Glycolysis and Gluconeogenesis Alice Skoumalov 1 Glycolysis Glucose

Glycolysis and Gluconeogenesis Alice Skoumalová

1. Glycolysis

Glucose: the universal fuel for human cells Sources: Ø diet (the major sugar in our diet) Ø internal glycogen stores Ø blood (glucose homeostasis) Glucose oxidation: Ø after a meal: almost all tissues Ø during fasting: brain, erythrocytes

Glycolysis: Ø oxidation and cleavage of glucose Ø ATP generation (with and without oxygen) Ø all cells Ø in the cytosol (the reducing equivalents are transferred to the electron-transport chain by the shuttle) ATP is generated: 1. via substrate-level phosphorylation 2. from NADH 3. from oxidation of pyruvate Regulation of glycolysis: 1. Hexokinase 2. Phosphofructokinase 3. Pyruvate Kinase Generation of precursors for biosynthesis: Ø fatty acids Ø amino acids Ø ribosis-5 -P

Anaerobic glycolysis Øa limited supply of O 2 Øno mitochondria Øincreased demands for ATP Lactic acidemia Øin hypoxia

Phosphorylation of glucose: Ø irreversible Glucose 6 -P: Ø cannot be transported back across the plasma membrane Ø a precursor for many pathways that uses glucose Hexokinases Glucokinase (liver, β-cell of the pancreas) Ø high Km

Michaelis-Menten kinetics

1. Conversion of glucose 6 -P to the triose phosphates 2. Oxidation and substrate-level phosphorylation

1. Conversion of glucose 6 -P to the triose phosphates essential for the subsequent cleavage • irreversible • regulation

2. Oxidation and substrate-level phosphorylation Substrate-level phophorylation

Summary of the glycolytic pathway: Glucosis + 2 NAD+ + 2 Pi + 2 ADP 2 pyruvate + 2 NADH + 4 H+ + 2 ATP + 2 H 2 O ∆G 0´ = - 22 kcal (it cannot be reversed without the expenditure of energy!)

Clinical correlations: Hypoxemia (lack of oxygen in tissues) ØAcute hemorrhage (hypotension, lost of erythrocytes) - anaerobic glycolysis - lactate formation, metabolic acidosis ØChronic obstructive pulmonary disease (an insuficient ventilation) - anaerobic glycolysis, lactate formation, metabolic acidosis - accumulation of CO 2, respiratory acidosis ØMyocardial infarction (lack of oxygen in myocardium) - anaerobic glycolysis, lactate formation - lack of ATP

Aerobic glycolysis: Ø involving shuttles that transfer reducing equivalents across the mitochondrial membrane

Glycerol 3 -phosphate shuttle:

Malate-aspartate shuttle:

Anaerobic glycolysis: dissociation and formation of H+ Energy yield 2 mol of ATP

Major tissues of lactate production: (in a resting state) Daily lactate production 115 (g/d) Erythrocytes 29 Skin 20 Brain 17 Sceletal muscle 16 Renal medulla 15 Intestinal mucosa 8 Other tissues 10

Cori cycle: Lactate can be further metabolized by: Ø heart, sceletal muscle Lactate dehydrogenase: a tetramer (subunits M and H)

Lactate dehydrogenase Pyruvate + NADH + H+ LD lactate + NAD+ 5 isoenzymes: Heart (lactate) Muscle (pyruvate)

Biosynthetic functions of glycolysis:

Clinical correlations: Long-intensity exercise (for example a sprint) - the need for ATP exceeds the capacity of the mitochondria for oxidative phosphorylation, anaerobic glycolysis → lactate formation, muscle fatigue and pain - a training → the amounts of mitochondria and myoglobin increase

Regulation

• tissue-specific isoenzymes (low Km, a high afinity) • glucokinase (high Km) • the rate-limiting, allosteric enzyme • tissue-specific isoenzymes Fructose 2, 6 -bis-phosphate: Ø is not an intermediate of glycolysis! Ø Phosphofructokinase-2: inhibited through phosphorylation - c. AMP-dependent protein kinase (inhibition of glycolysis during fasting-glucagon)

the liver isoenzyme - inhibition by c. AMP-dependent protein kinase (inhibition of glycolysis during fasting) Lactic acidemia: increased NADH/NAD+ ratio inhibition of pyruvate dehydrogenase

2. Gluconeogenesis

Gluconeogenesis: Ø synthesis of glucose from noncarbohydrate precursors → to maintain blood glucose levels during fasting Ø liver, kidney Ø fasting, prolonged exercise, a highprotein diet, stress Specific pathways: 1. Pyruvate → Phosphoenolpyruvate 2. Fructose-1, 6 -P → Fructose-6 -P 3. Glucose-6 -P → Glucose

Precursors for gluconeogenesis 1. lactate (anaerobic glycolysis) 2. amino acids (muscle proteins) 3. glycerol (adipose tissue)

Conversion of pyruvate to phosphoenolpyruvate 1. Pyruvate → Oxaloacetate Ø Pyruvate carboxylase 2. Oxaloacetate → PEP Ø Phosphoenolpyruvatecarboxykinase

Conversion of phosphoenolpyruvate to glucose 3. Fructose-1, 6 -P → Fructose-6 -P Ø Fructose 1, 6 -bisphosphatase (cytosol) 4. Glucose-6 -P → Glucose Ø Glucose 6 -phosphatase (ER)

Clinical correlations: Alcoholism - excessive ethanol consumption → increase NADH/NAD+ ratio that drive the lactate dehydrogenase reaction toward lactate - lack of precursors for gluconeogenesis → its inhibition - insuficient diet - reduced glucose in the blood, consumption of glycogen in the liver → hypoglycemia

Regulation of gluconeogenesis: Ø concomitant inactivation of the glycolytic enzymes and activation of the enzymes of gluconeogenesis 1. Pyruvate → PEP Phosphoenolpyruvate carboxykinase induced by glucagon, epinephrine, and cortisol 2. Fructose 1, 6 -P → Fructose 6 -P Fructose 1, 6 -bisphosphatase - inhibited by fructose 2, 6 -P 3. Glucose 6 -P → Glucose 6 -phosphatase - induced during fasting

Summary Glycolysis • Generation of ATP (with or without oxygen) • The role of glycolysis in different tissues • Lactate production • Regulation Gluconeogenesis • Activation during fasting, prolonged exercise, after a highprotein diet • Precursors: lactate, glycerol, amino acids • 3 key reactions: Pyruvate → PEP Fructose-1, 6 -P→ Fructose-6 -P Glucose-6 -P → Glucose • Regulation

Pictures used in the presentation: Marks´ Basic Medical Biochemistry, A Clinical Approach, third edition, 2009 (M. Lieberman, A. D. Marks)