Lecture 4: Glycolysis PDF

Lect 5: Regulation of Glycolysis Done By Biochemistry Department Regulatory enzymes of glycolysis Glycolysis is regulated at the three irreversible (Committed) steps that are catalyzed by the key regulatory enzymes: a) Hexokinase/ glucokinase b) Phosohophructokina...

Lect 5: Regulation of Glycolysis Done By Biochemistry Department Regulatory enzymes of glycolysis Glycolysis is regulated at the three irreversible (Committed) steps that are catalyzed by the key regulatory enzymes: a) Hexokinase/ glucokinase b) Phosohophructokinase-1 ( PFK-1) (the most important one) c) Pyruvate kinase( PK) 11/24/2024 4 Overview of Glycolysis 11/24/2024 5 Glucokinase & Hexokinase Glucokinase / Hexokinase IV ▪ Present in liver parenchymal cells and β cells of the pancreas ▪ In β cells, glucokinase functions as a glucose sensor, determining the threshold for insulin secretion. ▪ In the liver, the enzyme facilitates glucose phosphorylation during hyperglycemia. ▪ Have a high (Km), and a low affinity for glucose. Therefore, functions only after a carbohydrate-rich meal to effectively remove glucose flood and minimizing hyperglycemia. Mutations of glucokinase cause a rare form of diabetes, maturity 11/24/2024 onset diabetes of the young type 2 (MODY 2) 7 Glucokinase & Hexokinase Hexokinase ▪ It catalyze phosphorylation of glucose in most tissue --- broad substrate specificity ▪ Inhibited by the reaction product, glucose 6-phosphate ▪ Stimulated by increased glucose level ▪ Have a low (Km), and a high affinity for glucose. Therefore, perform efficient phosphorylation and subsequent metabolism of glucose even when tissue concentrations of glucose are low 11/24/2024 8 PFK-1 Regulation by energy levels : PFK-1 inhibition: High levels of ATP (allosteric inhibitor) Elevated levels of citrate (most potent activator) Low pH PFK-1 Activation: AMP, ADP and Fructose 2,6 biphosphate *Fructose 2,6-bisphosphate is the most potent activator of PFK-1 and is able to activate the enzyme even when ATP levels are high. 11/24/2024 10 Pyruvate Kinase (PK) ▪ PK is activated by fructose 1,6- bisphosphate, the product of the PFK-1 ATP reaction. ▪ Pyruvate Kinase is inhibited by excess ATP (decreases its ability to bind to PhosphoEnol pyruvate (PEP). Acetyl CoA 11/24/2024 11 HORMONAL REGULATION OF GLYCOLYSIS ▪ During fasting: There are elevated levels of glucagon and low levels of insulin. This inhibits all the regulatory enzymes of glycolysis hence inhibit glycolysis ▪ During the well-fed state: There are decreased levels of glucagon and elevated levels of insulin. This stimulates all the regulatory enzymes of glycolysis hence activates glycolysis. 11/24/2024 15 Inhibitors of Glycolysis Arsenate poisoning: ▪ Competes with Pi as a substrate for glyceraldehyde 3-phosphate dehydrogenase, ▪ Prevent net ATP and NADH production 11/24/2024 17 Inhibitors of Glycolysis Fluoride: ▪ Inhibits enzyme enolase. ▪ Water fluoridation reduces lactate production by mouth bacteria, decreasing dental caries. ▪ It is also used in blood collection tubes for glucose estimation. 11/24/2024 18 Lecture 6: Oxidative Carboxylation of Pyruvate & Kreb’s Cycle Dr. Uzma Asif Sherwani Medicine Program BMC Introduction: (Fate of Pyruvate) The fate of pyruvate depends on oxygen availability. When oxygen is present, pyruvate is oxidized to acetyl-CoA which enters the Krebs cycle Without oxygen, pyruvate is reduced to Lactate to oxidize NADH back to NAD+ 3 PDH is the link between glycolysis and TCA Cycle: ▪ The end product of glycolysis aerobically is pyruvic acid. ▪ Pyruvic acid can be oxidized in presence of oxygen to be converted into acetyl CoA that can be degraded through Citric acid cycle to produce more energy ▪ Pyruvate is transported into the mitochondria via special pyruvate transporters. ▪ In mitochondria, pyruvic acid is oxidatively decarboxylated into Acetyl CoA by pyruvate dehydrogenase enzyme complex (PDH) Coenzymes: 1. TPP (Thiamine PyroPhosphate) ; active form of vitamin B1 2. FAD ; active form of vitamin B2 3. NAD+ ; active form of vitamin B3 4. Coenzyme A (CoASH); active form of vitamin B5 5. Lipoic acid ; a member of vitamin B complex Pyruvate dehydrogenase complex (PDH) 1 molecule of pyruvate provide 1 NADH Products: ▪One NADH+ that provides 3 ATP by its oxidation through Electron Transport Chain (ETC) in mitochondria. ▪As aerobic glycolysis produces 2 pyruvates, so, extra 6 ATP are produced by PDH Regulation of PDH activity: 1. Feed back inhibition ▪ Inhibitor: Acetyl CoA and NADH+H+ (the products of PDH). ▪ Activator: Pyruvate and Ca++ ion. 2. Allosteric control ▪ Inhibitor: ATP ▪ Activator: AMP. In this way, the activity of the complex is reduced when the cell is rich in available energy. Wernicke-Korsakoff Syndrome: Thiamine or niacin deficiency can cause serious central nervous system problems. This is because brain cells are unable to produce sufficient ATP (via the TCA cycle) if the PDH complex is inactive. Wernicke-Korsakoff, an encephalopathy-psychosis syndrome due to thiamine deficiency, may be seen with alcohol abuse. May cause confusion, oculomotor dysfunction, and gait ataxia. Intravenous Thiamine (Vit B1) is initially administered as a treatment option.  AMBOSS QUESTION: A. Vitamin B12 B. Vitamin B1 C. Vitamin B6 D. Vitamin B9 Carbohydrate Metabolism: Kreb’s Cycle Citric acid cycle (CAC) - Tricarboxylic acid cycle (TCA)- Krebs cycle Definition: Series of reactions in requires CoA (vitamin B5 pantothenic acid) mitochondria, that catabolizes Acetyl- CoA, leading to production of energy. Site : only occurs in mitochondria. requires CoA (vitamin B5 pantothenic acid) Energetics & Energy production sites of TCA Cycle: The 2 acetyl CoA produced from oxidation of one glucose molecule ,when catabolized through CAC, they generate: 6 NADH, 2 FADH2 and 2 ATP and 4 CO2 are released Panthothenic Acid: Vitamin B5 (a component of Coenzyme A) Vitamin B5 deficiency may lead to distal paresthesias (tingling, prickling) and dysesthesias (burning sensation of the feet) requires CoA (vitamin B5 pantothenic acid) Citrate synthase Oxaloacetate --------------------→ Citrate; requires CoA (vitamin B5 pantothenic acid) Enzymes that require CoA as a cofactor (e.g, alpha-ketoglutarate dehydrogenase, pyruvate dehydrogenase) would be affected by pantothenic acid (Vit B5) deficiency because pantothenic acid is required for requires CoA (vitamin B5 synthesis of CoA. pantothenic acid) Significance of TCA Cycle: 1 -Energy production Significance of TCA Cycle: (amphibolic pathway) Catabolic Functions Anabolic Functions 1 Citrate → acetyl CoA →synthesis of fatty acids and It is the final common cholesterol. metabolic pathway for oxidation of carbohydrates, fats and proteins. 2 Succinyl CoA →heme synthesis. 3 Oxaloacetate →PEP which is converted to glucose (gluconeogenesis) 4 α- ketoglutarate → glutamic acid and oxaloacetate → Aspartic acid ( amino acids) 5 CO2 produced is used in buffer system. Regulation of TCA Cycle (CAC): ▪ TCA cycle is controlled by the regulation of several enzymes ▪ The most important of these regulated enzymes are ✓ Citrate synthase, ✓ isocitrate dehydrogenase and ✓ α-ketoglutarate dehydrogenase ▪ They are inhibited by high ATP and NADH+H concentration. Inhibitors of TCA Cycle: ▪ Fluoroacetate, a plant toxin that is used as a pesticide inhibits aconitase enzyme. ▪ Malonic acid inhibits succinate dehydrogenase enzyme and succinate is accumulated. Total ATP produced from complete oxidation of one molecule of glucose through Glycolysis, Oxidative Decarboxylation and CAC Lecture 4: Glycolysis Done By Biochemistry Department Glycolysis; Overview: 11/24/2024 7 Glycolysis: Definition: It is a biochemical pathway that aims at degradation of glucose to generate ATP. Glycolysis can proceed in presence of O2 (aerobically) and in its absence (anaerobically). ❖Aerobically, its end product is pyruvate. ❖Anaerobically, its end product is lactate. Site of glycolysis: Cytoplasm of all tissue cells 11/24/2024 8 Energy Investment Phase / Preparatory Phase: Where glucose (hexose) is activated and cleaved into two trioses. In this phase there is loss (investment) of 2 ATP. 11/24/2024 11 Energy Generation Phase / Payoff phase: Where the 2 trioses are oxidized and energy is gained. 11/24/2024 12 Net ATP Production in Glycolysis 11/24/2024 16  b. Importance of glycolysis in red blood cells: Effect of 2,3-bisphosphoglycerate on oxygen affinity 2,3-BPG binds to hemoglobin → conformational change → release of oxygen into tissue 11/24/2024 22 c. Pyruvate kinase deficiency cause ATP deficiency: ▪ Mature RBCs lack mitochondria and are, therefore, completely dependent on glycolysis for ATP production. ▪ ATP is required to meet the metabolic needs of RBCs and to fuel the ion pumps necessary for the maintenance of the flexible, biconcave shape that allows them to squeeze through narrow capillaries. ▪ The anemia observed in glycolytic enzyme deficiencies is a consequence of the reduced rate of glycolysis, leading to decreased ATP production. ▪ The resulting alterations in the RBC membrane lead to changes in cell shape and, ultimately, to phagocytosis by cells of the reticuloendothelial system, resulting in hemolytic anemia. 11/24/2024 23