CHAPTER 6.3. OXIDATIVE DEGRADATION OF GLUCOSE TO CO2

OXIDATIVE DEGRADATION OF GLUCOSE TO CO2 Oxidative degradation of glucose to CO2 and energy production takes place via two important pathways. 1. Glycolysis or Embden- Meyerhoff pathway is the major pathway for the utilization of glucose for the production of energy and is found in the cytosol of all cells. Glycolysis can function under aerobic and anaerobic conditions. Two molecules of pyruvate are produced. Pyruvate is then converted to Acetyl CoA. 2. In the second pathway, Citric acid cycle, acetyl CoA is further oxidized to CO2 and H2O in mitochondria

1. GLYCOLYSIS 0R EMBDEN- MEYERHOFF PATHWAY Glucose is converted into two molecules of pyruvate, chemical energy in the form of ATP is produced, and NADH- reduced coenzymes are produced. This metabolic pathway takes place in almost all cells. All of the enzymes of the glycolysis pathway are found in the extramitochondrial soluble fraction of the cell, the cytosol. They catalyze the reactions under aerobic and anaerobic conditions.   The over all reactions of glycolytic reactions are presented in below. Glycolysis Anaerobic condition Glucose → Pyruvate → Lactic acid Aerobic condition Pyruvate → CO2 + H2O Glycolysis is a ten step process in which every step is enzyme catalyzed. Details of individual steps with in the glycolytic pathway are now considered.

Step1: Phosphorylation of Glucose Glucose enters into the glycolytic pathway by phosphorylation to glucose 6- phosphate, accomplished by the enzyme hexokinase. However, in liver parenchyma cells and in pancreatic islet cells, this function is carried out by glucokinase. ATP is required as phosphate donor, and it reacts as the Mg-ATP complex.

Step 2: Converstion of Glucose 6 Phosphate to Fructose 6-Phosphate Glucose 6 phosphate is converted to fructose 6-phosphate by phosphoglucose isomerase, which involves an aldose-ketose isomerization.

Step 3: Phosphorlyation of Fructose 6-Phosphate to Fructose 1, 6-Bisphosphate This reaction is catalyzed by the enzyme phosphofructokinase to produce fructose 1, 6-bisphosphate from Fructose 6 phosphate. The phosphofructokinase reaction is irreversible under physiologic conditions.

Step 4: Cleavage of Fructose 1,6-Bisphosphate Fructose 1, 6-bisphosphate is split by aldolase (fructose1, 6-bisphosphate aldolase) into two triose phosphates, glyceraldehyde -3-phosphate and dihydroxyacetone phosphate.

Step 5: Inter conversion of triose phosphates Glyceraldehyde-3-phosphate and dihydroxy acetone phosphate are interconverted by the enzyme phosphotriose isomerase. At this stage 2 molecules glyceraldehyde 3-phosphate are formed. Dihydroxyacetone phosphate is also formed from glycerol of fat, which is phosphorylated to glycerol 3 phosphate and then to dihydroxy acetone phosphate

Step 6: Oxidation of Glyceraldehyde 3-Phosphate to 1, 3 Bisphosphoglycerate Glyceraldehyde-3-phosphate is converted to 1,3 bisphosphglycerate by glyceraldehyde 3-phosphate dehydrogenase using NAD+ as the coenzyme. Finally, by phosphorolysis, inorganic phosphate (pi) is added, forming1, 3 bisphosphoglycerate, and the free enzyme. Energy released during the oxidation is conserved by the formation of a high-energy sulfur group that becomes, after phosphorolysis, a high- energy phosphate group in position 1 of 1, 3 bisphosphoglycerate.

Step 7: Transfer of phosphate group from 1, 3 bisphosphoglycerate 1, 3-bisphosphoglycerate is oxidized to 3- phosphoglycerate by phosphoglycerate kinase. This high- energy phosphate is captured as ATP in a further reaction with ADP. Since two molecules of triose phosphate are formed per molecule of glucose undergoing glycolysis, two molecules of ATP are generated at this stage per molecule of glucose.

Step 8: Conversion of 3-Phosphoglycerate to 2-Phosphoglycerate In the next reaction 3-phosphoglycerate is converted to 2-phosphoglycerate by the enzyme phosphoglycerate mutase

Step 9: Dehydration of 2- Phosphoglycerate to Phosphoenolpyruvate The subsequent step is catalyzed by enolase that promotes the reversible removal of a water molecule from 2-Phosphoglycerate to form phosphoenolpyruvate

Step 10: Conversion of Phosphoenol pyruvate to pyruvate The high-energy phosphate of phosphoenol pyruvate is transferred to ADP by the enzyme pyruvate kinase to generate, at this stage, two molecules of ATP per molecule of glucose oxidized and enolpyruvate is formed. Enolpyruvate formed is converted spontaneousny to the keto form pyruvate. This is an irreversible step

Energy production a. Under aerobic condition Under aerobic condition, pyruvate is taken up into mitochondria, and after conversion to acety-CoA is oxidized to CO2 by the citric acid cycle. The reducing equivalents from the NADH+H+ formed in glycolysis are taken up into mitochondria for oxidation. Two triosephosphates are produced from each molecule of hexose metabolized.

Dihydroxyacetone phosphate←→ glyceraldehyde 3 phosphate Isomerase Dihydroxyacetone phosphate←→ glyceraldehyde 3 phosphate glyceraldehyde 3 phosphate dehydrogenase ↓ + NAD+Pi 1,3 diphosphoglyceric acid +NADH+H+ ↓ Phosphoenol pyruvate +ATP Enol pyruvate + ATP Since two molecules of triose phosphates are undergoing the above reaction totally four ATP molecules of produced.

Out of this 2 ATP molecules are used during the initial reactions. The NADH formed by the dehydrogenation of glyceraldehyde 3 phosphate is then rexoidised to NAD+ by O2 via Electron transport chain of mitochondria and produce 3ATP molecules. Since two molecules of glyceraldehyde 3 phosphate are involved in the above reaction six ATP molecules are produced. However, two ATP molecules are used up in the production of glucose-6-phosphate from glucose and fructose-1, 6-disphosphate from fructose-6-phosphate. Hence under aerobic glycolysis, the total number of ATP molecules produced is 10. Out of this 2 ATP molecules are used during the initial reactions. The net ATP production is 8.

b. Under anaerobic condition Pyruvate is reduced by the NADH+H+ to lactate, the reaction being catalyzed by lactate dehydrogenase. The reoxidation of NADH via lactate formation allows glycolysis to proceed in the absence of oxygen by regenerating sufficient NAD+ for another cycle of the reaction catalyzed by glyceraldehyde-3 phosphate dehydrogenase. Lactate dehydrogenase Pyruvate + NADH + H+ ↔ NAD+ + Lactate During fermentation, pyruvate is reduced by NADH to ethyl alcohol being catalysed by alcohol dehydrogenase.   Alcohol dehydrogenase Pyruvate + NADH + H+ ↔ NAD+ + Ethyl alcohol The conversion of two triose phosphates to lactic acid (or ethanol) yields four molecules of ATP. However, two ATP molecules are used up in the production of glucose-6-phosphate from glucose and fructose-1, 6-disphosphate from fructose-6-phosphate. The net production of ATP is thus only two ATP molecules/ mole of glucose

Fates of Pyruvate Three common fates for pyruvate are of prime importance: conversion into acteyl CoA, lactate, and ethanol.

Oxidation of Pyruvate to Acetyl CoA Pruvate derived from glucose by glycolysis, is dehydrogenated to yield acetyl-CoA and CO2 by pyruvate dehydrogenase, located in the mitochondria. CH3COCOOH+NAD++CoA-SH→CH3S~CoA+NADH+ CO2 Pyruvate Acetyl CoA Thiamine pyrophosphate, coenzyme A, lipoic acid and NAD+ are coenzymes needed for the reaction. The acetyl CoA is then fed into citric acid cycle and oxidized to CO2 and water.

2) CITRIC ACID CYCLE The reactions of citric acid cycle the following steps

1. Condensation of acety1- CoA with oxaloacetate to form citrate The initial condensation of acety1- CoA with oxaloacetate to form citrate is catalyzed by condensing enzyme, citrate synthase, which effects synthesis of a carbon to carbon bond between the methyl carbon of acety1-CoA and the carbony1 carbon of oxaloacetate.

2. Conversion of citrate to isocirtrate via cis-aconitate Citrate is converted to isocitrate by the enzyme aconitase (aconitate hydratase), which contains iron in the Fe2+ state in the form of an iron- sulfur protein (Fe:S) This conversion takes place in two steps: dehydration to cis-aconitate, some of which remains bound to the enzyme, and rehydration to iocitrate.

3. Dehydrogenation of isocitrate to oxalosuccinate Isocitrate undergoes dehydrogenation in the presence of isocitrate dehydrogenase to form oxalosuccinate. The linked oxidation of isocitrate proceeds almost completely through the NAD+ dependent enzyme isocitrate dehydrogenase.

4. Decarboxylation of oxalosuccinate to α -ketoglutarate There follows decarboxylation of oxalosuccinate to α -ketoglutarate, also catalyzed by isocitrate dehydrogenase. A CO2 molecule is liberated. Mn2+(or Mg2+) is an important component of the decarboxylation .

5. Decarboxylation of α-ketoglutarate to succiny1-CoA Next, α-ketoglutarate undergoes oxidative decarboxylation to form succinyl CoA The reaction is catalyzed by an α-ketoglutarate dehydrogenase complex, which requires cofactors thiamin pyrophosphote, lipoate, NAD+, FAD and CoA results in the formation of succiny1-CoA, a high- energy thioester and NADH.

6. Conversion of succinyl-CoA to succinate Succinyl-CoA is converted to succinate by the enzyme succinate thiokinase (succiny1CoA synthetase). High-energy phosphate (ADP+Pi →ATP) is synthesized at the substrate level because the release of free energy from the oxidative decarboxylation of α -ketoglutarate is sufficient to generate a high- energy phosphate..

7. Dehydrogenation of succinate to fumarate Succinate is metabolized further by undergoing a dehydrogenation catalyzed by succinate dehydrogenase to form fumerate. It is the only dehydrogenation in the citric acid cycle that involves the direct transfer of hydrogen from the substrate to a flavorprotein without the participation of NAD+.

8. Addition of water to furmarate to give malate Furmarase (furmarate hydratase) catalyzes the addition of water to furmarate to give malate. In addition to being specific for the L-isomer of malate, furmarase catalyzes the addition of the elements of water to the double bond of furmarate in the tans configuration.

9. Dehydrogenation of malate to form oxaloacetate Malate is converted to oxaloacetate by dehydrogenation catalysed by the enzyme malate dehydrogenase, a reaction requiring NAD+. Thus the citric acid cycle is completed. An acetyl group, containing two carbon atoms, is fed into the cycle by combining it with oxaloacetate. At the end of the cycle a molecule of oxaloacetate is generated. These reducing equivalents NADH+H+ and FADH2 are transferred to the respiratory chain in the inner mitochondrial membrane for reoxidation.

Respiratory chain in the inner mitochondrial membrane Pyruvate ADP+Pi→ATP ADP+Pi→ATP ADP+Pi→ATP Isocitrate ↑ ↑ ↑ α-ketoglutarate →NAD→FMN→CoQ→Cytb→CytC1→CytC→Cyta→ Cyta3 Malate

Isocitrate dehydrogenase Respiratory chain oxidation of NADH +H+ 3 ATP production in mitochondria Name of enzyme Reaction Catalyzed ATP molecules formed Isocitrate dehydrogenase Respiratory chain oxidation of NADH +H+ 3 α-Ketoglutrate dehydrogenase Respiratory chain oxidation of NADH +H+ Succinate thiokinase Phosphorylation at substrate level 1 Succinate dehydrogenase Resiphorylation chain oxidation of FADH2 2 Malate dehydrogenase Net 12

ATP production from glucose Glycolysis One mole. of glucose → 2 moles of pyruvate ATP produced = 8 Pyruvate → Acetyl CoA ATP produced = 2x3=6 Citric acid cycle Acetyl CoA →CO2 + H2O ATP produced = 12 x2=24 Net production =30+8=38