Lecture #1 Carbohydrates Metabolism (Glycolysis) PDF

CARBOHYDRATES METABOLISM GLYCOLYSIS DR. OMAIMA ALI 1  CONTENT 1- Introduction to metabolism 2- Digestion of carbohydrates 3- Types of carbohydrates 4- Glycolysis 5- Disorder of glycolysis 6- Regulation of glycolysis...

CARBOHYDRATES METABOLISM GLYCOLYSIS DR. OMAIMA ALI 1  CONTENT 1- Introduction to metabolism 2- Digestion of carbohydrates 3- Types of carbohydrates 4- Glycolysis 5- Disorder of glycolysis 6- Regulation of glycolysis 2 INTRODUCTION TO METABOLISM 3 METABOLISM: Total sum of the chemical reactions happening in a living organism. a. ANABOLISM PATHWAYS- Energy requiring biosynthetic pathways b. CATABOLISM PATHWAYS - Degradation of fuel molecules and the production of energy for cellular function c. AMPHIBOLIC PATHWAYS-ACTING As links between the anabolic and catabolic pathways Almost all reactions are catalyzed by enzymes The primary functions of metabolism are: a. Acquisition & utilization of energy b. Synthesis of molecules needed for cell structure and functioning (i.e. Proteins, nucleic acids, lipids, & CHO) 4 c. Removal of waste products  DIGESTION OF CARBOHYDRATES INTRODUCTION: Carbohydrates play a major role in our life and make us healthy. Every cells of human body needs energy to do its work. People don't eat glucose directly, they eat carbohydrates. Then their bodies convert the carbohydrates into glucose for energy and to glycogen for reserve energy. Dietary carbohydrate provided the body's great source of fuel for energy. The carbohydrates are compounds made of carbon (C), oxygen (O) and hydrogen (H). 5 CLASSIFICATION OF CARBOHYDRATES Carbohydrate Simple Complex Monosaccharide Disaccharide Polysaccharide Glucose Sucrose Fructose Maltose Starch 6 Galactose Lactose CARBOHYDRATES DIGESTION In mouth In stomach In intestine 7 In the mouth: Mastication (chewing): the process by which food is crushed and ground by teeth which stimulate the flow of saliva. Digestion of starch begins in mouth: salivary glands secretes saliva into the mouth to moisten the food. The salivary enzyme amylase begins digestion. Starch amylase small polyshcarrides + maltose (disacharride) In the stomach: Stomach acid inactivates salivary amylase, halting (inhibiting) starch digestion. 8 In the small intestine : The major CHO-digesting enzyme is the pancreatic amylase which will continue breaking polysaccharides. The pancreatic amylase in the duodenum breaks down all small polysaccharides into disaccharides. The disaccharide digestion begins at this point. There specific enzymes secreted from the intestinal glands breaks down specific disaccharides. The disaccharide enzymes on the surface of the small intestinal cells hydrolyze the disaccharides into monosaccharaides Maltose maltase Glucose + Glucose Sucrose sucrase Fructose + Glucose Lactose lactase Galactose + Glucose 9 Intestinal cells absorb these monosaccharaides. In the large intestine : Only fibers remain in the digestive tract. Fibers in large intestine attracts water, which softens the stool for passage without straining. Also, bacteria in the GI tract ferment some fibers. Most fiber passes intact through digestive tract to the large intestine. Here, bacterial enzyme digest fibers. Some fibers Bacterial Enzymes short-chain + fatty acids + gas +water 10 Carbohydrate digestion disorders:  Lactose intolerance:  A condition that results from inability to digest the milk sugar lactose, characterized by bloating, gas, abdominal discomfort, and diarrhea. Lactose intolerance differs than milk allergy, which is caused by an immune reaction to the protein in milk. It is a common condition that occurs when there is insufficient lactase to digest the disaccharide lactose found in milk and milk products. Because treatment requires limiting milk intake, other sources of riboflavin, vitamin D , and calcium must be included in diet. 11  Galactosemia:  Is a rare genetic metabolic disorder that affects an individual's ability to metabolize the sugar galactose properly. Lactose in food (such as dairy products) is broken down by the enzyme lactase into glucose and galactose.  In individuals with galactosemia, the enzymes needed for further metabolism of galactose are severely diminished or missing entirely, leading accumulation of galactose. resulting in hepatomegaly (an enlarged liver), cirrhosis, renal failure, cataracts, brain damage, and ovarian failure 12 Carbohydrate absorption: The end products of CHO digestion are monosaccharides: Glucose, Fructose, Galactose. The duodenum and upper jejunum absorb the bulk of the dietary sugars. Glucose is unique that it can be absorbed to some extent through the lining of mouth, but for most part nutrient absorption takes place in the small intestine. Glucose and galactose traverse the cell lining the small intestine by active transport. Fructose is absorbed by facilitated diffusion, which slows its entry and produces a smaller rise in blood glucose. 13 Mechanism of absorption: Insulin is not required for the uptake of glucose by intestinal cells. However, different sugars have different mechanisms of absorption. E.g. galactose and glucose are transported into the mucosal cells by an active, energy-requiring process that requires a concurrent uptake of sodium ions; the transport protein is the sodium-dependent glucose cotransporter 1 (SGLT-1). Fructose uptake requires a sodium-independent monosaccharide transporter (GLUT-5) for its absorption. All three monosaccharides are transported from the intestinal mucosal cell into the portal circulation by yet 14 another transporter, GLUT-2. Fate of glucose A-Oxidative fate: Major pathways: A. Glycolysis. B. Krebs' cycle. Minor pathways: A. Pentose shunt. B. Uronic acid pathway. B- Anabolic fates: Glycogenesis/glycogenolysis. Gluconeogenesis. Glycolipids, glycoproteins and Proteoglycans synthesis. 15 16 Glycolysis overview Site: It occurs in the cell cytosol of all tissues of the body. Classification: It could be aerobic or anaerobic A) Aerobic glycolysis: Pyruvate is the end product of glycolysis in cells with mitochondria and an adequate supply of oxygen. B) Anaerobic glycolysis: Lactate is the end product. It can occur without the participation of oxygen which allows the production of ATP in tissues that lack mitochondria. Glycolysis tissue distribution: 1- Cells devoid of mitochondria: Mature RBCs 2- Contracting muscles: due to occlusion of blood vessels by the muscular contraction that decreases oxygen 3- Tissue with less mitochondria: Kidney (medulla), testicles, leukocytes and white muscle fibers 18 4- Brain and gastrointestinal tract: also normally derive most of their energy from glycolysis. Reactions of Glycolysis:  The break down of glucose to 2 molecules of pyruvate by sequence of 10 reactions which can be divided into: A. Energy investment phase (Preparatory phase) (first 5 steps) B. Energy generation phase (Payoff phase) (last 5 steps) 19 20 1- Phosphorylation  Glucose is activated by phosphorylation to glucose 6-Phosphate by Hexokinase or Glucokinase This reaction is the first irreversible Requires ATP and Mg++ Glucose 6-phosphate is impermeable to cell membrane, so, glucose is trapped within the cell and is further utilized for energy purposes. 21 Differences between hexokinase and glucokinase 2- Isomerization  Glucose 6-Phosphate is isomerized to fructose 6-phosphate by isomerase. This reaction is reversible rearrangement of chemical structure forming ketose from aldose. 23 3- Phosphorylation  Fructose 6-Phosphate is phosphorylated to fructose 1,6-bisphosphate by the enzyme phoshofructokinase (PFK). Energy for this reaction is derived from ATP. PFK is the KEY enzyme of this pathway This is the second irreversible reaction. 24 4- Breakdown  The 6 C molecule fructose 1,6-bisphosphate is splits into two 3 C molecules: * Glyceraldehyde 3-phosphate * Dihydroxyacetone phosphate The enzyme is Aldolase This is reversible reaction. 25 5- Isomerization  Dihydroxyacetone phosphate is isomerized to glyceraldehyde 3-phosphate by the enzyme phosphotriose isomerase. In this reaction 2 molecules of glyceraldehyde 3-phosphate are formed. This is reversible reaction. 26 6- Oxidation  Glyceraldehyde 3-phosphate is dehydrogenated and phosphorylated to 1,3- bisphoshoglycerte (1,3-BPG) catalyzed by Glyceraldehyde 3-phosphate dehydrogenase (G3-P DH). Reducing equivalents are carried by NAD+. This is reversible reaction. This step is important as it is involved in formation of NADH+H + and a high energy 27 compound (1,3-BPG). 7- Substrate level phosphorylation  The energy of 1,3-bisphoshoglycerte (1,3-BPG) is used for the synthesis of ATP in this reaction and produce 3-phoshoglycerate in a reaction catalyzed by 3-phosphoglycerate kinase. This is an example of substrate level phosphorylation. This is reversible reaction. This step is inhibited by Arsenate. 28 8- Shift of phosphoryl group  3-phosphoglycerate is converted to 2-phosphoglycerate by phosphoglycerate mutase enzyme by shifting phosphate group from C3 to C2. This is reversible reaction. 29 9- Dehydration  2-phosphoglycerate is converted into phosphoenol pyruvate (PEP high energy compound) by the enzyme enolase, removing one water molecule. This is reversible reaction. Enolase requires Mg++ for the reaction. Fluoride irreversibly inhibit enolase thereby stops the whole glycolysis. Therefore fluoride is added to blood during estimation of blood sugar. 30 10- Substrate level phosphorylation  Phosphoenol pyruvate (PEP) converts into pyruvate by pyruvate kinase enzyme. Here also energy from PEP is used to produce ATP another substrate level phosphorylation Pyruvate kinase is another KEY enzyme. This is the third irreversible reaction. 31 Anaerobic condition  In anaerobic condition, pyruvate is reduced to lactate by lactate dehydrogenase (LDH) enzyme Here reducing equivalents (NADH+H+) is used which are synthesized in 5th step. 32 34 -ATP -ATP +2 NADH+H+ x 3= +6 ATP +2 ATP +2 ATP Aerobic Total= 10-2= 8 ATP Anaerobic total= 2 ATP  BIOENERGETICS OF (OR ENERGY YIELD FROM) GLYCOLYSIS:  Under anaerobic conditions: 1- Total ATP lost = 2 ATP as follows, One ATP in the activation of glucose to glucose-6-phosphate. One ATP in the activation of fructose-6-phosphate to fructose1,6-diphosphate. 2- Total ATP gained = 4 ATP as follows, 2 ATP by substrate level phosphorylation from 1,3-diphosphoglycerate 2 ATP from substrate level phosphorylation from phosphoenol pyruvate. 3- Net ATP gained = 4 ATP gained - 2 ATP lost = 2 ATP for the anaerobic oxidation of one mole of glucose into lactate. 36  Under aerobic conditions: 1- Total ATP lost = 2 ATP. 2- Total ATP gained = 10 ATP are generated as follows, 4 ATP (obtained by substrate level phosphorylation) + 2 NADH.H+ chain (produced from oxidation of glyceraldehyde-3-phosphate) 2 X 3 ATP = 6 ATP, after oxidation in the functioning respiratory 3- Net ATP gained = 8 ATP as follows, 10 ATP – 2 ATP = 8 ATP for the aerobic oxidation of one mole of glucose. 37  GLYCOLYSIS IN RBCS - Glucose is metabolized in RBCs through anaerobic glycolysis (that requires no mitochondria and no oxygen). - One molecule of glucose yields 2 molecules of ATP by one anaerobic glycolytic pathway. - In addition, 2 molecules of lactate are produced. - Lactate is transported to blood & in the liver it is converted to glucose. 38  GLYCOLYSIS IN RBCS 2,3-Bisphosphoglycerate Shuttle: - 1,3-bisphosphoglycerate is converted into 2,3- bisphosphoglycerate (2,3-BPG) by the action of bisphosphoglycerate mutase in RBCs which is then hydrolyzed into 3-phosphoglycerate by phosphatase enzyme without production of high energy phosphate (ATP). - Then 3-phosphoglycerate complete glycolysis. - 2,3-BPG is present in RBCs and promotes the release of oxygen from hemoglobin through its reduction of hemoglobin affinity to oxygen, thus 39 allow oxygen delivery to tissues in case of hypoxia.  DISORDER OF GLYCOLYSIS Pyruvate kinase (PK) deficiency - Inherited disease - The most common enzyme defect in glycolytic pathway - Reduced rate of glycolysis in RBCs & by this way will deprive RBCs of the only means for producing energy. - As a result, hemolytic anemia will be a consequence as RBCs will not be able to keep the biconcave flexible shape which allows it to squeeze through narrow capillaries with an end result of hemolysis (destruction of RBCs). 40  REGULATION (CONTROL) OF GLYCOLYSIS A. Allosteric Regulation. B. Covalent Modification Regulation C. Hormonal Regulation 41 A- Allosteric Regulation The three main key enzymes that regulate glycolysis are hexokinase and glucokinase phosphofructokinase, pyruvate kinase. 1. Regulation of hexokinase Hexokinase is inhibited by glucose 6-phosphate. 2. Regulation glucokinase (GK) It is inhibited indirectly by Fructose-6-P and is indirectly stimulated by glucose 3. Regulation of Pyruvate Kinase (PK) * Inhibited by high levels of ATP. * Activated by fructose 1,6-bisphosphate 4. Regulation of Phosphofructokinase (PFK) * Inhibited by ATP, citrate 42 * Activated by fructose 2,6-bisphosphate, and AMP B- Covalent Modification Regulation of pyruvate kinase(PK) Phosphorylated PK is inactive while dephosphorylated PK is active. form. Glucagon and Epinephrine stimulate Protein kinase A leading to enzyme phosphorylation. Insulin stimulates phosphoprotein phosphatase leading to enzyme dephosphorylation. 43 C- Hormonal Regulation 44  REFERENCES MLA. HARVEY, RICHARD A., PH. D. LIPPINCOTT'S ILLUSTRATED REVIEWS: BIOCHEMISTRY. PHILADELPHIA :WOLTERS KLUWER HEALTH, 2011. 45 46