Glycolysis and Gluconeogenesis
Alice Skoumalová
Metabolism of glucose - overview
1. Glycolysis
Glucose:
the universal fuel for human cells
Sources: diet (the major sugar in our diet) internal glycogen stores blood (glucose homeostasis)
Glucose oxidation: after a meal: almost all tissues during fasting: brain, erythrocytes
Glycolysis:
oxidation and cleavage of glucose
ATP generation (with and without oxygen)
all cells
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:
fatty acids amino acids ribosis-5-P
Anaerobic glycolysis
a limited supply of O2
no mitochondria increased demands for ATP
Lactic acidemia in hypoxia
Phosphorylation of glucose: irreversible
Glucose 6-P: cannot be transported back across the
plasma membrane a precursor for many pathways that
uses glucose
Hexokinases
Glucokinase (liver, β-cell of the pancreas)
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
• irreversible
• regulation
essential for the subsequent
cleavage
Substrate-level phosphorylation
Substrate-level phosphorylation
2. Oxidation and substrate-level phosphorylation
Summary of the glycolytic pathway:
Glucosis + 2 NAD+ + 2 Pi + 2 ADP
2 pyruvate + 2 NADH + 4 H+ + 2 ATP + 2 H2O
∆G0´ = - 22 kcal (it cannot be reversed without the expenditure of energy!)
Clinical correlations:
Hypoxemia (lack of oxygen in tissues)
Acute hemorrhage (hypotension, lost of erythrocytes)
- anaerobic glycolysis
- lactate formation, metabolic acidosis
Chronic obstructive pulmonary disease (an insuficient ventilation)
- anaerobic glycolysis, lactate formation, metabolic acidosis
- accumulation of CO2, respiratory acidosis
Myocardial infarction (lack of oxygen in myocardium)
- anaerobic glycolysis, lactate formation
- lack of ATP
Aerobic glycolysis:
involving shuttles that transfer reducing equivalents across the mitochondrial membrane
Glycerol 3-phosphate shuttle:
Malate-aspartate shuttle:
Anaerobic glycolysis:
Energy yield 2 mol of ATP
dissociation and formation of H+
Daily lactate production 115 (g/d)
Erythrocytes 29
Skin 20
Brain 17
Sceletal muscle 16
Renal medulla 15
Intestinal mucosa 8
Other tissues 10
Major tissues of lactate production:
(in a resting state)
Cori cycle:
Lactate can be further metabolized by: heart, sceletal muscle
Lactate dehydrogenase: a tetramer (subunits M and H)
Lactate dehydrogenase
Pyruvate + NADH + H+ lactate + NAD+LD
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
Fructose 2,6-bis-phosphate: is not an intermediate of glycolysis!
Phosphofructokinase-2: inhibited through phosphorylation - cAMP-dependent protein kinase (inhibition of glycolysis during fasting-glucagon)
• tissue-specific isoenzymes (low Km, a high afinity)
• glucokinase (high Km)
• the rate-limiting, allosteric enzyme• tissue-specific isoenzymes
the liver isoenzyme - inhibition by cAMP-dependent protein kinase (inhibition of glycolysis during fasting)
Lactic acidemia:
increased NADH/NAD+ ratio inhibition of pyruvate dehydrogenase
2. Gluconeogenesis
Gluconeogenesis:
synthesis of glucose from noncarbohydrate precursors → to maintain blood glucose levels during fasting
liver, kidney
fasting, prolonged exercise, a high-protein 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 Pyruvate carboxylase
2. Oxaloacetate → PEP Phosphoenolpyruvate-
carboxykinase
Conversion of phosphoenolpyruvate to glucose
3. Fructose-1,6-P → Fructose-6-P Fructose 1,6-bisphosphatase (cytosol)
4. Glucose-6-P → Glucose 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
3. Regulations
Type Mechanism Example
Substrate concentration Saturation kinetics (according Michaelis-Menten)
Glucokinase (activation after meal- high Km)
Allosteric Conformational changes induced by the binding of the allosteric efector
Enzymes of glycolysis and glukoneogenesis (allosteric effectors ATP, AMP, citrate)
Covalent modification Conformational changes induced by phosphorylation by proteinkinases
Phosphorylation og glycogensynthase and glycogenphosphorylase (glucagon)
Protein-protein interaction
Conformational changes as a result of different protein binding
Muscle glycogenphosphorylase (activated by Ca2+-calmodulin)
Proteolytic cleavage Activation by proteolytic cleavage of precursor molecule
Proteins of coagulation cascade (zymogens)
Enzyme synthesis Induction or repression of the enzyme synthesis
Enzymes of gluconeogenesis (induced during fasting)
Enzyme regulation:
Second messenger - cAMP:
GlucagonAdenylate-cyclase
Regulation of gluconeogenesis: concomitant inactivation of the glycolytic
enzymes and activation of the enzymes of gluconeogenesis
1. Pyruvate → PEPPhosphoenolpyruvate carboxykinase - induced by glucagon, epinephrine, and cortisol
2. Fructose 1,6-P → Fructose 6-PFructose 1,6-bisphosphatase - inhibited by fructose 2,6-P
3. Glucose 6-P → GlucoseGlucose 6-phosphatase - induced during fasting
Summary
Glycolysis• Generation of ATP (with or without oxygen)• The role of glycolysis in different tissues• Lactate production• Regulation (3 key enzymes)
Gluconeogenesis• Activation during fasting, prolonged exercise, after a high-
protein diet• Precursors: lactate, glycerol, amino acids• 3 key reactions: Pyruvate → PEP
Fructose-1,6-P→ Fructose-6-PGlucose-6-P → Glucose
• Regulation
Pictures used in the presentation:
Marks´ Basic Medical Biochemistry, A Clinical Approach, third edition, 2009 (M. Lieberman, A.D. Marks)