid cycle. In fact, there are four such reactions. Three hydride ions (hence, six electrons) are transferred to three NAD+ molecules, whereas one pair of hydrogen atoms (hence, two electrons) is transferred to a flavin adenine dinucleotide (FAD) molecule. These electron carriers yield nine molecules of adenosine triphosphate (ATP) when they are oxidized by O2 in the electron transport chain. In addition, one high-energy phosphate bond is formed in each round of the citric acid cycle itself.

The Reactions of the Citric Acid Cycle

Acetyl Coenzyme A condenses with Oxaloacetate to form Citrate

The cycle starts with the joining of a four-carbon unit, oxaloacetate, and a two-carbon unit, the acetyl group of acetyl CoA. Oxaloacetate reacts with acetyl CoA and H2O to yield citrate and CoA.

Acetyl-CoA + Oxaloacetate + H2O → Citrate + CoA

This reaction is catalyzed by the enzyme citrate synthase.

Citrate is Isomerized into Isocitrate

Citrate must be isomerized into isocitrate to enable the six-carbon unit to undergo oxidative decarboxylation. The isomerization of citrate is accomplished by a dehydration step followed by a hydration step. The result is an interchange of an H and an OH. In the reaction the citrate is first converted to an enzyme-bound intermediate known as cis-aconitate, which is then converted to isocitrate. The reaction is catalyzed by the enzyme aconitase.

Oxidation and Decarboxylation of Isocitrate to α -Ketoglutarate

We come now to the first of four oxidation-reduction reactions in the citric acid cycle. In this reaction, Isocitrate is oxidized, a pair of hydrogens and electrons are removed from the molecule and picked up by NAD+, which becomes NADH + H+. This reaction results in the formation of an enzyme-bound intermediate known as Oxalosuccinate. Next, a carbon dioxide is removed from oxalosuccinate, reducing the number of its carbons from 6 to 5, and resulting in the formation of α -ketoglutarate. This reaction is catalyzed by isocitrate dehydrogenase.

Isocitrate + NAD+ ? α –ketoglutatate + CO2 + NADH + H+

The Oxidative Decarboxylation of α –Ketoglutarate to Succinyl Coenzyme A

In this reaction a carbon dioxide is removed from α –ketoglutarate converting it to a four-carbon molecule. Coenzyme A is added to the succinate. Also, a pair of hydrogen ions and electrons is removed by NAD+ which becomes NADH + H+. Succinyl CoA is formed.

α –Ketoglutarate + NAD+ + CoA → succinyl CoA + CO2 + NADH + H+

Deacylation of Succinyl – CoA and Generation of a High-Energy Phosphate Bond

In this reaction, Coenzyme A is removed from succinyl CoA. The coenzyme A is attached to the rest of the molecule by way of an energy-rich bond. The energy derived from breaking this bond is used to phosphorylate a molecule of guanosine diphosphate (GDP).

Succinyl CoA + Pi + GDP ? succinate + GTP + CoA

The reaction is catalyzed by the enzyme succinyl CoA synthetase.

Guanosine triphosphate can be used to form a molecule of ATP. In this reaction, a phosphate group from GDP is transferred to ADP to form a molecule of ATP. The reaction is catalyzed by nucleoside diphosphate kinase.

GTP + ADP ? GDP + ATP

This is the only molecule of ATP that is produced directly in the cycle.

Oxidation of Succinate to Fumarate

In this reaction, succinate is oxidized to fumarate. The reaction is catalyzed by the enzyme succinate dehydrogenase. The hydrogen acceptor is FAD rather than NAD+. In succinate dehydrogenase, the flavin adenine dinucleotide (FAD) is covalently bound to the enzyme (denoted E-FAD). In the reaction two hydrogen atoms are removed from succinate and transferred to FAD, converting it to FADH2. Succinate is thereby converted to fumarate.

Succinate + E-FAD ? Fumarate + E-FADH2

The Hydration of Fumarate to L-Malate

The next step is the hydration of fumarate to form L-malate. The enzyme fumarase catalyzes the addition of H and OH to one side of the double bond of fumarate. This results in the formation of the L isomer of malate.

Fumarate + H2O ? L-Malate

Oxidation of Malate to Oxaloacetate

In the last reaction of the cycle, Malate is oxidized to form Oxaloacetate. This reaction is catalyzed by the enzyme malate dehydrogenase. In the reaction, NAD+ accepts a hydrogen ion and two electrons, which are equivalent to a hydride ion from Malate. The other hydrogen ion appears in the solvent as a proton (H+). Oxaloacetate is formed in the reaction. Notice that this brings us back to the beginning of the cycle. Oxaloacetate can combine with another molecule of acetyl CoA to begin another round of the cycle.

Malate + NAD+ ? Oxaloacetate + NADH + H+

Formation of ATP

As the glucose molecule is oxidized in the reactions of glycolysis, decarboxylation of pyruvate to Acetyl Coenzyme-A, and the Citric Acid Cycle, electrons are removed and picked up by NAD+ and FAD+ forming NADH and FADH2. The electrons are carried to the electron transport chain and travel along a series of electron transport molecules. As the electrons flow along the electron transport molecules, their energy is used to transport protons across the inner mitochondrial membrane, building up a concentration gradient of these ions across the membrane. This gradient can be used as a source of energy to generate the production of ATP. Oxidative phosphorylation is the process in which ATP is formed as a result of the transfer of electrons from NADH or FADH2 to O2 by a series of electron carriers. ATP is produced as the protons flow through an enzyme located in the inner mitochondrial membrane known as ATP synthase. There the energy is used to combine ADP with a phosphate group to form ATP. When the electrons reach the end of the electron transport chain, they combine with oxygen. Therefore oxygen is the final acceptor of the hydrogens and electrons that travel along the electron transport chain. The oxygen combines with hydrogen to form water.

The Electron Transport chain

Electrons are transferred from NADH to O2 through a chain of three large protein complexes called NADH dehydrogenase, bc1 complex and cytochrome oxidase complex.

Electron flow within these transmembrane complexes leads to the pumping of protons across the inner mitochondrial membrane.

The electrons of NADH enter the chain at NADH dehydrogenase. A carrier called ubiquinone then passes the electrons to the bc1 complex.

FADH2 enters the electron transport chain at Ubiquinone. Its electrons are transferred to FeS and then to ubiquinone where they enter the electron transport chain. Because the enzymes that transfer electrons from FADH2 to Q are not proton pumps like NADH reductase, less ATP is produced by the oxidation of FADH2 than from NADH.

As electrons flow through the bc1 complex, H+ ions are transported across the mitochondrial membrane.

Cytochrome oxidase, the last of the three proton-pumping assemblies of the respiratory chain, catalyzes the transfer of electrons from ferrocytochrome c (the reduced form) to molecular oxygen, the final acceptor.

4 Cyt c (+2) + 4 H+ + O2 → 4 Cyt c (+3) + 2 H2O

Four electrons are funneled into O2 reducing it to H2O while transporting protons from the matrix to the cytosolic side of the inner mitochondrial membrane.

oxidation and phosphorylation are coupled by a proton-motive force

The synthesis of ATP is carried out by a molecular assembly in the inner mitochondrial membrane. This enzyme complex is called ATP synthase.

The way that electron transport leads to the production of ATP was explained by the chemiosmotic hypothesis, developed by Peter Mitchell in 1961. According to this model, as electrons travel along the electron transport chain, protons are pumped across the inner mitochondrial membrane to the cytosolic side, producing a concentration gradient of protons that is higher on the cytosolic side and lower on the matrix side. The protons then flow down their concentration gradient across the inner mitochondrial membrane through an enzyme complex known as ATP synthase. The force created by the flow of protons, the proton-motive force, drives the synthesis of ATP from ADP + P by ATP synthase.

electrons from cytosolic nadh enter mitochondria by shuttles

The 2 electrons of NADH captured in glycolysis are brought into the mitochondrion by either of two electron shuttle systems. The first shuttle is the glycerol 3-phosphate shuttle. This shuttle transfers electrons to FAD. The second shuttle is the malate- aspartate shuttle. This shuttle transfers electrons to NAD. The amount of ATP produced depends upon the type of shuttle used. If the glycerol 3-phosphate shuttle is used and the electrons are passed to FAD, only about 1.5 ATPs are produced. If the malate-aspartate shuttle is used and the electrons are passed to NAD, 2.5 ATPs can be produced.

ATP YIELD FROM CELLULAR RESPIRATION

Glycolysis

There is a net gain of 2 ATP molecules from glycolysis.

2 NADH molecules produced by glycolysis generate 3 to 5 ATP molecules (If oxygen is available) depending on the shuttle used.

Pyruvate oxidation yields 5 ATPs

Citric acid Cycle

There are 2 ATP Molecules produced directly in the Citric Acid Cycle.

Oxidative phosphorylation, electron transport and chemiosmosis

6 NADH molecules from the Citric Acid Cycle produce 15 ATPs

2 FADH2 molecules from the Citric Acid Cycle produce 3 ATPs

Total = 30 to 32 ATPs

Study Guide The Production of Energy by the Cell Draft 3

Corrected 9/29/2016

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