Metabolism:

Key words Metabolism – definition Catabolism and anabolism – definition, example Identify/distinguish structure of coenzymes Identify structure of ATP

What is Metabolism? The study of the biochemical reactions in an organism, including the coordination, regulation and energy needs Definition: Metabolism is the sum total of the chemical reactions of biomolecules in an organism Metabolism consists of catabolism: the breakdown of larger molecules into smaller ones; an oxidative process that releases energy anabolism: the synthesis of larger molecules from smaller ones; a reductive process that requires energy Catabolism: the oxidative breakdown of nutrients Anabolism: the reductive synthesis of biomolecules

Terminology in Metabolism Metabolic pathway: A sequence of reactions, where the product of one reaction becomes the substrate for the next reaction. - either linear pathway or cyclic pathway - metabolic pathways proceed in many stages, allowing for efficient use of energy Metabolites: intermediates in metabolic pathway light Eg. 6 CO2(g) + 6 H2O(l) → C6H12O6(aq) + 6 O2(g) Anabolism photosynthesis C6H12O6 (aq) + 6O2 (g) → 6CO2 (g) + 6H2O Catabolism respiration

Metabolic pathway

Metabolic pathway: linear or cyclic

A Comparison of Catabolism and Anabolism • Metabolism is the sum total of the chemical reactions of biomolecules in an organism

Metabolism Metabolism involves the energy flow in the cell Photoautotroph via photosynthesis transfers the energy to heterotrophs Heterotrophs obtain the energy through oxidation/reduction of organic compounds (carbohydrate, lipid and proteins)  Food supplies the energy Energy = ATP

The Role of Oxidation and Reduction in Metabolism Oxidation-Reduction (redox) reactions are those in which electrons are transferred from a donor to an acceptor oxidation: the loss of electrons; the substance that loses the electrons is called a reducing agent reduction: the gain of electrons; the substance that gains the electrons is called an oxidizing agent Carbon in most reduced form- alkane Carbon in most oxidized form- CO2 (final product of catabolism) Reduced Oxidized

Oxidation and Reduction in Metabolism Reduction – gain e Oxidation – less e Oxidizing agent – e acceptor reducing agent – e donor

Metabolism: Features Metabolic pathway: Enzymes – multienzymes A group of noncovalently associated enzymes that catalyze 2 or more sequential steps in metabolic/biochemical pathway Metabolic pathway: Enzymes – multienzymes Coenzymes ATP – produced or used Regulation of metabolic pathway: Feedback inhibition or Feed-forward activation

Metabolism: Regulation Regulation of metabolic pathway: Feedback inhibition = product (usually ultimate product) of a pathway controls the rate of synthesis through inhibition of an early step (usually the first step) A  B  C  D  E  P Feed-forward activation = metabolite produced early in pathway activates enzyme that catalyzes a reaction further down the pathway E1 E2 E3 E4 E5 — E1 E2 E3 E4 E5 +

Coenzymes Coenzymes in metabolism: NAD+/NADH NADP+/NADPH FAD+/FADH2 Coenzyme A (CoASH) – activation of metabolites Electron carriers

NAD+/NADH: An Important Coenzyme Nicotinamide adenine dinucleotide (NAD+) is an important coenzyme Acts as a biological oxidizing agent The structure of NAD+/NADH is comprised of a nicotinamide portion. It is a derivative of nicotinic acid NAD+ is a two-electron oxidizing agent, and is reduced to NADH Reduced form, NADH carries 2 electrons

NADP+/NADPH: Also comprised of nicotinamide portion Nicotinamide adenine dinucleotide phosphate (NADP+) – oxidizing agent NADPH involves in reductive biosynthesis Differ with NAD+ at ribose (C2 contain a phosphoryl group, PO32- As electron carrier in photosythesis and pentose phosphate pathway Reduced form, NADPH carries 2 electrons Anabolism

The Structures Flavin Adenine Dinucleotide (FAD) FAD is also a biological oxidizing agent FAD – can accept one-electron or two-electron The terminal e acceptor (O2) can accept only unpaired e (e must be transferred to O2 one at a time) FADH carries 1 electron, FADH2 carries 2 electrons

FAD/FADH2 * FADH (semiquinone form) carries 1 electron, FADH2 (fully reduced hydroquinone form) carries 2 electrons * 1 1 Formation of fully reduced hydroquinone form bypass the semiquinone form

Coenzyme A in Activation of Metabolic Pathways A step frequently encountered in metabolism is activation activation: the formation of a more reactive substance A metabolite is bonded to some other molecule and the free-energy change for breaking the new bond is negative. Causes next reaction to be exergonic

Coenzyme A (CoASH) Coenzyme A – functions as a carrier of acetyl and other acyl groups Has sulfhydryl/thiol group Thioester bond CoASH Acetyl-CoA: is a “high-energy” compound because of the presence of thioester bond – hydrolysis will release energy

ATP- high energy compound ATP is essential high energy bond-containing compound Phosphorylation of ADP to ATP requires energy Hydrolysis of ATP to ADP releases energy nucleotide Phosphorylation: the addition of phosphoryl (PO32-) group/Pi (inorganic phosphate)

Metabolism: (2)

ATP- high energy compound ATP is essential high energy bond-containing compound Phosphorylation of ADP to ATP requires energy Hydrolysis of ATP to ADP releases energy nucleotide Phosphorylation: the addition of phosphoryl (PO32-) group/Pi (inorganic phosphate)

The Phosphoric Anhydride Bonds in ATP are “High Energy” Bonds bonds that require or release convenient amounts of energy, depending on the direction of the reaction Couple reactions: the energy released by one reaction, such as ATP hydrolysis, provides energy for another reactions to completion – in metabolic pathway Phosphoanhydride /

Couple reaction: example

Role of ATP as Energy Currency Phosphorylation of ADP requires energy from breakdown of nutrients (catabolism) The energy from hydrolysis of ATP will be used in the formation of products (anabolism)

Metabolism of Carbohydrate Catabolism Anabolism

Major pathways of carbohydrate metabolism. Fig 8.1 3rd ed

Key words Glycolysis, the fate for pyruvate Substrate-level phosphorylation and oxidative phosphorylation

Glycolysis Glycolysis is the first stage of glucose metabolism Glycolysis converts 1 molecule of glucose to 2 units of pyruvate (three C units) and the process involves the synthesis of ATP and reduction of NAD+ (to NADH) The pathway has 10 steps/reactions Glycolysis are divided into 2 stages/phases, Phase 1=1st 5 reactions Phase 2=2nd 5 reactions Linear pathway

Glycolysis has a net “profit” of 2 ATP per glucose Glycolysis are divided into 2 stages/phases, Phase 1=1st 5 reactions Energy investment – A hexose sugar (glucose) is split into 2 molecules of three-C metabolite (glyceraldehyde-3-phosphate = GAP). The process consume 2 ATP Phase 2=2nd 5 reactions Energy recovery – The two molecules of GAP are converted to 2 molecules of pyruvate with the generation of 4 ATP and 2 NADH. Overall equation – Glucose + 2 NAD+ + 2 ADP + 2Pi  2 pyruvate + 2 NADH + 2 ATP + 2 H2O + 4H+ Glycolysis has a net “profit” of 2 ATP per glucose

The Reactions of Glycolysis glucokinase 1 Phosphorylation of glucose to give glucose-6-phosphate Isomerization of glucose-6-phosphate to give fructose-6-phosphate Phosphorylation of fructose-6-phosphate to yield fructose-1,6-bisphosphate Cleavage of fructose-1,6,-bisphosphate to give glyceraldehyde-3-phosphate and dihydroxyacetone phosphate Isomerization of dihydroxyacetone phosphate to give glyceraldehyde-3-phosphate – isomerase enzyme Use ATP 2 Use ATP 3 phosphofructokinase 4 5

The Reactions of Glycolysis (Cont’d) Glyceraldehyde-3-P dehydrogenase Oxidation of glyceraldehyde-3-phosphate to give 1,3-bisphosphoglycerate Transfer of a phosphate group from 1,3-bisphosphoglycerate to ADP to give 3-phosphoglycerate Isomerization of 3-phosphoglycerate to give 2-phosphoglycerate Dehydration of 2-phosphoglycerate to give phosphoenolpyruvate Transfer of a phosphate group from phosphoenolpyruvate to ADP to give pyruvate oxidation 6 Electron acceptor – NAD+ transfer 7 Phosphorylation of ADP to ATP 8 isomerization dehydration 9 transfer 10 Phosphorylation of ADP to ATP

By kinase enzyme at step 1, 3, 7 and 10 Glycolysis Dephosphorylation of ATP Phosphorylation of ADP Oxidation of intermediates and reduction of NAD+ to NADH by dehydrogenase reactions - step 6 - glyceraldehyde-3-phosphate dehydrogenase By kinase enzyme at step 1, 3, 7 and 10

ATP production ATP is produced by phosphorylation of ADP - is through substrate-level phosphorylation Substrate-level phosphorylation – the process of forming ATP by phosphoryl group transfer from reactive intermediates to ADP 1,3-bisphosphoglycerate and phosphoenolpyruvate – “high-energy” intermediates/compounds Oxidative phosphorylation – the process of forming ATP via the pH gradient as a result of the electron transport chain. Glycolysis - Step 7 and 10

Fates of Pyruvate From Glycolysis Once pyruvate is formed, it has one of several fates In aerobic metabolism- pyruvate will enter the citric acid cycle, end product in aerobic metabolism CO2 and H2O In anaerobic metabolism- the pyruvate loses CO2 produce ethanol = alcoholic fermentation produce lactate = anaerobic glycolysis

Anaerobic Metabolism of Pyruvate Under anaerobic conditions, the most important pathway for the regeneration of NAD+ is reduction of pyruvate to lactate Lactate dehydrogenase (LDH) is a tetrameric isoenzyme consisting of H and M subunits; H4 predominates in heart muscle, and M4 in skeletal muscle In muscle, during vigorous exercise – demand of ATP  but O2 is in short supply  is largely synthesized via anaerobic glycolysis which rapidly generates ATP rather than through slower oxidative phosphorylation

Alcoholic Fermentation In anaerobic bacteria Two reactions lead to the production of ethanol: Decarboxylation of pyruvate to acetaldehyde Reduction of acetaldehyde to ethanol • Pyruvate decarboxylase is the enzyme that catalyzes the first reaction This enzyme require Mg2+ and the cofactor, thiamine pyrophosphate (TPP) • Alcohol dehydrogenase catalyzes the conversion of acetaldehyde to ethanol

NAD+ Needs to be Recycled to Prevent Decrease in Oxidation Reactions

Structure of cell Cytoplasm/ Cytosol

Where does the Glycolysis Take Place? Cytosol Glycolysis is universal!

= Krebs Cycle, Tricarboxylic acid Cycle (TCA) Citric Acid Cycle = Krebs Cycle, Tricarboxylic acid Cycle (TCA)

Metabolism: Features Metabolic pathway: Enzymes – multienzymes A group of noncovalently associated enzymes that catalyze 2 or more sequential steps in metabolic/biochemical pathway Metabolic pathway: Enzymes – multienzymes Coenzymes ATP – produced or used

Couple reaction: example

Coenzymes Coenzymes in metabolism: NAD+/NADH NADP+/NADPH FAD+/FADH2 Coenzyme A (CoASH) – activation of metabolites Electron carriers

Glycolysis has a net “profit” of 2 ATP per glucose Glycolysis are divided into 2 stages/phases, Phase 1=1st 5 reactions Energy investment – A hexose sugar (glucose) is split into 2 molecules of three-C metabolite (glyceraldehyde-3-phosphate = GAP). The process consume 2 ATP Phase 2=2nd 5 reactions Energy recovery – The two molecules of GAP are converted to 2 molecules of pyruvate with the generation of 4 ATP and 2 NADH. Overall equation – Glucose + 2 NAD+ + 2 ADP + 2Pi  2 pyruvate + 2 NADH + 2 ATP + 2 H2O + 4H+ Glycolysis has a net “profit” of 2 ATP per glucose

Fates of Pyruvate From Glycolysis Once pyruvate is formed, it has one of several fates In aerobic metabolism- pyruvate will enter the citric acid cycle, end product in aerobic metabolism CO2 and H2O In anaerobic metabolism- the pyruvate loses CO2 produce ethanol = alcoholic fermentation produce lactate = anaerobic glycolysis Glycolysis – in cytoplasm

Key words Definition – citric acid cycle Explain the citric acid cycle Distinguish between glycolysis and citric acid cycle Understand -oxidation – catabolism of lipid

Citric acid cycle Requires aerobic condition Amphibolic (both catabolic & anabolic) Serves 2 purposes: Oxidize Acetyl-CoA to CO2 to produce energy (ATP & reducing power of NADH & FADH2)-involved in the aerobic catabolism of carbohydrates, lipids and amino acids Supply precursors for biosynthesis of carbohydrates, lipids, amino acids, nucleotides and porphyrins

= Tricarboxylic acid Cycle Citric Acid Cycle = Krebs Cycle = Tricarboxylic acid Cycle (TCA)

TCA Circular pathway Two-carbon unit needed at the start of the citric acid cycle The two-carbon unit is acetyl-CoA Involves 8 reactions The overall reaction from 1 acetyl-CoA produce 3 NADH, 1 FADH2, 2 CO2 and 1 GTP (equivalent to 1 ATP)

Pyruvate is converted to Acetyl-CoA – activation of pyruvate Pyruvate dehydrogenase complex is responsible for the conversion of pyruvate to acetyl-CoA Five enzymes in complex Requires the presence of cofactors TPP (thymine pyrophosphate), FAD, NAD+, and lipoic acid and coenzyme A (CoA-SH) The overall reaction of the pyruvate dehydrogenase complex is the conversion of pyruvate, NAD+, and CoA-SH to acetyl-CoA, NADH + H+, and CO2 Oxidation of pyruvate and reduction of NAD+ 3C Pyruvate = pyruvic acid 2C Thioester, high energy compound

Coenzyme A (CoASH) Coenzyme A – functions as a carrier of acetyl and other acyl groups Has sulfhydryl/thiol group Thioester bond CoASH Acetyl-CoA: is a “high-energy” compound because of the presence of thioester bond – hydrolysis will release energy

Features of TCA Circular pathway Mitochondrial matrix Electron acceptor – NAD+ and FAD Circular pathway Two-carbon unit needed at the start of the citric acid cycle The two-carbon unit is acetyl-CoA Involves 8 reactions The overall reaction from 1 acetyl-CoA produce 3 NADH, 1 FADH2, 2 CO2 and 1 GTP (equivalent to 1 ATP) X 2 How about 1 molecule of glucose?

Citric acid cycle - features Oxidation decarboxylation - CO2 leaves at step 3 and 4 Oxidation of intermediates and reduction of NAD+ to NADH by dehydrogenase reactions - step 3, 4 and 8 - isocitrate dehydrogenase - α-ketoglutarate dehydrogenase - malate dehydrogenase Oxidation of intermediates and reduction of FAD+ to FADH2 by succinate dehydrogenase reaction - step 6 Phosphorylation of GDP to GTP – step 5

Where does the Citric Acid Cycle Take Place? In eukaryotes, cycle takes place in the mitochondrial matrix In prokaryotes? Cytoplasm

The Central Relationship of the Citric Acid Cycle to Catabolism TCA involves 8 series of reactions that oxidizes the acetyl group of acetyl-CoA to 2 molecules of CO2 and the energy is conserves in NADH, FADH2 and “high-energy” compound, GTP Acetyl-CoA – synthesize from pyruvate (glycolysis product) Guanosine – Tri-Phosphate

Aerobic catabolism NADH, FADH2 from glycolysis and TCA will enter the Electron Transport Chain (ETC) to produce more ATP (oxidative phosphorylation) 1 NADH = 2.5 ATP, 1 FADH2 = 1.5 ATP ETC take place in mitochondria - inner membrane (eukaryotes) In ETC In prokaryotes? Plasma membrane

Oxidation of Pyruvate Forms CO2 and ATP Aerobic metabolism is more efficient than anaerobic metabolism

Citric acid cycle - amphibolic Amphibolic (both catabolic & anabolic) Serves 2 purposes: Oxidize Acetyl-CoA to CO2 to produce energy (ATP & reducing power of NADH & FADH2)-involved in the aerobic catabolism of carbohydrates, lipids and amino acids Supply precursors for biosynthesis (anabolism) of carbohydrates, lipids, amino acids, nucleotides and porphyrins Replenish TCA- catabolism of amino a. and fatty a. Anabolic pathway Require aerobic condition

Differences between glycolysis & TCA cycle Glycolysis is a linear pathway; TCA cycle is cyclic Glycolysis occurs in the cytosol and TCA is in the mitochondrial matrix Glycolysis does / does not require oxygen; TCA requires oxygen (aerobic)

Lipids are Involved in Generation and Storage of Energy The oxidation of fatty acids (FA)in triacylglycerols are the principal storage form of energy for most organisms Their carbon chains are in a highly reduced form The energy yield per gram of fatty acid oxidized is greater than that per gram of carbohydrate oxidized

Catabolism of Lipids - triacylglycerol Lipases catalyze hydrolysis of bonds between fatty acid and the rest of triacylglycerols Phospholipases catalyze hydrolysis of bonds between fatty acid and the rest of phosphoacylglycerols May have multiple sites of action

Catabolism of fatty acid - -Oxidation -Oxidation: a series of reactions that cleaves carbon atoms two at a time from the carboxyl end of a fatty acid The complete cycle of one -oxidation requires four enzymes/steps Take place in mitochondria matrix Spiral pathway 1 round of -oxidation = yield 1 NADH, 1 FADH2 and 1 acetyl-CoA

METABOLISM REVISION

Catabolism: the oxidative breakdown of nutrients Anabolism: the reductive synthesis of biomolecules Catabolism – features Release energy (ADP ATP) Oxidizing agent (NAD+, FAD) Anabolism – features Use energy (ATP  ADP) Reducing agent (NADH ,FADH2) Metabolism – the sum total of biochemical reaction carried out by organism

Metabolism Metabolism involves the energy flow in the cell Photoautotroph via photosynthesis transfers the energy to heterotrophs Heterotrophs obtain the energy through oxidation/reduction of organic compounds (carbohydrate, lipid and proteins)  Food supplies the energy Energy = ATP

Major pathways of carbohydrate metabolism. Fig 8.1 3rd ed

Glycolysis Linear pathway Glycolysis is the first stage of glucose metabolism Glycolysis converts 1 molecule of glucose to 2 units of pyruvate (three C units) and the process involves the synthesis of ATP and reduction of NAD+ (to NADH) The pathway has 10 steps/reactions Glycolysis are divided into 2 stages/phases, Phase 1=1st 5 reactions Phase 2=2nd 5 reactions

Fates of Pyruvate From Glycolysis Once pyruvate is formed, it has one of several fates In aerobic metabolism- pyruvate will enter the citric acid cycle, end product in aerobic metabolism CO2 and H2O In anaerobic metabolism- the pyruvate loses CO2 produce ethanol = alcoholic fermentation produce lactate = anaerobic glycolysis Glycolysis – in cytoplasm

ATP – energy carrier / an energy transfer agent ATP- high energy compound ATP – energy carrier / an energy transfer agent ATP is essential high energy bond-containing compound Phosphorylation of ADP to ATP requires energy Hydrolysis of ATP to ADP releases energy nucleotide Phosphorylation: the addition of phosphoryl (PO32-) group/Pi (inorganic phosphate)

Coenzyme A (CoASH) Coenzyme A – functions as a carrier of acetyl and other acyl groups Has sulfhydryl/thiol group Thioester bond CoASH Acetyl-CoA: is a “high-energy” compound because of the presence of thioester bond – hydrolysis will release energy

TCA Circular pathway Two-carbon unit needed at the start of the citric acid cycle The two-carbon unit is acetyl-CoA Involves 8 reactions The overall reaction from 1 acetyl-CoA produce 3 NADH, 1 FADH2, 2 CO2 and 1 GTP (equivalent to 1 ATP)

Citric acid cycle - amphibolic Amphibolic (both catabolic & anabolic) Serves 2 purposes: Oxidize Acetyl-CoA to CO2 to produce energy (ATP & reducing power of NADH & FADH2)-involved in the aerobic catabolism of carbohydrates, lipids and amino acids Supply precursors for biosynthesis (anabolism) of carbohydrates, lipids, amino acids, nucleotides and porphyrins Replenish TCA- catabolism of amino a. and fatty a. Anabolic pathway Require aerobic condition

Where does the Citric Acid Cycle Take Place? In eukaryotes, cycle takes place in the mitochondrial matrix In prokaryotes? Cytoplasm