Phosphorylation of Glucose • There are TWO enzymes GlucokinaseGlucose + ATP Glucose–6–phosphate + ADP HexokinaseGlucose + ATP Glucose–6–phosphate + ADP
Trapping of Glucose by phosphorylation
Glucokinase enzyme GlucokinaseGlucose + ATP Glucose-6-phosphate + ADP
Comparison between Hexokinase & GlucokinasNo Factor Hexokinase Glucokinase1 Substrate All Hexoses Glucose only2 Distribution All tissues Liver only Product Inhibited by Not inhibited by3 inhibition Glucose-6-phosphate Glucose-6-phosphate Km for Low Km High Km4 glucose (High affinity) (Low affinity) Effect of Not affected Activated5 Insulin Effect of Not affected Activated6 Carbohydrate Effect of Not affected Inhibited7 Starvation
Comparison between Glucokinase & Hexokinase Tissue-specific distribution of the two enzymes ensures that: At low blood glucose concentrations, liver is prevented from utilizing glucose until the nutrient requirements of other tissues are satisfied
(Ethanol + CO2)(Lactate Fermentation)
Stages of Glycolysis < TARGET="display">
Stages of Glycolysis < TARGET="display">
Stages of Glycolysis
Substrate-LevelPhosphorylation Rx 7 and 10
PFK is the regulatory (key) enzyme in glycolysis!• The second irreversible reaction of glycolysis• Large negative ∆G, means PFK is highly regulated• PFK is regulated by: – Citrate is an allosteric inhibitor – ATP also inhibits PFK, while AMP activates PFK – Fructose-2,6-bisphosphate is allosteric activator – PFK increases activity when energy status is low – PFK decreases activity when energy status is high
1 2 3 5 4 6Metabolic Significance of Glycolysis9 8
• The three irreversible enzymes (Glucokinase, Phosphofructokinase, Pyruvate kinase) are under the control of Insulin• Insulin induces the synthesis of:1 < TARGET="display">23
Regulation of Glycolysis1- After carbohydrate meal:Blood glucose level Stimulates insulinsecretion Increases synthesis ofglucokinase, phosphofructokinase & pyruvate kinase Enhances glycolysis2- During fasting:Blood glucose level Inhibits insulinsecretion & stimulates glucocorticoid secretionIncreases the synthesis of the four enzymes thatreverse glycolysis (Stimulate gluconeogenesis)
3- Pasteur effect: Increased oxygen inhibits glycolysis, since increased citrate and ATP or increased ATP/ADP ratio which inhibit phosphofructokinase (the rate limiting enzyme of glycolysis) Decreased ATP/ADP ratio or increased ADP, AMP & Pi activates phosphofructokinase4. Glycolysis inhibited by iodoacetate, fluroacetate & arsenite, since they inhibit Kreb’s cycle
Fate of Pyruvate 2 Pyruvate Pyruvate Decarboxylase Pyruvate DH Lactate DH & Alcohol DHIn Mitochondria In Cytoplasm
< TD>For regeneration of NAD Lactate Dehydrogenase
Different forms of Lactate Dehydrogenase • Lactate DH in Heart & Muscles: Heart (H4) Muscle (M4) LDH1 LDH5 H H M M H H M M • Lactate DH in different tissues: H3M H2M2 HM3 LDH2 LDH3 LDH4 H H H H H M H M M M M M
1 Rapoport-Luebering Cycle in RBCs (R-L Cycle) Mutase 1,3- Diphosphoglycerate 2,3- Diphosphoglycerate (2,3-DPG) 3-Phoshoglycerate ADP kinase Phoshatase 7 Pi ATP 3- PhosphoglycerateComplete glycolysis • To meet this deficiency in ATP synthesis, glycolysis rate in RBCs increases.
1 Rapoport-Luebering Cycle in RBCs (R-L Cycle)
3 Pyruvate Kinase Deficiency in RBCs1. The genetic deficiency of pyruvate kinase in RBCs leads to hemolytic anemia.2. This is due to inhibited (reduced rate of) glycolysis and lowered level of ATP synthesis.3. So the rate of synthesis of ATP is inadequate to meet the energy needs of the cell to maintain the structural integrity of erythrocytes.
Two Shuttle Pathways forOxidation of Cytoplasmic NADH1. Malate shuttle (Dicarboxylic Acid Shuttle).2. Glycerol phosphate shuttle.
• Pyruvate Dehydrogenase: • A multienzyme complex has 3 Functions: 1. Decarboxylation: Removal of CO2 (Decarboxylase, TPP as coenzyme). 2. Oxidation of the remaining two-carbon compound and reduction of NAD+ (Dehydrogenase, CoASH, FAD & NAD+). 3. Trans-acetylating function: Attachment of CoA with a high energy thio-ester bond to form Acetyl CoA (Transacetylase, Lipoic acid).Pyruvate + NAD+ + CoASH Acetyl CoA + NADH + CO2
Pyruvate DHPyruvate Acetyl CoA 1 FAD 2 CoA-SH NADH + CO2 3 NAD+ 4 TPP 5 Lipoic acid
Regulation of Pyruvate DH - NADH & ATP Pyruvate DHPyruvate Acetyl CoA - - 1- Product inhibition2- Covalent modification(Protein kinase) + (directly) 3- Insulin
Carboxylation of Pyruvate - Pyruvate DHPyruvate Acetyl CoA + Insulin + Allosterically - Pyruvate CarboxylasePyruvate Oxaloacetate Biotin ATP + CO2 ADP + Pi
-Alanine Pantoic acid( )( ) CH3 OH
Ketone 3Bodies 4 1 5 Cholesterol 2 Sources & Fate of Acetyl CoA
Tricarboxylic Acid Cycle (TCA)Citric Acid Cycle, Kreb’s Cycle Third Stage of Metabolism
Comments & Biological Significance of TCA Cycle• It is the final pathway for oxidation (3rd stage) of all foodstuffs to CO2 + H2O + Energy.• It is important for the interconversion of carbohydrates, fats & proteins.• All reactions are reversible except: Citrate synthase, Isocitrate DH & -Ketoglutarate DH.• The rate limiting enzyme is Citrate synthase.
Comments & Biological Significance of TCA Cycle• Mitochondrial isocitrate DH is NAD+ linked, while cytoplasmic isocitrate DH is NADP+ linked.• TCA is the major source of succinyl Co A which used for: – Heme synthesis. – Ketolysis. – Detoxication reactions.
Regulation of Kreb’s Cycle1. Insulin activates pyruvate DH & inhibits pyruvate carboxylase, thus directing pyruvate towards complete oxidation through kreb’s cycle.2. Acetyl Co A inhibits pyruvate DH & activates pyruvate carboxylase, thus directing pyruvate & glucose towards formation of oxaloacetate to combine with excess Acetyl Co A for the optimal activity of kreb’s cycle.• During starvation (glucose supply is low & fat oxidation provides excess Acetyl Co A), so oxaloacetate is required.
Regulation of Kreb’s Cycle3. So, Kreb’s cycle is inhibited by: a) Starvation (No carbohydrates). b) Diabetes mellitus (No insulin). c) Anaerobic conditions (No oxygen).4. It is inhibited in vitro by fluroacetate & iodoacetate which form flurocitrate & iodocitrate that inhibits aconitase.5. Malonic acid is a competitive inhibitor of succinate dehydrogenase.6. Arsenite inhibits Kreb’s cycle.
Effect of Fluroacetate on TCA• Flurocitrate inhibits aconitase
Metabolic Significance of Kreb’s Cycle 1 4 2 3
Sources of Oxaloacetate1. Pyruvate, in mitochondria (Pyruvate carboxylase).2. Malate, in mitochondria, by malate DH.3. Citric acid, in cytoplasm (ATP-Citrate lyase).4. Aspartic acid, by transamination in both cytoplasm & mitochondria.
COO COOCOO CH2 COO CH2CH2 CH2 CH2 CH2HC NH3+ + C O C O + HC NH3+COO COO COO COOaspartate -ketoglutarate oxaloacetate glutamate Aminotransferase (Transaminase)
Fate of Oxaloacetate1. Aspartic acid, by transamination.2. Citric acid, by citrate synthase.3. Malate, in mitochondria, by malate DH.4. Phosphoenolpyruvate by reversal glycolysis (Gluconeogenesis), in cytoplasm, by PEP Carboxykinase.
Fate of CO21. Excretion through lungs (main fate).2. Combined with ammonia to form urea.3. Combined with ammonia to form pyrimidine.4. Enters in the formation of C6 of purines.5. Fixation into organic acids (Carboxylation): 1. Pyruvic acid + CO2 Oxaloacetic acid. 2. Acetyl Co A + CO2 Malonyl Co A. 3. Propionyl Co A + CO2 Methylmalonyl Co A.