Carbohydrates Metabolism PDF

MLT 212 Taibah University College of Applied Medical Sciences Chapter 3 Medical Laboratories Technology Carbohydrates Metabolism Dr: Saber Eweda 1446 (2025) Objectives After studying t...

MLT 212 Taibah University College of Applied Medical Sciences Chapter 3 Medical Laboratories Technology Carbohydrates Metabolism Dr: Saber Eweda 1446 (2025) Objectives After studying this chapter, the students should be able to: 1. Know the digestion and absorption of carbohydrates 2. Identify the fate of absorbed carbohydrates 3. Undrastand the reactions, enzymes and regulation of glycolysis 4. Explain the enzymatic reactions and energy produced from Krebs cycle 5. Know the the biochemical pathway and importance of HMP shunt 6. Identify the biochemical reactions of glycogenesis and glycogenolysis 7. Expalin the reactions and importance of gluconeogenesis Carbohydrates Digestion Starch, dextrins, Isomaltose Starch Maltose, Lactose, Lactose Sucrose MOUTH -amylase sucrose Cellulose Low pH stops action of salivary amylase STOMACH Pancreatic -amylase PANCREAS SMALL Isomaltose, Maltose INTESTINE Lactose, Sucrose Portal circulation Glucose To LIVER Fructose Enzymes bound to mucosal Galactos cell membrane (isomaltase, e maltase, lactase, sucrase) 3 Cellulose ❖ Cellulose is not digested in the intestine of man and passes in the stool as such. Dietary fibers have several beneficial effects: 4 Absorption ❖ Polysaccharides and oligosaccharides are not absorbable. ❖ Monosaccharides are principally absorbed from the jejunum. ❖ Glucose is absorbed at the rate of about 1g/kg B.W./h. ❖ Absorbed sugar goes through the portal blood to the liver. ❖ The absorption of hexoses is accelerated by thyroxine. ❖ Insulin has no effect on the absorption of sugars. 5  Fate of absorbed sugars A. Uptake by tissues ❖ The uptake of glucose by tissues occurs by facilitated diffusion, i.e. requires a carrier protein known as glucose transporter (GLUT). B. Utilization by tissues Uptake by the tissues: Two types of glucose transporters( GLUT) are recognized: 1. Insulin-independent glucose transporter I. GLUT1 facilitates uptake of glucose by the RBCs. II. GLUT2 helps the rapid uptake and release of glucose by liver and -cell of pancreas. Also present in kidney and intestine. III. GLUT3 present in brain, kidney and placenta. 2. Insulin dependent glucose transporter ❖ GLUT 4 is present in skeletal muscle, heart and fat cells. Insulin promotes the recruitment of these glucose transporters from intracellular pool to cell membrane. Also increases the activity of each transport. Utilization by tissues: The liver converts fructose and galactose into glucose, which undergoes one of the following fates: 1. Oxidation 1. Major pathways: Glycolysis, followed by oxidation of pyruvate to acetyl CoA which enters the Krebs' cycle. 2. The pentose phosphate pathway (HMP). 3. The uronic acid pathway. 2. Conversion to substances of biologic importance as ribose, fructose, galactose, glucuronic acid, aminosugars and amino acids. 3. Storage in the form of 1. Glycogen (Glycogenesis) 2. Triacylglycerols (lipogenesis) 4. Excretion in the urine: excretion by the kidney if it exceeds 180 mg%, called renal threshold.  GLYCOLYSIS ❖ Glycolysis means the degradation of glucose by a series of enzyme-catalyzed reactions to yield: Two molecules of the pyruvate (aerobic glycolysis, in presence of oxygen). Or, Two molecules of lactate (in absence of oxygen, called anaerobic glycolysis or lactic fermintation). ❖ During glycolysis, the free energy released from glucose is conserved in the form of ATP and NADH. ❖ Glycolysis takes place in the cytosol of cells. Glycolysis is of physiological importance in:- 1. Tissues with no mitochondria: RBCs, cornea and lens. 2. Tissues with few mitochondria: testis, leucocytes, retina, skin and GIT. 3. Tissues undergo frequent oxygen lack: skeletal muscle especially during exercise. Glycolysis occurs in ten reactions steps: 1. The first five reactions called the preparatory phase and, 2. The second five reactions steps reaction constitute the payoff phase. During glycolysis, three types of chemical reactions are occur: 1. Degradation of the carbon skeleton of glucose to yield pyruvate. 2. Phosphorylation of ADP to ATP by high-energy phosphate compounds formed during glycolysis. 3. Transfer of a hydride ion to NAD+, forming NADH. ATP Formation Coupled to Glycolysis ❖ Each NADH molecules enter the electron transport chain gives 3 ATP molecules. So, 6 ATP are produced from the 2 NADH. ❖ So, aerobic glycolysis produces 8 ATP molecules (2 ATP + 6 ATP from the 2 NADH) Regulation of glycolysis ❖ The rate of glycolysis in liver is regulated to meet major cellular needs which include: 1. The production of ATP. 2. The formation of building blocks for biosynthetic reactions. 3. To lower blood glucose ❖ one of the major functions of the liver. When blood sugar falls, glycolysis is stopped in the liver to allow the reverse process, gluconeogenesis and vice versa. Enzymes of Glycolysis Hexokinase ………catalyzes irreversible step (regulation point) Phosphoglucose Isomerase Phosphofructokinase …….rate limiting enzyme (the main regulation step) and also catalyzes irreversible step. Aldolase Triosephosphate Isomerase Glyceraldehyde-3-P Dehydrogenase & Phosphoglycerate Kinase Phosphoglycerate Mutase Enolase Pyruvate Kinase ………catalyzes irreversible step (regulation point) In glycolysis, the reactions catalyzed by hexokinase, phosphofructokinase, and pyruvate kinase are effectively irreversible in most organisms. In metabolic pathways, such enzymes are sites of control or regulation of glycolysis. 1- Hexokinase enzyme ❖ Hexokinase is inhibited by its product glucose-6-phosphate. ❖ The liver also contains Glucokinase enzyme which catalyzes the same reaction of hexokinase.  Glucokinase has a high KM for glucose while hexokinase has low Km. Glucokinase is active only at high glucose concentration.  One effect of insulin in liver is activation of transcription of the gene that encodes the Glucokinase enzyme.  Glucokinase is not subject to product inhibition by glucose-6- phosphate. 2- Phosphofructokinase ❖ Phosphofructokinase is the rate-limiting step of the Glycolysis. 1. Phosphofructokinase is allosterically inhibited by ATP.  At low concentration, the substrate ATP binds only at the active site and activate the enzyme.  At high concentration, ATP binds also at a low-affinity to regulatory site, promoting the tense conformation and inhibition of enzyme. 2. Fructose 2,6-bisphosphate (F2,6BP) is a very potent activator of phosphofructokinase (PFK-1). 2. AMP is an allosteric activator for PFK-1 enzyme. 3. Citrate inhibits phosphofructokinase. Citrate in the cytosol is converted to acetyl-CoA for fatty acid and cholesterol synthesis. 3- Pyruvate kinase 1. Both of its own substrate PEP and fructose 1,6- bisphosphate enhance its activity. Thus, glycolysis is driven to operate faster when more substrate is present. 2. ATP is a negative allosteric inhibitor. This accounts for parallel regulation with PFK 1. 3. Alanine, a negative allosteric modulator (inhibitor) Fates of Pyruvate  Lactate Dehydrogenase O O− O O− C NADH + H+ NAD+ C C O HC OH CH3 CH3 pyruvate lactate ❖ Lactate Dehydrogenase catalyzes reduction of the keto in pyruvate to a hydroxyl, yielding lactate, as NADH is oxidized to NAD+. ❖ Cell membranes contain carrier proteins that facilitate transport of lactate. ❖ Skeletal muscles ferment glucose to lactate during exercise, when the exertion is brief and intense.  Pyruvate Alcohol Decarboxylase Dehydrogenase O O− CO2 H NADH + H+ NAD+ H C O C H C OH C O CH3 CH3 CH3 pyruvate acetaldehyde ethanol ❖ Some anaerobic organisms metabolize pyruvate to ethanol, which is excreted as a waste product. ❖ Lactate released to the blood may be taken up by other tissues, or by skeletal muscle after exercise, and converted via Lactate Dehydrogenase back to pyruvate. ❖ Pyruvate may be oxidized in Krebs Cycle (in liver) or converted back to glucose via gluconeogenesis 25  Krebs Cycle (Citric Acid Cycle) Tricarboxylic Acid Cycle (Krebs-Hansselet Pathway) ❖ It is the final common pathway for the oxidation of carbohydrates, amino acids, and fatty acids to CO2 and H2O. ❖ It is the chemical transformation process by which the acetyl- CoA undergoes oxidation to CO2 and H2O. ❖ This oxidation provides energy for the production of the majority of ATP in most animals, including humans. ❖ The cycle occurs totally in the mitochondria (close to the reactions of electron transport) which oxidize the reduced coenzymes 26 (NADH & FADH2) produced by the cycle. ❖ The TCA cycle is thus an aerobic pathway, because O2 is required as the final electron acceptor. ❖ The citric acid cycle also participates in a number of important synthetic reactions. For example, the formation of glucose from the carbon skeletons of some amino acids, and it provides building blocks for the synthesis of some amino acids and heme. ❖ Firstly, Pyruvate is oxidized to acetyl-CoA and CO2 by the pyruvate dehydrogenase (PDH) complex. ❖ This reaction is an oxidative decarboxylation (irreversible oxidation process) in which the carboxyl group is removed from pyruvate as a molecule of CO2 and producing acetyl-CoA. ❖ The NADH formed in this reaction gives up a hydride ion to the respiratory chain ❖ The pyruvate dehydrogenase complex require 5 coenzymes; NAD+, FAD+, CoA, thyamine pyrophosphate (TPP), Lipoate. Reactions of the Citric Acid Cycle 1 2 3 8 7 4 6 5 29  1 1 30  2a Dehydration 1 2 2b Hydration 31  1 2 3 NAD+ NADH 3 Oxidative decarboxylation 32  1 2 3 4 NAD+ NADH 4 Oxidative decarboxylation 33  1 2 3 4 5 5 Substrate-level 34 phosphorylation  1 2 3 4 6 5 6 Dehydrogenation 35  1 2 3 7 7 4 Hydration 6 5 36  1 2 8 3 7 4 NAD+ 6 5 NADH 8 Dehydrogenation 37 ENERGY PRODUCED BY THE TCA CYCLE 38 Regulation of the Citric Acid Cycle NADH GTP, ATP ❖ The pyruvate dehydrogenase enzyme complex (PDH) activity is turned off (inhibited) when fuel is available in the form of fatty acids and acetyl-CoA and when the cell’s [ATP]/[ADP] and [NADH]/[NAD+] ratios are high. ❖ But it is turned on (activated) again when energy demands are high and the cell requires greater flux of acetyl-CoA into the citric acid cycle. ❖ The enzyme also controlled by second level of regulation called covalent protein modification. The PDH complex is inhibited by reversible phosphorylation of a specific Ser residue on one of the enzyme. ❖ The flow of metabolites through the citric acid cycle is under strong regulation. Three factors govern the rate of flux through the Krebs cycle: 1. Substrate availability. 2. Inhibition by accumulating products. 3. Allosteric feedback inhibition of the enzymes that catalyze early steps in the cycle. ❖ Each of the three strongly exergonic steps in the cycle (those catalyzed by citrate synthase, isocitrate dehydrogenase, and α-ketoglutarate dehydrogenase can become the rate-limiting step under some circumstances. These enzymes are regulated as illustrated in the previous diagram. 42 The complete oxidation of glucose yields 38 ATP Glucose (C6H12O2 ) + 6 O2 6 CO2 + 6 H2O + 38 ATP Glucose Glycolysis 2 + (2x3) = 8 ATP 2 pyruvate In the cytosol Inside mitochondria 2 pyruvate Oxidative 2x3 = 6 ATP decarboxylation 2 Acetyl CoA 2x12= 24 ATP Total 38 ATP Krebs' cycle 43  Pentose Phosphate Pathway ❖ It is the oxidation of glucose 6-phosphate to pentose phosphates. It occur in the cytoplasm of cells. It is an alternative pathway for glucose oxidation where ATP is neither produced or utilized. pentose phosphate pathway has 2 phases The non-oxidative phase of the pathway ❖ Transketolase enzyme catalyzes the transfer of a two-carbon fragment from a ketose donor to an aldose acceptor. ❖ Transaldolase enzyme catalyzes the removal of three-carbon fragment from ketose and condensed with aldose. 47 Glycogen synthesis (Glycogenesis) ❖ Glycogen synthesis takes place in virtually all animal tissues but is especially prominent in the liver and skeletal muscles. ❖ The starting point for synthesis of glycogen is glucose 6-phosphate derived from free glucose by hexokinase enzyme. ❖ The glucose 6-phosphate is converted to glucose 1-phosphate by phosphoglucomutase enzyme. ❖ The glucose-1-phosphate is converted to UDP-glucose by the action of UDP-glucose pyrophosphorylase. This step is the key step (rate limiting step) for glycogen synthesis. ❖ The glycogen synthase catalyzes the transfer of the glucose residue from UDP-glucose to a non-reducing end of a branched glycogen molecule. ❖ Glycogen synthase cannot make the (α 1-6) bonds found at the branch points of glycogen. ❖ These branches are formed by the glycogen-branching enzyme (glycosyl- (4- 6)-transferase). ❖ The glycogen-branching enzyme catalyzes transfer of a terminal fragment of 6 or 7 glucose residues from the nonreducing end of a glycogen branch having at least 11 residues to the C-6 hydroxyl group of a glucose residue at a more interior position of the same or another glycogen chain (thus creating a new branch). ❖ Further glucose residues may be added to the new branch by glycogen synthase. 51  Glycogen Breakdown (glycogenolysis) ❖ In skeletal muscle and liver, the glucose units of the outer branches of glycogen are cleaved and enter the glycolytic pathway by action of three enzymes: glycogen phosphorylase, glycogen debranching enzyme, and phosphoglucomutase. ❖ Glycogen phosphorylase catalyzes the cleavage of (α1-4) glycosidic linkage between two glucose residues at a nonreducing end of glycogen removing the terminal glucose residue as α-D-glucose 1- phosphate. ❖ Glycogen phosphorylase acts repetitively on the nonreducing ends of glycogen branches until it four glucose residues remain in the branch, then it where its action stops. ❖ Then the debranching enzyme catalyzes the transfer of three glucose residues from the branch to another nonreducing end. ❖ The single glucose residue remaining at the branch point (in α 1-6) linkage) is then released as free glucose by the same enzyme. ❖ Once the branch is removed, glycogen phosphorylase act again. 54 55 56  Gluconeogenesis ❖ It is the formation of glucose from non-carbohydrate sources as lactate, pyruvate, glycerol, some amino acids. This pathway consumes energy. ❖ It is essential for providing the body with glucose during fasting. ❖ It is essential for removing the waste products of other tissues as lactate. ❖ Occur in cytoplasm and mitochondria in liver mainly (90%) and kidney (10%). ❖ Steps involve the reversal of glycolysis but the following enzyme responsible to reverse of the 3 irreversible glycolysis enzymes. 58 59  Illustrative Questions What are the meaning for the following scientific terms (define)? 1. Anaerobic glycolysis 2. Krebs cycle 3. Gluconeogenesis Explain how: 1. The cell regulate the pyruvate dehydrogenase enzyme complex. 2. The complete oxidation of one glucose molecule gives 38 ATP. What are the biological importance of the following pathways: 1. Glycolysis 2. Krebs cycle 3. Hexose monophosphate pathway (HMP) 4. gluconeogenesis Give reasons for the following: 1. GLUT 4 is insulin-dependent glucose transporter. 2. Glycolysis is of physiological importance in RBCs. 3. Phosphofructokinase-1 (PFK-1) is allosterically regulated by ATP. Give two differences between glycolysis and HMP. Regarding the function, what is the difference between liver glycogen and muscle glycogen. 61