OS 201 Human Cell Biology: Metabolism of Carbohydrates - UPCM 2029 PDF

OS 201: HUMAN CELL BIOLOGY Metabolism of Carbohydrates UPCM 2029 | Dr. Albert R. Tiotuyco | LU3 A.Y. 2024-2025 Other Carbohydrate Metabolic Pathways OUTLI...

OS 201: HUMAN CELL BIOLOGY Metabolism of Carbohydrates UPCM 2029 | Dr. Albert R. Tiotuyco | LU3 A.Y. 2024-2025 Other Carbohydrate Metabolic Pathways OUTLINE ○ Fructose Metabolism I. Overview B. Non-oxidative Phase ○ Galactose Metabolism II. Glycolysis C. Significance ○ There are other metabolic pathways since glucose is not the only A. Overview D. Clinical Correlation: carbohydrate. B. Steps of Glycolysis G6PD II. GLYCOLYSIS III. Gluconeogenesis I. Fructose and Galactose A. Overview Metabolism Catabolism or breakdown of glucose to pyruvate (or lactate during B. Steps of A. Fructose Metabolism anaerobiosis) Gluconeogenesis in Liver and Other Key pathway for organisms to extract energy from nutrients C. Diabetic Ketoacidosis Tissues ○ Since it is an energy-producing pathway IV. Glycogen Metabolism B. Galactose Metabolism Two phases: A. Overview C. Clinical Applications ○ Phase 1: “Priming phase” B. Glycogenesis VII. References Prepares the molecule for provision of energy C. Glycogenolysis VIII. Appendix Energy is invested V. Hexose Monophosphate Steps 1-5 Pathway ○ Phase 2: “Energy producing phase” or “oxidation phase” A. Oxidative Phase Energy is released Step 6 onwards Two types: Learning Objectives ○ Aerobic glycolysis At the end of the session, the student should be able to: ○ Anaerobic glycolysis ○ Explain the central role played by glucose in metabolism Site: cytoplasm ○ Describe the steps in glycolysis End product/s: ○ Compare anaerobic glycolysis and aerobic glycolysis ○ Aerobic: pyruvate (substrate, end-products, number of ATPs generated, etc.) ○ Anaerobic: lactate ○ Enumerate the rate limiting steps of glycolysis and their significance in regulation From Batch 2028 Trans: Glycolysis Summary ○ Describe the steps in gluconeogenesis, glycogenolysis and Net synthesis : 2 ATPs glycogenesis 10 steps ○ Explain the Hexose Monophosphate Pathway (HMP) and its ○ 3 irreversible steps/Regulatory Points significance in metabolism Step 1: Phosphorylation of Glucose ○ Describe the metabolism of other carbohydrates (fructose, Step 3: Phosphorylation of F6P galactose) Step 10: Transfer of a Phosphate Group (from PEP to ATP to produce Pyruvate) I. OVERVIEW 3 key rate-limiting enzymes ○ Step 1: Hexokinase ○ Step 3: Phosphofructokinase (PFK-1) ○ Step 10: Pyruvate kinase (PKs) A. AEROBIC GLYCOLYSIS There are a total of 10 steps in (aerobic) glycolysis. STEP 1: PHOSPHORYLATION OF GLUCOSE Figure 2. Phosphorylation of Glucose Figure 1. Overview of Glucose Metabolism. Substrate: Glucose Figure Notes: ATP-requiring step (energy requiring step) Glucose is converted into glucose-6-phosphate via 1/3 of the irreversible regulatory steps in glycolysis phosphorylation with the enzyme, kinase. Priming by phosphorylation at C6 carbon ○ Kinase: enzyme that adds a phosphate group to an original End product: Glucose-6-phosphate (G6P) substance Enzyme: Hexokinase The glucose-6-phosphate is like a key central molecule which can ○ The enzyme is inhibited by G6P undergo different pathways ○ Classic example of product inhibition of an enzyme ○ Glycolysis: glucose-6-phosphate → pyruvate ○ Present in many tissues, notably the muscle cells Glucose can be used to produce energy ○ Can exist in many isoforms (I - IV) Pyruvate Hexokinase IV, also known as glucokinase, present in the liver, ○ End product in aerobic glycolysis has unique properties ○ A very important molecule as it can be made into: Ensures the glucose enters the liver cell and does not go out Amino acids into the bloodstream Acetyl-COA The added phosphate group makes it too big to leave the cell ○ Can enter the citric acid cycle via the transporter ○ Can be used as initial substance in the creation of fatty acids Table 1. SUMMARY OF ENZYMES IN THE NON-OXIDATIVE PHASE OF HEXOSE Lactate MONOPHOSPHATE PATHWAY Ethanol, an alcohol Hexokinase Glucokinase ○ Gluconeogenesis: pyruvate → glucose-6-phosphate Present in all tissues Present in liver & pancreatic β Glucose can also be synthesized from pyruvate cells ○ Glycogen Metabolism Low Km (high affinity) High Km (low affinity) Glycogenesis: glucose-6-phosphate → glycogen Low Vmax High Vmax Glycogenolysis: glycogen → glucose-6-phosphate No effect by insulin Inducible by insulin ○ Alternative Glucose Pathway Substrate specificity: Substrate specificity: Pentose Phosphate Pathway (or Hexose Monophosphate Glucose, Fructose Galactose Primarily glucose, has a very low Pathway): glucose-6-phosphate → ribose-5-phosphate affinity for fructose, but NO ○ Ribose-5-phosphate: sugar found in nucleic acids affinity for galactose Carbohydrates are also critical in nucleic acid metabolism Allosterically inhibited by G6P NOT allosterically inhibited by G6P Trans 2 TG5: Cabatingan, Cabrera, Cadag, Cagayan, Camacho, Camama, Campo TH: Punongbayan 1 of 12  Physiological role: Basal levels of Physiological role: Accumulation STEP 5: ISOMERIZATION OF DHAP G6P for glycolysis and ATP of high intracellular levels of G6P production for conversion to glycogen & TAGs Km is a product concentration in which a reaction is at its maximum velocity. High Km means low affinity in which you need a lot of product for the enzyme to attach. The same is true for the inverse which is low Km. Glucokinase has a high Km while hexokinase has a low Km ○ Low Km in hexokinase enzyme of the muscle cells since a lot of energy is needed for it to function, and any slight depletion in Figure 6. Isomerization of DHAP energy will prompt the hexokinase to attach to the glucose Substrate: DHAP entering the cell to prevent it from going out ○ All DHAP is converted to G3P ○ The muscle cells have the ‘preferential treatment’ in having ○ GAP (G3P) is the substrate for step 6 so all the DHAP is the glucose over the liver cells eventually and readily converted to GAP Insulin has no effect on hexokinase End product: glyceraldehyde 3-phosphate (G3P) ○ Even if you have high or low levels of insulin, if the muscle cells Enzyme: Triose phosphate isomerase need an energy source, glucose enters the muscle cell. This reaction completes the first phase of glycolysis, which is the energy consuming process of the glycolysis. STEP 2: ISOMERIZATION OF G6P STEP 6: OXIDATION of G3P Figure 7. Oxidation of G3P Substrate: G3P Figure 3. Isomerization of G6P End product: NADH, H+, and 1,3-Bisphosphoglycerate (1,3-BPG) Substrate: G6P ○ First energy generating step in glycolysis via oxidative Involves a shift of the carbonyl oxygen from C1 to C2, thus phosphorylation converting an aldose into a ketose ○ NADH + H+ is produced from NAD+ ○ NOT a decarboxylization reaction ○ Requirements: inorganic phosphate group and Carbon was not removed; the shape was just changed oxidized NAD+ Reversible step ○ In this process, NAD+ is reduced to coenzyme NADH by the H– ○ Forward reaction is favored since F6P is readily consumed from glyceraldehydes 3-phosphate. End product: Fructose 6-Phosphate (F6P) ○ Since 2 moles of glyceraldehyde 3-phosphate are formed from Enzyme: Phosphoglucoisomerase one mole of glucose, 2 NADH are generated in this step. Enzyme: Glyceraldehyde-3-phosphate dehydrogenase STEP 3: PHOSPHORYLATION OF F6P NADH is a reducing equivalent that is hydrogen rich and is needed in the electron transport chain. NADH in this step may be transported to ETC or used in anaerobic glycolysis in the conversion of pyruvate to lactate. STEP 7: TRANSFER OF A PHOSPHATE GROUP Figure 4. Phosphorylation of F6P Substrate: F6P ○ Phosphorylated in the 1st carbon ATP-requiring step 2/3 of the irreversible regulatory steps in glycolysis End product: Fructose 1,6-bisphosphate (F-1,6-BP) Enzyme: Phosphofructokinase-1 (PFK1) ○ An allosteric enzyme Figure 8. Transfer of a Phosphate Group ○ Rate-limiting Substrate: 1,3-BPG ○ With substrate level phosphorylation of ADP → ATP Bisphosphate: Phosphate groups are attached to two different ○ Since two moles of 1,3-bisphosphoglycerate are formed from atoms. one mole of glucose, 2 ATPs are generated in this step Biphosphate: Phosphate groups are attached to each other. Energy generating step via substrate level phosphorylation of ADP Ex: ATP End product: 3-Phosphoglycerate (3PG) and ATP ○ The mixed anhydride (a high energy bond) provides the energy needed to produce ATP STEP 4: CLEAVAGE OF F-1,6-BP Enzyme: Phosphoglycerate kinase STEP 8: ISOMERIZATION OF 3-PHOSPHOGLYCERATE Figure 5. Cleavage of F-1,6-BP Substrate: F-1,6-BP ○ 6-carbon sugar F-1,6-BP is converted to 2 three-carbon isomers: Figure 9. Isomerization of 3-phosphoglycerate dihydroxyacetone phosphate (a ketose) Substrate: 3PG glyceraldehyde 3-phosphate (an aldose) ○ Reversible isomerization reaction where the phosphate group is The remaining steps in glycolysis involve three-carbon units, rather transferred from C-3 to C-2 than six carbon units End product: 2-Phosphoglycerate (2PG) End product: Dihydroxyacetone phosphate (DHAP) and Enzyme: Phosphoglycerate mutase glyceraldehyde 3-phosphate (G3P) Enzyme: Aldolase OS 201 Metabolism of Carbohydrates 2 of 12  STEP 9: DEHYDRATION OF 2-PHOSPHOGLYCERATE Figure 10. Dehydration of 2-phosphoglycerate Substrate: 2PG ○ Converted to a high energy molecule ○ Reversible Reaction End product: Phosphoenolpyruvate (PEP) and water ○ A double bond is created between C2 and C3 with the release of H2O Enzyme: Enolase STEP 10: TRANSFER OF A PHOSPHATE GROUP Figure 13. Steps in Glycolysis Figure 11. Transfer of a Phosphate Group Substrate: PEP ○ Converted to a high energy molecule End product: Pyruvate and ATP 3/3 of the irreversible step in glycolysis High energy phosphoenol bond provides energy for the substrate level phosphorylation of ADP Enzyme: Pyruvate Kinase ○ Most common enzyme deficiency (enzymopathy) of anaerobic Figure 14. Fate of Pyruvate glycolysis. III. GLUCONEOGENESIS Pyruvate is NOT a carbohydrate. Synthesis of glucose from non-carbohydrate precursors Does not contain an -OH Gluconeogenic substrates include: Pyruvate is a product of carbohydrate metabolism ○ Amino acid (Gluconeogenic amino acids e.g. Alanine) ○ Pyruvate ○ Lactate B. ANAEROBIC GLYCOLYSIS ○ Glycerol STEP 11: REDUCTION ○ TCA OR Krebs Cycle intermediates Most of gluconeogenic enzymes are similar to glycolytic enzymes EXCEPT these essentially irreversible enzymes: ○ Hexokinase [Step 1], ○ Phosphofructokinase (PFK) [Step 3], and ○ Pyruvate kinase [Step 10] Therefore, gluconeogenesis can be considered as a reversal of glycolysis EXCEPT for the 3 irreversible glycolytic steps. Figure 12. Reduction in Anaerobic Conditions Substrate: Pyruvate ○ Fermentation step in the absence of oxygen ○ Energy-requiring step ○ Pyruvate is reduced to lactate (gaining electrons) Electrons came from NADH and H+ (which is electron-rich) ○ Requirements: NADH and H If the NADH, H+ produced in Step 6 does not go to electron transport chain, it is used here Thus, no net production of NADH and H+ End product: Lactate and NAD+ Enzyme: Lactate Dehydrogenase Reversible ○ But forward reaction is favored LEORA ○ Lose Electron ○ Oxidation ○ Reducing Agent GEROA ○ Gain Electron ○ Reduction ○ Oxidizing Agent Products of aerobic glycolysis ○ Pyruvate ○ NADH and H+ ○ ATP ○ H2O Figure 15. Overview of gluconeogenesis. Enlarged figure in the appendix. OS 201 Metabolism of Carbohydrates 3 of 12  Information from Batch 2028 Trans Recall OAA needs to form a bond with Acetyl-CoA for the first step of the Krebs cycle to happen OAA is reduced to malate in the mitochondrial matrix ○ Catalyzed by malate dehydrogenase ○ This enzyme converts malate to OAA in the Krebs cycle but OAA to malate for the gluconeogenic pathway Malate leaves the mitochondria to be oxidized back to OAA in the cytosol Pyruvate carboxylase is found in the mitochondria while phosphoenolpyruvate carboxykinase occurs in the cytosol STEP 2: FRUCTOSE 1,6-BISPHOSPATE TO FRUCTOSE 6-PHOSPHATE Figure 16. Gluconeogenesis and glycolysis Figure 19. Conversion of fructose 1,6-bisphosphate to fructose 6-phosphate Hydration and phosphate removal of Fructose 1,6-bisphosphate Information from Batch 2028 Trans to Fructose 6-Phosphate catalyzed by Fructose Gluco = Glucose; Neo = New; Genesis = Beginning 1,6-Bisphosphatase (F-1,6-BP) ○ “Beginning of new glucose” Substrate: F-1,6BP ○ 90% of gluconeogenesis occurs in the liver producing glucose End product: Fructose-6-phosphate from non-carbohydrate sources Enzyme: Fructose-1,6-bisphosphatase Stimulus: low glucose level or fasting state ○ Presence of this enzyme determines whether tissue is capable of The body wants to increase the glucose level back to 4-6 performing gluconeogenesis mmol/L The reaction occurs in the cytoplasm ○ Usually happens during fasting (post-absorptive) and starvation states STEP 3: GLUCOSE 6-PHOSPHATE TO GLUCOSE ○ α-cells in pancreas release glucagon, cortisol, noradrenaline ○ Results in increase in glucose Why do we need to produce glucose specifically? ○ Main source of energy of the brain Cannot utilize energy from fatty acids Ketones are only backup sources ○ Maintenance of intermediates of the citric acid cycle STEP 1: PYRUVATE TO PHOSPHOENOLPYRUVATE Figure 20. Glucose 6-phosphate to glucose Hydration and phosphate removal of Glucose 6-Phosphate to Glucose catalyzed by Glucose 6-phosphatase Substrate: G6P End product: Glucose Enzyme: Glucose-6 Phosphatase Glucose 6-phosphatase is found in the liver and kidney but not in muscles The reaction occurs in the smooth endoplasmic reticulum Glucose is no longer trapped and can finally leave the cell → Figure 17. Conversion of pyruvate to phosphoenolpyruvate increases blood glucose The reverse for the 10th step of glycolysis SUMMARY STEP 1A: Carboxylation Table 2. SUMMARY OF GLUCONEOGENESIS Pyruvate is carboxylated (i.e. a CO2 from mitochondrion was added) Step Substrate Enzyme Product to oxaloacetate (OAA) catalyzed by pyruvate carboxylase Substrate: Pyruvate 1A Pyruvate Pyruvate carboxylase Oxaloacetate (OAA) End products: Oxaloacetate (OAA) (Mitochondria) ○ Recall OAA needs to form a bond with Acetyl-CoA for the first PEP carboxykinase step of the Krebs cycle to happen 1B OAA PEP (Cytoplasm) Enzyme: Pyruvate carboxylase Fructose-1,6-bisphosphatase 2 F-1,6BP Fructose-6-phosphate STEP 1B: Decarboxylation and Phosphorylation (Cytoplasm) Glucose-6 Phosphatase OAA is then decarboxylated (i.e. a CO2 was removed and released to 3 G6P (Smooth ER) Glucose cytoplasm) and phosphorylated to phosphoenolpyruvate (PEP) by phosphoenolpyruvate carboxykinase Substrate: OAA Which of the following are considered to be substrates for End products: PEP gluconeogenesis? Enzyme: PEP carboxykinase Substrate Answer/Explanation Occurs in the cytosol GTP is the phosphate donor Yes: It can be converted back to pyruvate and then Lactate glucose through gluconeogenesis (Cori cycle). Yes: It can be converted to pyruvate, serving as a Alanine key gluconeogenic precursor. Yes: It can be converted to dihydroxyacetone Glycerol phosphate (DHAP), an intermediate in gluconeogenesis. No: It is purely ketogenic and cannot be converted Leucine into glucose. No: It is primarily used for fatty acid synthesis and Acetyl-CoA ketone body formation, not for glucose production. No: Cholesterol is involved in membrane structure Figure 18. Carboxylation of pyruvate Cholesterol and steroid synthesis but does not participate in gluconeogenesis. OS 201 Metabolism of Carbohydrates 4 of 12 Glucogenic vs Ketogenic Amino Acids ○ Glucose is used to produce about 34-36 ATP molecules ○ Animals store glucose in the form of glycogen in fed (postprandial)/absorptive states (0-4 hours after eating) ○ Plants store glucose in the forms of starch and cellulose Starch is broken down by the human body in the postabsorptive state via amylase Not as branched as glycogen Body does not have enzymes to break cellulose down ○ Branched polymer of ⍺-D-glucose connected via ⍺-1,4 glycosidic bonds with branches at ⍺-1,6 position ○ Occurs mainly in the liver and muscle, with modest amounts in the brain Muscle glycogen is a ready source of glucose-1-phosphate for the muscles Liver glycogen is a reserve source for maintenance of blood glucose in the fasting state Figure 21. Glucogenic and Ketogenic Amino Acids Glucogenic Amino Acids: Can be converted into glucose. Examples include alanine, arginine, and aspartate. Ketogenic Amino Acids: Converted into ketone bodies for energy. Examples include leucine and lysine. Information from Batch 2028 Trans DIABETIC KETOACIDOSIS Buildup of substances called ketones (i.e. ketone bodies) to dangerous levels in the body Ketone bodies (beta-hydroxybutyrate and acetoacetate) are synthesized from acetyl CoA in the excess of glycerol and fatty acids relative to carbohydrates and proteins Among type I diabetic patients who lack insulin, ketone bodies become the source of energy, especially of the brain ○ Ketone bodies do not produce glucose ○ Low insulin is a trigger for the continuous breakdown of fatty acids ○ Ketones are oxidized to produce ATP and produce ○ H+ as a by-product making cellular environment acidic E.g. One mole of acetoacetate is converted to 2 acetyl CoAs ○ Oxidized completely to CO2 (acidifying the cell) and H2O in the brain mitochondria via the TCA ○ After the ETC, it will yield 2 GTPs and 22 ATPs in total The buildup of acid leads to diabetic ketoacidosis Figure 23. Scheme of glycogen synthesis and degradation IV. GLYCOGEN METABOLISM (S1): Glucose 6-phosphate is formed from glucose by hexokinase in most cells, and glucokinase in the liver. A. OVERVIEW ○ It is a metabolic branch point for the pathways of glycolysis, the Glycogen is the main storage form of glucose in animals. pentose phosphate pathway, and glycogen synthesis. Glucose enters the cell, and it is phosphorylated. (S2): Uridine diphosphate glucose (UDP-G) is synthesized from ○ It is converted into Glucose-1-Phosphate, which eventually glucose 1-phosphate. becomes glycogen. ○ UDP-glucose is the branch point for glycogen synthesis and Glycogen, when you need it, will undergo degradation to produce other pathways that require the addition of carbohydrate units. Glucose-1-Phosphate and Glucose-6-Phosphate. (S3): Glycogen synthesis is catalyzed by glycogen synthase and the ○ Glucose-6-Phosphate only becomes glucose in the liver. branching enzyme. There is an enzyme called Glucose-6-Phosphatase, which is (D1): Glycogen degradation is catalyzed by glycogen phosphorylase only found in the liver. and a debrancher enzyme. Glucose-6-Phosphatase will not be found in the muscle cell (D2): Glucose 6-phosphatase in the liver (and, to a small extent, the because the glucose found in the muscle (stored as glycogen) kidney) generates free glucose from glucose 6-phosphate. is used by the muscle; it is not used by other tissues. ○ ATP, adenosine triphosphate; Pi, inorganic phosphate; PPi, It provides sugar for itself because it is highly pyrophosphate; UTP, uridine triphosphate. energy-requiring. The liver, on the other hand, will produce the sugar for other tissues. Information from Batch 2028 Trans Figure 24. Pathways of glycogenesis and glycogenolysis in the liver Figure 22. Overview of glycogen metabolism OS 201 Metabolism of Carbohydrates 5 of 12 Figure Explanation from Harper’s Illustrated Biochemistry STEP 4: INITIATION OF GLYCOGEN SYNTHESIS ○ Insulin decreases the level of cAMP only after it has been raised Substrate: UDPG by glucagon or epinephrine; that is, it antagonizes their action. ○ UDPG attaches to a specific tyrosine residue in the glycogen ○ Glucagon is active in heart muscle but not in skeletal muscle. primer or glycogenin ○ Glucan transferase and debranching enzymes appear to be two End product: Glycogen primer separate activities of the same enzyme. Enzyme: Glycogenin ○ 37-kDa protein B. GLYCOGENESIS ○ Acts as an enzyme that catalyzes the attachment of glucose molecules to itself (autocatalysis) Biosynthesis of glycogen ○ Glucose is attached to the side chain hydroxyl group of Glycogen Synthase – Forms ɑ 1-4 linkages tyrosine-194 in glycogenin Branching Enzyme – Forms ɑ 1-6 linkages ○ The glycogenin and the attached glucose assembly becomes the Glycogenin – enzyme that acts as a primer for initial glucose substrate for glycogen synthase in the next step attachment; contains a tyrosine residue where UDP-G attaches ○ Glycogen cannot be produced on its own; there has to be a backbone, scaffold, or foundation. STEP 5: CHAIN EXTENSION Enzyme: Glycogen synthase Information from Batch 2028 Trans Glycosidic bonds are formed between C1 of the glucose of ○ Also known as glycogen synthesis UDP-glucose and C4 of the terminal glucose of glycogen ○ Occurs when there is excess glucose molecules ○ Liberates UDP ○ Occurs mainly in the muscles and liver cells ○ Addition of glucose to the chain is always at the non-reducing end ○ Chain of glucose continues to elongate as successive 1→4 STEP 1: PHOSPHORYLATION linkages are formed Also known as glycogen synthesis Glucose enters the cell via GLUT2 (hepatocytes) and GLUT4 STEP 6: BRANCHING (myocytes and adipocytes) Enzyme: Glycogen branching enzyme Substrate: Glucose Occurs to synthesize a more compact glycogen molecule End product: Glucose-6-Phosphate (G6P) Chains of at least 11 glucose residues long begin to branch Enzyme: Hexokinase or glucokinase Branching enzyme transfers at least 6 glucose residues from the ○ Hexokinase in the muscle non-reducing end of the ⍺(1→4) chain to neighboring chain ○ Glucokinase in the liver ○ Forms an α-1,6 glycosidic bond This is done to “trap” the glucose so it cannot return to the Branches may elongate further by addition of glucose residues and bloodstream (since the receptor for a phosphorylated glucose will further branching be different from the normal glucose molecule) ○ Increases number of non-reducing ends for further elongation and degradation STEP 2: ISOMERIZATION ○ Increases solubility of glycogen Substrate: G6P End product: Glucose 1-Phosphate (G1P) ○ Substrate used in glycogen synthesis Enzyme: Phosphoglucomutase ○ Phosphoglucomutase is phosphorylated in the process STEP 3: FORMATION OF UDP-GLUCOSE AND PYROPHOSPHATE Substrate: G1P, Uridine triphosphate (UTP) End product: Uridine diphosphate glucose (UDPG) and pyrophosphate (PPi) Enzyme: UDPG pyrophosphorylase ○ Has high affinity for G1P ○ Present in relatively large amounts Forward reaction (production of UDPG) is favored due to hydrolysis of PPi by pyrophosphatase ○ This removes PPi as a reactant for a possible reverse reaction Figure 27. Creating Branches with Glycogen Figure 25. Uridine Diphosphate Glucose (UDPG) Figure 26. UDPG Synthesis Figure 28. Transfer of Branching OS 201 Metabolism of Carbohydrates 6 of 12  ○ Due to glucose-6-phosphatase ○ Glucose exits the cell via GLUT2 (hepatocytes) and GLUT4 (myocytes and adipocytes) Blood glucose levels increase PHOSPHORYLATION REACTION Basic glycogenolysis reaction is phosphorylation reaction ○ Performed by glycogen phosphorylase ○ Requires presence of inorganic phosphates ○ Activity is only present in linear molecules of glycogen Figure 29. Glucose Residues Added on the Non-Reducing Ends of Glycogen Can only cleave α-1,4 linkages in the glycogen backbone When there are branch points (i.e., α-1,6 linkages at the branches), glycogen phosphorylase cannot perform the removal of glucose It is important to know this [pathway] because there are metabolic moiety disorders where there is branching enzyme deficiency. ○ Due to steric constraints ○ It is rare, but it can happen, especially in the pediatric age ○ Enzyme can only act on residues at least four (4) glucose units group. away from branch points ○ What happens when there is branching enzyme deficiency? In Fig. 31, the activity of the debranching enzyme will remove the No branches → Glycogen will become linear → Will take up final glucose more space than necessary → Liver and muscle cells will Glucosidase part of debranching enzyme will remove the final become enlarged (not in a good way) glucose ○ Branch point is eliminated, glycogen phosphorylase can resume formation of G1P Figure 30. Glycogen Storage Diseases Figure 31. Phosphorylation of glycogen Table 3. SUMMARY OF GLYCOGENESIS Reactant Product Enzyme Activation of glucose for glycogenesis Glucose G6P Hexokinase (muscle) or glucokinase (liver G6P G1P Phosphoglucomutase G1P + UTP UDPGIc + PPi UDPGIc pyrophosphorylase Glycogen synthesis: initiation, elongation, branching Glycogenin + Glucose Glycogen primer Glycogenin (from UDPGIc) Glycogen primer + Longer chain of Glycogen synthase pre-existing glycogen glycogen molecule Straight chain of Branched chain Amylo-(1,4→1,6) glycogen molecule of glycogen -transglucosidase C. GLYCOGENOLYSIS Catabolism of glycogen to produce glucose-1-phosphate ○ Process by which glycogen is broken down Takes place in hepatocytes and myocytes where glycogen is present Activated during the fight-or-flight response (sympathetic nervous system) Figure 32. Process of glycogen debranching Utilizes debranching process Information from Batch 2028 Trans Also known as glycogen degradation Substrate: Glycogen End products ○ Glucose-1-phosphate (G1P) ○ Small amounts of dephosphorylated glucose (due to the debranching enzyme) Main enzymes: Glycogen phosphorylase and glucose 6-phosphatase BASIC STEPS Glycogen is phosphorylated into G1P ○ Action of glycogen phosphorylase and debrancher enzyme ○ Small amounts of unphosphorylated glucose are liberated due to the debrancher enzyme Majority of end products of glycogen degradation via glycogen phosphorylase is still G1P G1P is further converted into G6P ○ In myocytes, G6P produced is shunted to glycolysis to provide energy due to the high energy requirement Figure 33. Process of glycogen debranching G6P is dephosphorylated to liberate glucose OS 201 Metabolism of Carbohydrates 7 of 12  Step 1: ○ Substrate: Glycogen ○ Enzyme: Glycogen Phosphorylase ○ Product: Glucose-1-phosphate (G1P) and a shorter glycogen chain) ○ Majority of end products of glycogen degradation via glycogen phosphorylase is still G1P Step 2: ○ Substrate: Glycogen (at the branch point) ○ Enzyme: Transferase ○ Product: A linear glycogen chain and a small oligosaccharide ○ In myocytes, G6P produced is shunted to glycolysis to provide energy due to the high energy requirement Step 3: ○ Substrate: Oligosaccharide (from the previous step) ○ Enzyme: α-1,6- Glucosidase and water ○ Product: Free glucose (from the hydrolysis of the α-1,6 bond) and a further shortened glycogen chain without any phosphate group ○ Due to glucose-6-phosphatase ○ Glucose exits the cell via GLUT2 (hepatocytes) and GLUT4 (myocytes and adipocytes) ○ Blood glucose levels increase Main end point of debranching is to remove the branches Figure 35. Reactions occurring during the oxidative phase of the hexose monophosphate pathway Information from Batch 2028 Trans STEP 1: Dehydrogenation Substrate: G6P, NADP+ ○ G6P is dehydrogenated at the hydroxyl group of C-1 to form 6-phosphogluconolactone End product: 6-phosphogluconolactone, NADPH, H+ Enzyme: Glucose 6-phosphate dehydrogenase ○ Deficiency in this enzyme can lead to the hemolysis of red blood cells of susceptible individuals STEP 2: Lactone hydrolysis Substrate: 6-phosphogluconolactone ring ○ 6-phospho-gluconolactone ring is hydrolyzed to open the ring End product: 6-phosphogluconate (6PG) Enzyme: Lactonase STEP 3: Oxidation of 6-phosphogluconate Substrate: 6PG End product: D-ribulose 5-phosphate, NADPH, H+ CO2 is released Enzyme: 6-phosphogluconate dehydrogenase Figure 33. Summary of Glycogen Metabolism STEP 4: Isomerization V. HEXOSE MONOPHOSPHATE PATHWAY/SHUNT Substrate: D-ribulose 5-phosphate One of the pathways that G6P from glucose may undergo End product: D-ribose 5-phosphate Alternative names: pentose phosphate pathway (PPP) and ○ End product of oxidative phase phosphogluconate pathway ○ Can enter the non-oxidative phase or used to synthesize Occurs in cytosol nucleotides While it produces reducing equivalents used in reductive Enzyme: Phosphopentose isomerase synthesis, no net energy is produced or consumed Two phases: Oxidative and Non-oxidative B. NON-OXIDATIVE PHASE Series of interconversion reactions producing a variety of monosaccharides with 3-7 carbons Catalyzed by various enzymes: transketolases, transaldolases, epimerases, isomerases Once D-ribose 5-phosphate is produced, and if it is NOT utilized in nucleotide synthesis, it enters the non-oxidative phase Interconvert sugars into other sugars having different number of carbons Glyceraldehyde-3-phosphate can be used in glycolysis Figure 34. Hexose Monophosphate Pathway A. OXIDATIVE PHASE G6P is converted into ribose 5-phosphate (R5P) + CO2 + 2 NADPH + H+ ○ R5P may proceed to the non-oxidative phase or nucleotide synthesis ○ NADPH and H+ are important in reductive synthesis, especially in lipogenesis (e.g. synthesis of cholesterol, fatty acids) ○ NADPH also acts like NADH as reducing equivalent Electron-rich, capable of donating them Involves a decarboxylation reaction Irreversible redox steps/reactions NADPH is different from NADH ○ NADPH is not used in the electron transport chain, unlike NADH ○ NADPH is used in reductive synthesis Figure 36. Reactions occurring during the non-oxidative phase of the hexose E.g. cholesterol synthesis, fatty acid synthesis, etc. monophosphate pathway OS 201 Metabolism of Carbohydrates 8 of 12  Information from Batch 2028 Trans Table 4. SUMMARY OF ENZYMES IN THE NON-OXIDATIVE PHASE OF HEXOSE MONOPHOSPHATE PATHWAY ENZYME ACTION Transketolase Transfer of two-carbon fragment from ketose donor to aldose acceptor Transaldolase Transfer of three-carbon fragment from ketose donor to aldose acceptor Epimerase Catalyze interconversion of ribose 5-phosphate (R5P) and xylulose-5-phosphate Isomerase Catalyze formation of glucose 6-phosphate (G6P) from fructose 6-phosphate (F6P) C. SIGNIFICANCE Figure 38. Fructose metabolism Production of NADPH and H+ (reducing equivalents) used for FRUCTOSE METABOLISM IN THE LIVER reductive synthesis of lipids (lipogenesis) Production of important intermediates for nucleotide synthesis Fructose metabolism occurs at a faster rate relative to glucose (e.g., ribose-5-phosphate) metabolism. Production of phosphorylated sugars with 3-7 carbons that may ○ Why? participate in carbohydrate metabolism pathways Triose production from fructose-1-phosphate bypasses phosphofructokinase-1, the major rate-limiting step in glycolysis. [LIPPINCOTT, HARPER’S] D. CLINICAL CORRELATION STEP 1: Phosphorylation of Fructose by Fructokinase Information from Batch 2028 Trans Fructose must be phosphorylated to enter the pathways of G6PD Deficiency intermediary metabolism. [LIPPINCOTT] X-linked recessive disorder that manifests itself as anemia Table 5. Phosphorylation of Fructose by Fructokinase ○ Uncommon but should be one of the differentials in patients with anemia COMPONENT DESCRIPTION Patients are usually asymptomatic SUBSTRATE Fructose, ATP ○ May manifest only when triggers challenge the body ○ Stressors include infections, stress, medication (e.g. Fructokinase ENZYME sulfonamides), food (fava beans), etc. - Phosphorylates fructose at C1 Cell is unable to maintain sufficient levels of reduced glutathione PRODUCT Fructose-1-phosphate, ADP, H+ (GSH) Result in a decrease in NADPH supply resulting in diminished reduced GSH ○ GSH is important in maintaining the integrity of RBCs ○ PPP is the only way for RBCs to maintain reduced GSH Hexose monophosphate pathway is important in having sufficient GSH levels, protecting fragile red blood cells from oxidative stress Figure 39. Phosphorylation of Fructose into F1P by fructokinase. [Andrey K; 2028 Trans] Phosphorylation of fructose into fructose-1-phosphate (F1P) may be accomplished by either hexokinase or fructokinase. [LIPPINCOTT] ○ HOWEVER, hexokinase has a low affinity (High Km) for fructose. [LIPPINCOTT] ○ MOREOVER, fructokinase has a low Km and high Vmax. [LIPPINCOTT] Fructokinase rapidly phosphorylates fructose as it enters the cell. [MARKS’] This enzyme is also found in the liver, the organ that processes most of the dietary fructose. [LIPPINCOTT] Figure 37. Effect of G6PD on erythrocyte response to oxidative stress FROM LIPPINCOTT Unless the intracellular concentration of fructose becomes unusually high, hexokinase is saturated with and phosphorylates VI. FRUCTOSE AND GALACTOSE METABOLISM glucose rather than fructose. Fructose (from plants) and galactose (from milk) do not have specific catabolic pathways unlike glucose (via glycolysis). [2028 TRANS] STEP 2: Aldol Cleavage of Fructose-1-phosphate A. FRUCTOSE METABOLISM Fructose is a sugar that you find in a lot of food products Fructose-1-phosphate is not phosphorylated into ○ E.g. fruits, honey, etc. fructose-1,6-bisphosphate as is fructose-6-phosphate. [LIPPINCOTT] Fructose is almost completely metabolized in the liver. ○ Instead, it is cleaved by aldolase B into DHAP and ○ Other tissues capable of fructose metabolism include the glyceraldehyde. intestine, testis, kidney, skeletal muscle, adipose tissue, and the brain. Table 6. Aldol Cleavage of Fructose-1-phosphate Majority of fructose is converted into glucose (29-54%) COMPONENT DESCRIPTION ○ 25% converted to lactate ○ 15-18% converted to glycogen SUBSTRATE fructose-1-phosphate ○ Essential fructosuria (EF) ○ GALE (UDP Galactose 4’-epimerase) ○ Rare, benign genetic disorder[MARKS’] converts UDP-Gal to UDP-Glu; deficiency may lead to Aldolase B deficiency -> Hereditary fructose intolerance (HFI) galactosemia ○ Can be fatal[MARKS’] QUESTION: What accounts for the difference in the effects of EF (benign) and HFI (possibly fatal)? ○ EF -> cannot make F1P -> accumulates fructose (which can be transformed into other molecules, not just F1P) No toxic metabolites of fructose accumulate in the liver; patient remains nearly asymptomatic[MARKS’] ○ HFI -> no aldolase B -> F1P not cleaved -> F1P accumulates F1P accumulation leads to: ↓ inorganic phosphate -> ↓ ATP production[LIPPINCOTT] Figure 42. Overview of Galactose Metabolism ○ ↓ gluconeogenesis -> hypoglycemia, vomiting[LIPPINCOTT] ○ ↓ protein synthesis -> decrease in blood-clotting factors and other essential protein[LIPPINCOTT] ○ Release of inhibition of AMP deaminase -> converts AMP to IMP (inosine monophosphate)[MARKS’] IMP -> degraded into uric acid -> uric acid accumulates -> formation of uric acid crystals[MARKS’] Inhibition of glycogen phosphorylase -> inhibition of glycogenolysis -> hypoglycemia[MARKS’] STEP 3: Phosphorylation of Glyceraldehyde Table 7. Phosphorylation of glyceraldehyde COMPONENT DESCRIPTION SUBSTRATE Glyceraldehyde, ATP ENZYME Triose kinase Glyceraldehyde-3-phosphate (G3P) Figure 43. Enzymes in Galactose Metabolism - Proceeds to the glycolytic pathway PRODUCTS ADP H+ STEP 1: Phosphorylation of Galactose Galactose must be phosphorylated before it can be further metabolized[LIPPINCOTT] Table 8. Phosphorylation of Galactose by Galactokinase COMPONENT DESCRIPTION SUBSTRATE Galactose, ATP Figure 41. Phosphorylation of glyceraldehyde into G3P by triose kinase. [Andrey K; 2028 Trans] Galactokinase (GALK) DHAP, a product from step 2, can also be converted into G3P via the - Phosphotransferase that catalyzes the first action of triose P isomerase. committed step in galactose metabolism The generated G3P molecules can be metabolized by a number of ENZYME - Phosphorylates galactose at C1 pathways, including the glycolytic pathway. - Destabilizes galactose and traps it inside the ○ Hence, fructose eventually joins the glycolytic pathway through cell (similar to the case of fructose G3P. phosphorylation by fructokinase)[2028 TRANS] Galactose-1-phosphate Information from Batch 2028 Trans PRODUCT ADP H+ FRUCTOSE METABOLISM IN OTHER TISSUES (NON-LIVER) Much easier in non-liver cells since fructose is converted by hexokinase into fructose-6-phosphate (F6P), instead of F1P. This is notable because of its easy entry into the glycolytic pathway wherein F6P is the product of the second step of glycolysis (isomerization) Phosphorylation of Fructose into Fructose-6-phosphate Figure 44. Phosphorylation of Galactose into Galactose-1-phosphate by galactokinase. [Holden et al.; 2028 Trans] STEP 2: Formation of Uridine Diphosphate-Galactose Figure 42. Phosphorylation of fructose into F6P by hexokinase. [Andrey K; 2028 Trans] Galactose cannot enter the glycolytic pathway unless it is first SUBSTRATE: Fructose, ATP converted to uridine diphosphate (UDP)-galactose[LIPPINCOTT] Enzyme: Hexokinase ○ Transfers phosphoryl from ATP to C6 of fructose Product: Fructose-6-phosphate, ADP, H+ OS 201 Metabolism of Carbohydrates 10 of 12 Table 9. Phosphorylation of Galactose by Galactokinase COMPONENT DESCRIPTION SUBSTRATE Gal1P, UDP-glucose Galactose 1-phosphate uridyltransferase (GALT) - Converts Galactose-1-phosphate to ENZYME Glucose-1-phosphate Figure 47. Transformation of G1P into G6P. [Holden et al.; 2028 Trans] - Transfers a phosphoryl group from UDP-glucose into phosphate region of C. CLINICAL APPLICATIONS Gal1P[2028 TRANS] UDP-galactose - Undergoes epimerization (see step 3) Information from Batch 2028 Trans PRODUCT Glucose-1-phosphate (G1P) FRUCTOSE METABOLISM - Can be isomerized into G6P to enter glycolysis Fructose is common in sodas and candy. It is converted into (see step 4)[LIPPINCOTT] DHAP as its metabolic entry. ○ Fructose metabolism involves only the third irreversible step of glycolysis, which means that the metabolic pathway is easier. ○ Because the energy-investing parts of glycolysis are the first five steps, fructose metabolism invests much less energy because it mostly skips these stps. Therefore, compared to glycolysis: ○ Pyruvate is easier to produce ○ Acetyl-CoA is easier to produce Excess acetyl-CoA is converted into fatty acids and cholesterol, because acetyl-CoA can only be stored as such. It cannot partake in gluconeogenesis. Beta-oxidation of lipids also yields acetyl-CoA, which is then converted back into fatty acids. Fructose is much more conducive to weight gain. Figure 45. Uridine diphosphate-galactose formation. [Holden et al.; 2028 Trans] The reaction involves oxidation, and then reduction, at C4, with NAD+ as a coenzyme.[HARPER’S] GALACTOSE METABOLISM Essentially an Exchange reaction[LIPPINCOTT] Problems in the enzymes of Galactose Metabolism ○ What goes into the reaction: ○ GALK (Galactokinase) UDP-glucose Deficiency will cause galactose accumulation -> ↑ galactitol -> galactose-1-phosphate obscure lens opacity (cataracts) ○ What are produced: ○ GALT (Galactose 1-P uridyltransferase) UDP-galactose Absent in classic galactosemia (condition that can be fatal if Glucose-1-phosphate undiagnosed in neonates) ○ GALE (UDP-Galactose 4’-epimerase) Deficiency may lead to galactosemia STEP 3: Epimerization of UDP-Galactose For UDP-galactose to enter the mainstream of glucose metabolism, it must first be isomerized to its C-4 epimer, UDP-glucose[LIPPINCOTT] Table 10. Phosphorylation of Galactose by Galactokinase COMPONENT DESCRIPTION SUBSTRATE UDP-galactose UDP-hexose 4-epimerase (UDP-galactose ENZYME 4’-epimerase; GALE) - Transforms UDP-gal into UDP-glu UDP-glucose - Can participate in biosynthetic reactions (e.g., PRODUCT glycogenesis) as well as in the GALT reaction (step 2 of galactose metabolism)[LIPPINCOTT] Figure 48. Metabolism of Galactose. [LIPPINCOTT] Figure 46. Epimerization of UDP-galactose. [Holden et al.; 2028 Trans] This reaction is freely reversible ○ Glucose can be converted to galactose -> galactose is NOT a dietary essential[HARPER’S] STEP 4: Transformation of G1P into G6P Table 11. Phosphorylation of Galactose by Galactokinase COMPONENT DESCRIPTION Figure 49. Metabolism of galactose. [MARKS’] SUBSTRATE Glucose-1-phosphate (G1P) VII. REFERENCES Phosphoglucomutase Formation and Degradation of Glycogen. (2022). Retrieved from ENZYME - Transfers phosphoryl group of G1P from C1 to https://basicmedicalkey.com/formation-and-degradation-of-glycogen/ C6[2028 TRANS] Lippincott Illustrated Reviews: Biochemistry 8E. Wolters Kluwer. Peet, A. (2012). Marks' Basic Medical Biochemistry. Lippincott Williams & Glucose-6-phosphate Wilkins. - Enters the glycolytic pathway as the substrate Rodwell, V.W., Bender, D.A., Botham, K.M., Kennelly, P.J., &; Weil, P.A. (2018). PRODUCT to be catalyzed into F6P (glycolysis step 2: Harper’s Illustrated Biochemistry. McGraw-Hill Education. isomerization)[2028 TRANS] Tiotuyco, A. R. (2024). An Overview of Carbohydrate Metabolism. UPCM 2028 Trans (2023). Metabolism of Carbohydrates OS 201 Metabolism of Carbohydrates 11 of 12  VIII. APPENDIX Figure 15. Overview of gluconeogenesis. Figure 30. Glycogen Storage Diseases OS 201 Metabolism of Carbohydrates 12 of 12