Biochemistry Tutorial 1: Digestion and Absorption of Carbohydrates (PDF)

Biochemistry – Winter 2024/1 Tutorial 1: Digestion and Absorption of Carbohydrates Mohamed M. Naguib, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] n.ma...

Biochemistry – Winter 2024/1 Tutorial 1: Digestion and Absorption of Carbohydrates Mohamed M. Naguib, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] [email protected] Tutorial 1 October 14, 2024  Module Assessment ❖ Mock In-class Quizzes ✓ There are no submission requirements. ✓ Part of the homework will be solved/discussed in the tutorial. ✓ Zero grades for in-class quizzes. ❖ Grading System (Two Final Exams) ✓ First Exam (after 5 Weeks) → 50% ✓ Second Exam (after the last Week) → 50% ❑ To pass this module, you are expected to achieve a minimum overall pass grade of 40% for the module. 2  Module Topics Week no. Topic W 01 Classification of Carbohydrates Digestion and Absorption of Carbohydrates W 02 Metabolism of Carbohydrates I (Glycolysis & Kreb’s cycle) W 03 Metabolism of Carbohydrates II (Glycogen Metabolism & Gluconeogenesis) Hormonal Control of Blood Glucose W04 Lipid Structure and Classification Lipids Digestion, Absorption & Transportation in Blood W 05 Lipid Metabolism (β-Oxidation of Fatty Acids & Ketone Bodies Metabolism) W 06 Classification of Amino acids & Protein Structure Protein Digestion & Absorption W 07 Enzymes I: Basic Concepts Clinical Enzymology W 08 Enzyme Kinetics II: Factors Affecting Enzyme Activity Water & Electrolyte Balance W 09 Water & Fat-Soluble Vitamins W 10 Nucleic Acids Acid-Base Balance 3  Biochemistry – Definition ✓ Biochemistry is a sub-discipline of both Biology and Chemistry. It can be defined as the Chemistry of Life. Carbohydrates Monosaccharides (Sugars) Living Organism Macromolecules Amino acids Proteins (Polymers) Others Water Fatty acids Lipids (Organic acids, & Glycerol (Fats) Wastes, Non- Vitamins Minerals and protein amino Electrolytes Nucleic Acids (DNA and RNA) Nucleotides acids) Micromolecules 4 Lecture 1 The First Class of Biomolecules Is Carbohydrates 5  Carbohydrates – Sources The Main sources of carbohydrates are either from Plant or Animal. A. Plant Source B. Animal Source In Plants, carbohydrates are formed via Animals use plant carbohydrates in photosynthesis process in which carbon dioxide and water the formation of their own are transformed into glucose that is can be; carbohydrates, such as; ✓ Used in the synthesis of other monosaccharides Such ✓ Excess glucose is stored as as fructose and galactose. Glycogen (in liver and muscles). ✓ Used in the synthesis of disaccharides (2 ✓ Glucose and galactose is used to Monosaccharides) Such as maltose and sucrose. form Lactose (in milk). ✓ Used in the synthesis of cellulose (polymer of glucose) in plant cell walls. ✓ Stored in plant in form of starch (polymer of glucose). 6 Lecture 1  Carbohydrates in Diet: Classification The Major Source of Carbohydrates is Found in Plants Dietary Carbohydrates Polysaccharides Disaccharides Monosaccharides Starch Glycogen Cellulose Maltose Sucrose Lactose Galactose Glucose Fructose Brain Grape Fruit The storage The storage A structural Malt sugar Cane Milk sugar sugar sugar sugar carbohydrat carbohydrates carbohydrate that (table) that es in plants in animal in that is involved consists of sugar that consists of (inside the liver and in the formation 2 glucose consists of Glucose chloroplasts muscles of cell walls of molecules. Glucose and Doesn’t need digestion as starch (animal starch) plant cells and Galactose. granules) (dietary fibers) Fructose. 7  Carbohydrates Digestion Digestion Process It is the process by which foods (Carbohydrates, Proteins, Lipids) are broken down into their monomers (Monosaccharides, Amino acids, Fatty acids and Glycerol respectively) to facilitate their absorption. It occurs mainly in Mouth & Duodenum. Dietary Carbohydrates Digestible Non-Digestible - Chief source of Energy (4 C/g). - Non-Calorigenic ** Examples: - They exert various beneficial - Starch & Glycogen. effects on the body such as; - Sucrose, Lactose & Maltose. promoting normal gut motility, Preventing constipation, improves glycemic level control in diabetic patients. ** Examples: - Dietary Fibers (Cellulose). - Pectins & Gums. 8  Digestion: Sites & Enzymes ✓ Digestion of carbohydrates occurs at 3 Sites (Mouth, Duodenum, and Upper jejunum of Small Intestine) with the aid of Digestive Enzymes (Glycosidases), which help to speed up hydrolysis of carbohydrates, namely: 1. Salivary Amylase (secreted by Salivary glands). 2. Pancreatic Amylase (secreted by Pancreas). 3. Intestinal Brush Border Enzymes (special enzymes found in microvilli of the Small Intestine that complete digestion). 9  Digestive Enzymes Action 1. Salivary Amylase Salivary Amylase Starch & Glycogen Dextrin + Maltotriose + Maltose & Isomaltose 2. Pancreatic Amylase Pancreatic Amylase Starch & Glycogen Limit Dextrin + Maltotriose + Maltose & Isomaltose 3. Intestinal Brush Border Enzymes (5 Enzymes) Maltase Maltotriose or Maltose 3 Glucose or 2 Glucose Isomaltase Isomaltose 2 Glucose Lactase Lactose Glucose & Galactose Sucrase Sucrose Glucose & Fructose Dextrinase Dextrin & Limit Dextrin Glucose molecules 10  Digestion: Process & End Products - Digestion starts in Mouth, stops in Stomach, the completed in Duodenum & Upper Jejunum of Small Intestine as Follows. Site Mouth Stomach Duodenum Upper Jejunum Dietary Salivary Pancreatic Maltase Isomaltase Lactase Sucrase Dextrinase X carbohydrates Amylase Amylase Starch Dextrin No Limit Dextrin Glucose & Maltotriose digestion. Maltotriose 3 Glucose molecules Glycogen Maltose/ Amylase Maltose/ 2 Glucose Isomaltose stops due Isomaltose 2 Glucose No Digestion to the Due to lack of cellulase enzyme in humans Cellulose Cellulose high Lactose acidity (pH Glucose & 1.5-2.5). Galactose Sucrose Glucose & Fructose Glucose & No Digestion As no need for digestion (Monosaccharides) Glucose & Fructose Fructose - End products of carbohydrate digestion are Monosaccharides (Glucose, Fructose, & Galactose) and Undigested Cellulose. 11  Carbohydrates Absorption Absorption Process It is the process of taking the products of digestion (monomers) across the gastrointestinal (GI) tract walls and into circulation (blood or lymph). It occurs in Ileum. ✓ By the end of digestion, monosaccharides are readily absorbed through the intestinal mucosal cells into the blood stream. Mechanisms responsible for the absorption of monosaccharides: 1. Active Transport, along with sodium by SGLT1 (sodium glucose transporter) against concentration gradient (i.e. from lower to higher concentration that needs ATP) → e.g. Glucose & Galactose. 2. Facilitated (Passive) Transport, by GLUT5 (Glucose transporter 5) with concentration gradient (i.e. from higher to lower concentration) → e.g. Fructose. Brush border (luminal) membrane Baso-lateral membrane  Brain Storming ! Question: Why Glucose has been added to Oral Rehydration Therapy? The answer lies in the transport mechanism (absorption) of glucose. Cause glucose & sodium share the same transport mechanism (Co-transport); Where this glucose allows the intestine to absorb sodium, which in turn drags water to the intestinal cells and hence to the body. This, in turn, is useful in the treatment of diarrhea. 13  Clinical Significance 1. Disorder of Digestion 2. Disorder of Absorption Lactose Intolerance SGLT Deficiency Cause: Lactase Deficiency Cause: Congenital Deficiency of SGLT (Sodium Glucose Transporter). Clinical Features: Diarrhea due to Osmotic Effect of lactose - Clinical Features: Bloating due to Diarrhea. Fermentation of lactose. Management: Avoid milk intake- Yogurt is added to diet. Figure: Lactose Intolerance 14 THE END  Biochemistry – Winter 2024/1 Tutorial 2:Classification of Carbohydrates Presented by Mohamed M. Naguib, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] [email protected] Tutorial 2 October 14, 2024 Tutorial-1: Classification of Carbohydrates ❑ Tutorial Contents: A. Carbohydrates: Definition, Origin, & Functions: B. General Carbohydrate Classification: C. Monosaccharide: Classification & Nomenclature. Structural Aspects of Monosaccharides. Structural Representations of Monosaccharides. D. Oligosaccharides: Dissaccharides, Glycosidic Linkage Formation, and Examples. Trisaccharides. E. Polysaccharides: Classification (Homopolysaccharides & Heteropolysaccharides). Examples. 2 Tutorial -1 First Class of Biomolecules  A. Definition of Carbohydrates Definition: - Carbohydrates consists of Carbon, Hydrogen and Oxygen atoms, with the empirical formula Cn(H2O)n. Hence, they are called Hydrates of Carbon. - Carbohydrates are Polyhydroxy Aldehydes or Ketones or substances that yield such compounds on hydrolysis.  A. Origin of Carbohydrates Origin Carbohydrates are formed by plants where the (CO2) is taken from air and water (H2O) from soil to form sugar in a process called Photosynthesis. Sources Carbohydrates are largely distributed in both plant and animal tissues. Few common types of carbohydrates are milk, bread, popcorn, potatoes, maze, etc. 5  A. Functions of Carbohydrates ✓ Carbohydrates perform numerous roles in living organisms as: 1. Carbohydrates are the dietary sources of energy (4 C/g) for all organisms. 2. These are storage form of energy in the form of glycogen & starch. 3. Carbohydrates are structural components that participate in the structure of cell membranes, cell walls of bacteria, and fibrous cellulose of plants. 4. Cellular recognition by CHO linked to proteins (glycoproteins) e.g. antigens. 6  B. Classification of Carbohydrates Carbohydrates are classified into Monosaccharides, Oligosaccharides, and Polysaccharides as shown in the following figure. 7  B. Classification of Carbohydrates I. Monosaccharides (Simple Sugars; only 1 sugar) 1. Trioses, C3H6O3, e.g. dihydroxyacetone and glyceraldehydes. 2. Tetroses, C4H8O4, e.g. erythrose. 3. Pentoses, C5H10O5, e.g. deoxyribose and ribose. 4. Hexoses, C6H12O6, e.g. galactose, glucose and fructose. II. Oligosaccharides (2-10 sugars) 1. Disaccharides, C12H22O11, e.g. lactose, maltose and sucrose. 2. Trisaccharides, C18H32O16, e.g. maltotriose. 3. Tetrasaccharides …etc. III. Polysaccharides (Complex Sugars; > 10 sugars) 1. Homopolysaccharides, e.g., cellulose, glycogen, and starch. 2. Heteropolysaccharides, e.g., glycosaminoglycans (GAGs). 8  C. Monosaccharides Definition - Defined as carbohydrate consisting of one saccharide unit. - It cannot be further hydrolysed to give simpler units. 9  C. Monosaccharides Common Monosaccharides a- Trioses: The simplest monosaccharises are the two three carbon trioses; Dihydroxyacetone (a ketotriose) and Glyceraldehyde (an aldotriose). b- Hexoses: The most common monosaccharides are hexoses; Glucose (grape sugar), Fructose (fruit sugar), and Galactose (brain sugar). 10  Monosaccharides Classification & Nomenclature A. Classification according to the Placement B. Classification according to the of its carbonyl group (C=O) Number of carbon atoms: o Triose: 3 carbons. ❑ If the carbonyl group is an aldehyde, the o Tetrose: 4 carbons. monosaccharide is an Aldose; C=O at the end o Pentose: 5 carbons. of the chain (e.g. Glucose). o Hexose: 6 carbons. o Heptose: 7 carbons. ❑ If the carbonyl group is a ketone, the monosaccharide is a Ketose; C=O in the middle of the chain (C2) (e.g. Fructose). Nomenclature according to A & B Aldohexose Ketohexose 11  Structural Aspects Monosaccharides Sugars exhibit various forms of isomerism due to their optical properties. Stereoisomeric Terms What are Chiral carbons, Isomers, Enantiomers, and Epimers? 12  C. Monosaccharides Chiral Carbon Atom Chiral carbon centers are carbon atoms that are attached to four different substituents. It is also referred to as asymmetric carbon atoms.  C. Monosaccharides Isomers ✓ Isomers are compounds that have the Same chemical formula but Different structural formula. They all have the same chemical formula C6H12O6 but different structure. 14  C. Monosaccharides Enantiomers: D vs L Designation ✓ A special type of isomerism is found in the pairs of structures that are mirror images of each other. These mirror images are called Enantiomers, and the two members of the pair are designated as D- and L-sugars. These two aldotetroses are enantiomers. They are stereoisomers that are mirror images of each other. 15  Enantiomers (follow) D vs L Designation For sugars with more than one chiral center, D- (Right) or L- (Left) refers to the asymmetric C farthest from the aldehyde or keto group.  C. Monosaccharides Epimers ✓ Stereoisomers that differ only in configuration about One Chiral Center (with the exception of the carbonyl carbon). e.g. Glucose and mannose are C2 epimers 17  C. Monosaccharides Structural Representations of Monosaccharides Long-Chain Structure Ring Structure Fisher projection: Haworth projection: Straight chain Simple ring in representation perspective In Fisher projection, aldehyde or ketone presents at the top, When the chain cyclises (closes) and CH2OH group presents at to make a ring it forms the the bottom. Last Hydroxyl Haworth structure. Most of sugars group to the right are D and to present in solutions in the the left are L sugars. Haworth projection, as it is more stable. 18  C. Monosaccharides Haworth Projections of Monosaccharides 19  D. Oligosaccharides Definition ▪ Carbohydrates that yield 2-10 monosaccharides units on hydrolysis. ▪ According to the number of monosaccharides units present in the molecule, they are further subclassified into disaccharides, trisaccharides, tetrasaccharides, etc. 20  D. Oligosaccharides Disaccharides ✓ Disaccharides are formed by the combination of two monosaccharides by a Glycosidic bond (linkage). ✓General Formula is Cn (H2O)n-1.  D. Oligosaccharides Formation of Disaccharides Glycosidic Bond Formation This bond is formed when a water molecule is removed, (-OH) from the anomeric carbon of one sugar and (–H) from any other carbon of the second sugar. (Condensation Reaction) 22  Common Disaccharides ✓ The most common disaccharides forms are Sucrose, Maltose, Lactose, and Isomaltose. ✓ Each composed of 2 monosaccharides and linked by Glycosidic linkage. ✓ The 2 Numbers preceding the glycosidic bond refers to Location of each of the 2 Carbons involved in the Glycosidic Bond. Formation of Common Disaccharides Basically… Sucrose Glucose + Fructose (linked by 1,2-glycosidic bond) Maltose Glucose + Glucose (linked by 1,4-glycosidic bond) Lactose Galactose + Glucose (linked by 1,4-glycosidic bond) Isomaltose Glucose + Glucose (linked by 1,6-glycosidic bond) 23  D. Oligosaccharides Trisaccharides = 3 Monosaccharides ✓ Plenty of trisaccharides occur free in nature. ✓ Maltotriose is most commonly produced by the action of the digestive enzyme (Salivary Amylase) on the starch amylose. ✓ Maltotriose unit consists of 3 glucose connected via 1,4 glycosidic bond. 24  E. Polysaccharides Definition: They are Complex carbohydrates consisting of more than 10 monosaccharide units connected together to form long chains (branched or unbranched). 25  E. Polysaccharides Classification Polysaccharides are classified into Homopolysaccharides and Heteropolysaccharides according to the Type of monosaccharide units present in the polymer. (e,g. Starch, (e.g. Glycogen, and Glycosaminoglycans; Cellulose) GAGs) 26 E. Homopolysaccharides ✓ The most common homopolysaccharides are Glycogen, Starch, and Cellulose. ✓All Starch, Glycogen, and Cellulose are polymeric Forms of Glucose. ✓ Starch & Glycogen are Storage / Dynamic Forms (They are formed and broken depending on needs). ✓ Cellulose is Structural / Indigestible Form. 27  E. Homopolysaccharides 1. Starch (Storage polysaccharide) - It is the glucose storage polymer in plants. - It is composed of 2 types of glucose polymers, amylose and amylopectin. Amylose is an unbranched glucose polymer with α(1 4) linkages. While, Amylopectin is a branched glucose polymer with mainly α(1 4) linkages, but it also has branches formed by α(1 6) linkages. - The branches occur every 20 residues, produce a compact structure (i.e. store larger quantity in smaller surface area) & provide multiple chain ends at which enzymatic cleavage can occur. 28  E. Homopolysaccharides 2. Glycogen (Storage polysaccharide) - It is the glucose storage polymer in animals, is similar in structure to Amylopectin. But glycogen has more α(1 6) branches. - Branches occur every 10 residues. The highly branched structure permits rapid glucose release (mobilization) from glycogen stores, e.g., in muscle during exercise. Over and above, the importance of branches is to store larger quantity in smaller surface area. - Both glycogen and starch are branched but glycogen (animal) is much highly branched than starch (plant); as 29  Brain Storming …! ✓ What is the advantage of storing polymeric glucose, as starch in plants or glycogen in animals, instead of monomeric glucose ? This is done to not Upset the Osmotic Balances in the Cell. A. Glucose monomers are Soluble in water and thus can cause the cell to become hypertonic. This will result in the entry of excess water within the cells, exert more osmotic pressure and cause cells to lyse. B. On the other hand, Glycogen is Insoluble in water and therefore stays inert. It does not cause any imbalance in osmotic pressure. i.e. Glucose storage in polymeric form (Glycogen) minimizes osmotic effects. 30  E. Homopolysaccharides 3. Cellulose (Structural polysaccharide) - A major structural constituent of plant cell walls, consists of long linear unbranched chains of glucose polymer attached by β (1,4) Cellulose (Fibers) linkages. Cellulose chains are straight & rigid, and pack with a crystalline arrangement in thick bundles – microfibrils (Fibers). - Humans are unable to digest cellulose because they lack appropriate enzymes (cellulase) to break down this complex substance. It is only digested and utilized by ruminants (cows, deers, giraffes, camels) due to presence of microorganisms in their gut that have cellulase enzyme. - These indigestible Cellulose fibres in humans aid the smooth movement of the intestinal tract. Furthermore, rigid cellulose fibres form the cell wall of plant cells. 31 E. Homopolysaccharides 32 E. Heteropolysaccharides 33 THE END  Biochemistry – Winter 2024/1 Tutorial 3: Metabolism of Carbohydrates Presented by Mohamed M. Naguib, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] [email protected] Tutorial 3 October 21, 2024  Tutorial 3: Metabolism of Carbohydrates ❑ Tutorial 3 Contents: I. Carbohydrate Metabolism: Basic Overview II. Carbohydrate Catabolism: 3 Stages of Cellular Respiration. A. Glycolysis - Definition, Types, and Importance. - Glycolytic Pathway. - Stepwise Explanation of Glycolysis (10 Steps). * Energy Investment Phase (1st 5 Steps). * Payoff Phase (last 5 Steps). - Overall Balance Sheet of Glycolysis (Energy Yield in Glycolysis). - Metabolic Fate of Pyruvate. - Clinical Significance of Glycolysis. 2 2  Metabolic Map of Carbohydrates: Basic Overview Carbohydrate metabolism denotes the various biochemical processes responsible for the formation, breakdown and interconversion of carbohydrates in living organisms. The carbohydrate metabolism takes place by 2 main steps: 1- Catabolism. 2- Anabolism. 3 Metabolic Reactions (Definitions)  Carbohydrate Catabolism Stages of Cellular Respiration ✓ One of the major Catabolic reactions inside our body is Cellular Respiration. ✓ Three Main Stages of cellular respiration are; Stage 1: Glycolysis. Stage 2: Krebs Cycle. Stage 3: Electron Transport Chain.  Stage 1: Glycolysis Definition - It is also called Embden-Meyerhof Pathway. - It is defined as the sequence of reactions breaking glucose (6C) into 2 pyruvate (3C) molecules with the production of ATP. - It occurs in the Cytosol.  Glycolysis: Types & Importance Types: - This process occurs either in presence of oxygen (Aerobic glycolysis), hence glucose is catabolized to Pyruvate which then converted into acetyl CoA, OR in the absence of oxygen (Anaerobic glycolysis), hence glucose is catabolized to Pyruvate which then converted into lactate. -Aerobic glycolysis accurs in all tissues like Liver, Kidney, etc. - While, Anaerobic glycolysis occur in RBCs and Muscles. Importance: -Main Pathway of Energy Production. - Energy production during anaerobic glycolysis (e.g. during vigorous exercise).  Glycolytic Pathway 10 Enzyme-Catalyzed Reactions - It takes place in cytosol. Hence prior to these steps are carried out, glucose is transported from extracellular space of the cell into intracellular cytosol by GLUTs (glucose transporters) embedded in the cell membrane. 8 Glycolysis: Two Phases Phase 1 (Energy Investment Phase 2 (Payoff Phase; 4 ATPs Phase; 2 ATPs are consumed). & 2 NADHs are produced).  Step-by -Step Glycolysis (1st reaction) * Step 1: Glucose is phosphorylated at C6 to Irreversible produce Glucose-6-phosphate Reaction (Partially-energized molecule) which is trapped inside the cell. Irreversible step. Catalyzed by Hexokinase. One ATP is consumed.  Step-by -Step Glycolysis (3rd reaction) * Step 3: Fructose-6-phosphate is phosphorylated into Fructose-1,6-bisphosphate (Fully- Irreversible energized molecule) that is unstable and Reaction goes down glycolytic breakdown reactions to release more energy. AMP Irreversible step. Rate-limiting step. Catalyzed by Phosphofructokinase. One ATP is consumed. Rate-Limiting Step  Phase I Energetics 1- First Phosphorylation 2- Isomerization 3- Second Phosphorylation 4- Cleavage  Step-by -Step Glycolysis (last reaction) Irreversible Reaction * Step 10: In this last step, phosphoenol pyruvate is converted into pyruvate. Also, it is called as Substrate level phosphorylation. Ireversible step. Catalyzed by Pyruvate Kinase. 2 ATPs are produced.  Phase II Energetics 5- Phosphorylation & dehydrogenation 6- First ATP Generation 7- Isomeration 8- Dehydration 9- Second ATP Generation  Overall Balance Sheet of Glycolysis (Energy Yield of Glycolysis) * Overall Balance Sheet (Equation) of Glycolysis * *Net Reaction* Glucose + 2 ADP + 2 Pi + 2 NAD+ 2 Pyruvate + 2 ATP + 2 NADH  Metabolic Fates of Pyruvate A. Under Aerobic Conditions: In a link oxidative decarboxylation reaction Pyruvate converts to Alcoholic Acidic Acetyl-CoA then enters krebs Fermentation Fermentation cycle to be finally broken down to CO2 and H2O. B. Under Anaerobic Conditions: Pyruvate undergoes a fermentation reaction and converts to either Lactate (Acidic fermentation) or Ethanol (Alcoholic fermentation). 16  Metabolic Fate of Pyruvate (Anaerobic) There are 2 types of fermentation: A- Acidic Fermentation (Pyruvate converts to Lactate). B- Alcoholic Fermentation (Pyruvate converts to Ethanol). Clinical Significance of Glycolysis 18 THE END  Biochemistry – Winter 2023/4 Tutorial 4: Kreb‘s Cycle Mohamed Mohamed Naguib, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] [email protected] Tutorial 4 October 21, 2024  Carbohydrates Metabolism: Basic Overview Absorbed Glucose in blood Enter the body cells by the Glucose can be aid of glucose transporters stored in liver & (GLUTs) skeletal muscles as glycogen Case 1: Glucose level meets normal cellular Under aerobic condition, requirement. glucose oxidation is completed Splitting of glucose in mitochondria by Kreb’s into pyruvate by cycle Glucose can be Glycolysis stored in adipose (in cytosol) Under anaerobic condition, tissues as glycolysis is followed by triacylglycerol Fermentation (Acidic or (TAG) Alcoholic) 2  Fates of Pyruvate Glucose (6C) Glycolysis (in cytosol) Under Aerobic Condition 2 Pyruvates Under Anaerobic Condition O2 (3C) O2 “Link Reaction” “Acidic Fermentation Reaction” Pyruvate is transported In cytosol, pyruvate is reduced to lactate (3C) into mitochondria via pyruvate transporter by LDH enzyme which requires NADH then it will be oxidized to Acetyl-CoA (2C) in muscles during vigorous exercise Pyruvate Lactate Dehydrogenase (PDH) Dehydrogenase (LDH) Pyruvate Acetyl-CoA (2C) Pyruvate Lactate Oxidative + CO2 Reduction Decarboxylation NAD+ NADH + H+ NADH + H+ NAD+ 3 Complete Stages of Cellular Respiration... Stage Pre-Stage Stage Stage I II II III 4  Stage 2: Kreb‘s Cycle It is also known as Citric acid cycle or Tricarboxylic acid (TCA) cycle. TCA cycle consists of a series of reactions in mitochondria which catalyzes the oxidation of acetyl CoA (2C) to CO2 and H2O in aerobic conditions giving out energy. TCA reactions occur in a cyclic manner and generate large amounts of ATP since the enzymes of the cycle are located in the mitochondria facilitating the transfer of reducing equivalents (NADH and FADH2) from the Kreb’s cycle to the respiratory chain (ETC, electron transport chain), the enzymes of which are also located in the inner mitochondrial membrane. It is the final common pathway by which much of the free energy is liberated during the oxidation of carbohydrate, lipids and proteins through formation of 2 carbon compound acetyl CoA. 5 Reactions of Kreb‘s Cycle Glucose or Fatty acid or Amino acid Oxidative Decarboxylations TCA Cycle Substrate level production of ATP Since A high energy molecule is formed at the substrate level without the involvement of the respiratory 6 Tutorial 22 Tutorial chain.  Energy Production During Kreb‘s Cycle Mechanisms of ATP Generation 1. Substrate-Level 2. Oxidative Phosphorylation Phosphorylation Direct ATP Generation from ADP Indirect ATP Generation from & Pi. Energized Molecules (NADH & FADH2). Takes place during both Glycolysis & Krebs Cycle (Produce total of 4 Most of ATPs generated during ATP/1 Glucose). cellular respiration (3 stages) proceed this mechanism. 7  Biological Significance of Kreb‘s Cycle (Amphibolic Nature) ❑ Citric acid cycle plays a Dual Role and is important both in the oxidation as well as synthetic processes due to its Amphibolic Nature as follows. ✓ It is Catabolic for the oxidation of carbohydrates, lipids and proteins whereby these substances are completely oxidized to CO2 and H2O and release a large amount of energy. ✓ It is also important for the Anabolic reactions as various intermediates of the cycle are used for the biosynthesis of nonessential amino acids, lipids, nucleotides and glucose. 8  Energy Yield of Kreb‘s Cycle Reaction Energy produced as Number of ATPs Isocitrate to α-ketoglutarate NADH + H+ 3 α-ketoglutarate to succinyl CoA NADH + H+ 3 Succinyl CoA to succinate (Substrate level ATP 1 phosphorylation) Succinate to fumarate FADH2 2 Malate to oxaloacetate NADH + H+ 3 Total ATP yield 12 ❑ Since 2 molecules of pyruvate are produced during glycolysis and it yields 2 molecules of acetyl CoA which in one cycle yields 12 ATPs, so 2 acetyl CoA in two cycles will give 24 molecules of ATPs. 9 Energy Yield from Glucose Oxidation (Glycolysis+Krebs Cycle) OR Total ATP yield = 36 or 38 10 Shuttle System: From Cytosol to Mitochondria “There are two NADH shuttles that are responsible for transporting the cytosolic NADH into the mitochondria to enter the oxidative phosphorylation pathway” NADH-Shuttles 1. Glycerol Phosphate-Shuttle 2. Malate Aspartate –Shuttle Function: The glycerol phosphate Function: The malate aspartate shuttle enables the transfer of shuttle results in the regeneration of electrons from NADH generated NADH utilized in the mitochondria and from glycolysis to complex II its subsequent transfer to complex I (coenzyme Q) in the electron in the electron transport chain giving transport chain giving 2 ATP 3 ATP molecules. molecules. Localizatin: Skeletal muscle and Localizatin: Liver, kidney, and the brain utilize glycerol phosphate heart utilize malate aspartate shuttle and liberate 2 ATP from shuttle, and yield 3 ATP per mole of NADH → 36 ATPs from glucose. NADH → 38 ATPs from glucose. 11  Stage 3: Electron Transport Chain (ETC) ✓ Energy-rich molecules, such as glucose, are metabolized by a series of oxidation reactions ultimately yielding carbon dioxide and water (H2O). ✓ The metabolic intermediates of these reactions donate electrons to specific coenzymes, NAD+ and FAD, to form the energy-rich reduced forms, NADH and FADH2. ✓ These reduced coenzymes can, in turn, each donate a pair of electrons to a specialized set of electron carriers, collectively called the electron transport chain (ETC). ✓ As electrons are passed down the ETC, they lose much of their free energy that drives the production of ATP from ADP and inorganic phosphate (Pi). ✓ The coupling of electron transport with ATP synthesis is called oxidative Phosphorylation that proceeds continuously in all tissues that contain mitochondria. 12  ETC: From NADH & FADH2 To ATP Actually, energy yield from oxidative phosphorylation (ETC) is not fully understood yet. In this connection, there are two relative theories that have been emerged to calculate the number of ATPs equivalent to either NADH or FADH2. Conventional Theory Chemo-Osmotic Theory NADH = 3 ATP NADH = 2.5 ATP FADH2 = 2 ATP FADH2 = 1.5 ATP Total ATP yield from glucose oxidation = Total ATP yield from glucose oxidation = 36 or 38 ATP 30 or 32 ATP 13 THE END  Biochemistry – Winter 2024/1 Tutorial 5: Carbohydrates Metabolism (II) Nourhan Ihab Elfar, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] [email protected] Mohamed Mohamed Naguib, PhD Assistant Professor of Biochemistry [email protected] Tutorial 5 November 3, 2024  Gluconeogenesis Definition & Site of Gluconeogenesis: ✓ Gluconeogenesis is the synthesis of glucose or glycogen from noncarbohydrate compounds. The major precursors for gluconeogenesis are lactate, pyruvate, glucogenic amino acids, propionate and glycerol. ✓ Gluconeogenesis occurs mainly in the Cytosol, although some precursors are produced in the mitochondria. ✓ Gluconeogenesis mostly takes place in Liver and, to some extent, in Kidneys. Importance of Gluconeogenesis: 1. Brain and central nervous system, erythrocytes, testes and kidney medulla are exclusively dependent on glucose for continuous supply of energy. 2. Glucose is the only source of supplying energy to the skeletal muscle & RBCs under anaerobic conditions. 2  Gluconeogenesis Reactions of Gluconeogenesis ✓ Gluconeogenesis closely resembles the reversed pathway of glycolysis, although it is not the complete reversal of glycolysis. ✓ Mainly 3 irreversible reactions of glycolysis are different from gluconeogenesis. The rest of the reactions are common for both glycolysis and gluconeogenesis as follows. 3  Gluconeogenesis Vs. Glycolysis The specific The specific enzymes of enzymes of gluconeogenesis glycolysis Hexokinase Glucose-6-phosphatase 1. It is present in liver and kidney. 2. It is absent from muscle and adipose tissue. Phosphofructokinase Fructose-1,6-bisphosphatase Pyruvate kinase Pyruvate carboxylase Phosphoenolpyruvate (PEP) carboxykinase Diagram Demonstrating the 3 Steps that differ in Gluconeogenesis and Glycolysis. 4  Gluconeogenesis from Lactate – Cori Cycle Cori cycle: ✓ It is a metabolic pathway in which lactate Gluconeogenesis produced by Anaerobic Glycolysis in exercising muscles is taken up by liver, oxidized to pyruvate that is eventually converted to glucose (Gluconeogenesis). ✓ Glucose is then released into circulation, returns back to muscles, and is cyclically metabolized. Anaerobic Glycolysis ✓ Cori cycle links Gluconeogenesis with Anaerobic Glycolysis. ✓ Significance: Cori cycle prevents Lactate Acidosis (Toxicity) by converting lactate to pyruvate, that is then converts into glucose. 5  Glycogen Metabolism ✓ If glucose is not needed immediately for ATP production, it is combined with many other molecules of glucose to form a long chain molecule called Glycogen. Glycogen Structure: ✓ Glycogen is a branching glucose polymer that is composed of glucose units linked together by (1→4) glycosidic bonds with more (1→6) branches than that are present in starch. Glycogenin ✓ At the center of glycogen, there is a core protein called Glycogenin that initiates glycogen synthesis. 6  Glycogen Metabolism Glycogen metabolism involves Two metabolic pathways that are; I. Glycogenesis. II. Glycogenolysis. Both of which is finely regulated to achieve blood glucose homeostasis. The synthesis (Glycogenesis) and break down (Glycogenolysis) of glycogen are not simply the reversal of one series of reactions. Instead, each process is an entirely separate metabolic pathway catalyzed by a different set of enzymes as follows. 7  I. Glycogenesis ✓ Glycogenesis is the synthesis of glycogen from glucose. HK Glucose Glucose-6-P ✓ Glycogenesis takes place in well fed state (i.e. when ATP ADP Phosphoglucomutase glucose is high). Glucose-1-P UTP UDP-glucose ✓ It occurs in Cytosol of mainly Liver and Muscle. The phosphorylase liver contains about 4-6% of glycogen as per its weight UDP-Glucose (Glucose donor) i.e., 72-108 g and the muscle contains about 0.7% of + Glycogen primer (n) its weight, i.e., about 245 g. Glycogen synthase UDP ✓ Glycogenesis requires UDP-Glucose (energized glucose donor) & Glycogen primer (Glycogenin + 8 glucose Elongated Glycogen (n+1) residues) that act as glucose acceptor. Repeated steps ✓ Glycogen synthase is the rate limiting enzyme in Elongated Glycogen chain (n+11) glycogenesis. It is responsible for glycogen chain elongation by making α(1,4) glycosidic bond. Branching enzyme ✓ Branching enzyme is responsible for creating branches by making α(1,6) glycosidic bond. Branched Glycogen chain  II. Glycogenolysis ✓ Glycogenolysis is the breakdown of glycogen. Glycogen (n) Glycogen Pi ✓ Glycogenesis takes place in starving state & exercising muscles. Phosphorylase ✓ Liver and Muscles are the major sites of glycogenolysis (Cytosol). + Glucose-1-P + Glycogen (n-1) ✓ The key enzyme of glycogenolysis is Glycogen phosphorylase. Repeated steps ✓ Glycogen phosphorylase catalyzes Phosphorylytic removal of glucose + from glycogen as glucose-1-phosphate. This enzyme hydrolyses the Glucose-1-P + Limit dextrin molecules terminal α(1,4) glycosidic bonds. Debranching enzyme ✓ A stepwise removal of glucose units continues until only four glucose units are left away from α(1,6) branch point; such formed molecule is H2O Debranching known as limit dextrin. enzyme + Free Glucose Debranched Glycogen ✓ Debranching enzyme catalyzes the Hydrolytic removal of free Glycogen glucose unit by splitting α(1,6) glycosidic linkage (Branch point). Phosphorylase (1 : 10) ~ Glucose + Glucose-1-P ✓ Glycogenolysis generates glucose-1-phosphate & free glucose (~ 10:1 Phosphoglucomutase ratio). Glucose-6-P  II. Glycogenolysis: Liver Vs. Muscles Fate of Glucose-6-phosphate Liver Muscles Glucose-6-P Glucose-6-P glycogenolysis No Glucose-6-Phosphatase Glucose-6- Phosphatase Glucose glycogenolysis Cannot be converted into ** Liver glycogen serves as a Glucose & hence, Glucose-6-P source of free glucose molecules is trapped inside muscles that is then exported to circulation for Maintenance of the Anaerobic Glycolysis glycolysis blood glucose concentration particularly between meals & Pyruvate + ATP (Energy) during early stages of fasting. ** Muscle glycogen serves for Energy production (through glycolysis) during muscle contraction (exercising muscles). 10 THE END  Biochemistry – Winter 2024/1 Tutorial 6: Regulation of Blood Glucose Nourhan Ihab Elfar, PhD Nihal Moustafa Mansour, PhD Assistant Professor of Biochemistry Assistant Professor of Biochemistry [email protected] [email protected] Mohamed Mohamed Naguib, PhD Assistant Professor of Biochemistry [email protected] Tutorial 6 November 3, 2024  Regulation of Blood Glucose Level ✓ The blood glucose level is maintained within the normal physiological range of 70-120 mg/100 ml in the post-absorptive state. ✓ It can be varied up and down in normal (non-pathological) states, hence there are various factors that act to regulate blood glucose level. Maintenance of the blood glucose level is regulated at 3 levels 1. Metabolic Level: 3. Hormonal Level: A balance between different 2. Renal Threshold: - Hypoglycemic hormone. metabolic pathways Threshold of the - Hyperglycemic involving various tissues. kidneys (180 mg/dl). hormones. 2  1. Regulation of Blood Glucose By Metabolic Pathways A). Fed State B). Fasting State (Glucose is High) (Glucose is Low) Pathways that Lower blood glucose Pathways that Increase blood glucose 1. After Carbohydrate meal: 1. During early fasting periods (2-6 hrs): *Under aerobic condition *Under anaerobic condition *Only liver glycogen (Not muscle glycogen) Complete glucose Incomplete glucose can be participated in maintaining normal oxidation into oxidation into lactate blood glucose level through (Glycogenolysis). CO2 & H2O. occurs through (Glycolysis). 2. After Carbohydrate rich meal (excess glucose): 2. During Late fasting periods (7-18 hrs; when glycogen stores are exhausted): *Glucose can be stored in liver & skeletal muscles as glycogen via (Glycogenesis). *Glucose can be synthesized in liver and kidney from non-carbohydrate precursors (Gluconeogenesis). 3. If glucose level exceeds storage capacity in liver & muscles: *Glucose can be stored as lipids in adipose tissues via (Lipogenesis) (but stored lipids aren’t converted into glucose in humans). 1. Regulation of Blood Glucose By Metabolic Processes (Summery) Various Metabolic Pathways Regulate Blood Glucose as Follows. ✓ Glycolysis and glycogenesis Lower blood glucose while glycogenolysis and gluconeogenesis Increase blood glucose concentration. ✓ In the fasting state or on a low carbohydrate diet when blood glucose level is reduced, gluconeogenesis and glycogenolysis are stimulated in the liver and add glucose to the blood stream. ✓ In the fed state, when blood glucose concentration increases, its cellular transport in the liver and utilization by the glycolytic reactions is increased. Glycogenesis is also stimulated in the liver and muscle. 4 2. Regulation of Blood Glucose By Kidneys Kidney Threshold (up to 180 mg/dl) ✓ If blood glucose concentration is 70-180 mg/dl, blood glucose is continuously filtered by the glomeruli and is completely reabsorbed by the renal tubules. ✓ However, when blood glucose concentration 180 exceeds 170-180 mg/dl, renal tubules fail to reabsorb all the filtered glucose, and hence glucose starts to appear in urine (Glycosuria). No 5 3. Regulation of Blood Glucose By Hormones Several hormones regulate blood glucose concentration. Hormonal Regulation A. Hypoglycemic B. Hyperglycemic Action Action Insulin is the only Various hormones hormone which lowers increase blood glucose blood glucose level (hyperglycemic (hypoglycemic hormones) such as; hormone). - Glucagon. - Epinephrine. - Glucocorticoids. - Growth hormone. 6  3. Regulation of Blood Glucose By Hormones A. Glucose is High (Hypoglycemic Hormonal Control) A.1. Insulin ✓ Insulin is released into blood stream under the direct influence of Hyperglycemia. ✓ It is produced by the β-Cells of the Islets of Langerhans of the pancreas. ✓ Insulin lowers blood glucose concentration by 4 actions: a-Increasing the uptake of glucose by the extrahepatic tissues for its oxidation. b-Promoting glycolysis and glycogenesis, both in liver and muscles. c-Suppressing glycogenolysis in liver and muscles. d-Suppressing gluconeogenesis in liver and kidneys. 7 3. Regulation of Blood Glucose By Hormones B. Glucose is Low Hyperglycemic Hormonal Control B.1. Glucagon ✓ Its synthesis is stimulated under the influence of Hypoglycemia. ✓ It is produced by the α-Cells of the Islets of Langerhans of the pancreas. ✓ It acts only in the liver and promotes glycogenolysis. ✓ It also promotes gluconeogenesis from lactate and amino acids. 8 Tutorial 6  3. Regulation of Blood Glucose By Hormones B.2. Epinephrine (Adrenaline) ✓ Epinephrine is regarded as the First line of defense against Hypoglycemia. ✓ It is secreted by Adrenal Medulla. ✓ It promotes glycogenolysis in both liver and muscle and promotes gluconeogenesis in liver. ✓ The hormone inhibits the release of insulin from pancreas, thereby, decreasing the transport and utilization of glucose in different tissues. ** Important Note: The Signs of Adrenaline (Epinephrine) Action in response to hypoglycemia are presented in: - Excessive sweating. - Hypertension (High blood pressure). - Increased heart beats. 9 3. Regulation of Blood Glucose By Hormones B.3. Glucocorticoids ✓ Glucocorticoids are the steroids secreted by Adrenal Cortex and are antagonists to insulin. ✓ Cortisol is the major glucocorticoid present in blood. ✓ Glucocorticoids promote gluconeogenesis in liver. ✓ Glucocorticoids also facilitate the action of other hyperglycemic hormones (Permissive Effect) such as glucagon, epinephrine and growth hormone. 10  3. Regulation of Blood Glucose By Hormones B.4. Growth Hormone ✓ Its secretion is stimulated under the influence of Hypoglycemia. ✓ Growth hormone is secreted by Anterior Pituitary Gland. ✓ Growth hormone also decreases glucose uptake and its utilization by the muscle and promotes lipolysis in the adipose tissue (for energy production). *** Important Note: ✓ Growth hormone stimulate Lipolysis during prolonged fasting as in case of Intermittent Fasting. 11 Lecture 3  "Clinical Significance“ Dysregulation of Blood Glucose (Abnormal Elevation of Blood Glucose) Diabetes Mellitus... 12  Diabetes Mellitus ✓ Diabetes mellitus (DM) is defined as a state of chronic hyperglycemia caused by a relative or absolute deficiency of insulin. ✓ There are Two Major Clinical Classes of the disease; 1. Insulin dependent (Type 1) diabetes mellitus. 2. Non-insulin dependent (Type 2) diabetes mellitus. 13  Type I Diabetes Mellitus Insulin-Dependent Diabetes Mellitus (IDDM) or Type I DM ✓ Nomenclature: It is called Juvenile-Onset Diabetes because it usually appears in childhood or in younger age group (less than 40 years of age). ✓ Etiology: There is an absolute deficiency of insulin due to a gradual depletion of the β-cells of the pancreas because β-cells are destroyed by an autoimmune process. ✓ Symptoms: Patients with IDDM can usually be recognized by the abrupt appearance of polyuria (frequent urination), polydypsia (excessive thirst), and polyphagia (excessive hunger), often triggered by stress or an illness → These symptoms are usually accompanied by fatigue, weight loss and weakness. ✓ Treatment: Insulin therapy is recommended for most people with type 1 diabetes. 14  Type II Diabetes Mellitus Non-insulin-Dependent Diabetes Mellitus (NIDDM) or Type II DM ✓ Nomenclature: It is called Maturity-Onset Diabetes since this usually occurs in middle-aged people (more than 40 years old), who are often obese. ✓ Etiology: The occurrence of the type II disease is almost completely determined by genetic factors. It is can be produced due to low insulin levels or non- responsive high insulin levels. Inspite of high levels of insulin, glucose levels are poorly controlled because of the lack of normal response to insulin (Insulin Resistance). ✓ Symtoms: NIDDM develops gradually without obvious symptoms. The metabolic alterations are milder than those for the IDDM. ✓ Treatment: Losing weight, eating well and exercising can help manage the disease. If diet and exercise aren't enough to control blood sugar, diabetes medications or insulin therapy may be recommended. 15 THE END