Week 12: Glucogenesis and Glycogen Metabolism

Fates of Glucose

• Glycolysis: A 10-step metabolic pathway for oxidation yielding ATP, NADH, and pyruvate.

• Synthesis of structural polymers: Used to produce extracellular matrix and cell wall polysaccharides.

• Storage: Stored in the form of glycogen and starch.

• Pentose Phosphate Pathway: Glucose is alternatively oxidised by this pathway.

Pentose Phosphate Pathway (PPP) + Erythrocytes

• Glucose 6-Phosphate as Starting Point:

  • Oxidation of Glucose 6-phosphate (G6P, 6C) yields ribose 5-phosphate (R5P, 5C) and NADPH.

  • Operates mostly in the cytoplasm of liver and adipose (fat) cells, and red blood cells (RBC), but not in skeletal muscle.

  • Ribose 5-phosphate: Serves as a precursor for RNA, DNA, and coenzymes.

  • NADPH: Drives fatty acid synthesis, cholesterol, and steroid hormones in the liver, adrenal glands, and gonads.

 

• Phases of PPP:

  • Oxidative phase: Glucose 6-phosphate is converted to 6-Phosphogluconate, Ribulose 5-phosphate, and Ribose 5-phosphate, releasing \(\text{CO}_2\) and generating NADPH.

  • Non-oxidative phase: Uses transketolase and transaldolase to convert Ribulose 5-phosphate into Fructose 6-phosphate and Glyceraldehyde 3-phosphate.

 

• Importance in Erythrocytes:

  • Erythrocytes rely heavily on the PPP for NADPH production because they lack mitochondria and cannot generate NADPH through other pathways.

  • NADPH is essential for regenerating reduced glutathione (2 GSH) from oxidized glutathione (GSSG) via glutathione reductase, which protects erythrocytes from oxidative damage.

  • Adequate NADPH levels help maintain the structural integrity and function of erythrocytes.

Gluconeogenesis Overview

• Purpose: Produces glucose from pyruvate or non-carbohydrate precursors:

  • Certain amino acids (carbon skeletons only).

  • Triacylglycerols (glycerol only).

• Occurrence: Occurs in all animals, plants, fungi, and microorganisms.

• Tissue Location: In mammals, gluconeogenesis occurs in the liver and kidneys.

• Essential Fuel Source: Glucose is the only energy source used by the brain (with the exception of ketone bodies during times of fasting), testes, erythrocytes, and the kidney medulla.

Gluconeogenic Substrates

• Key Precursors:

  • C3 Precursors: Lactate, pyruvate, glycerol, and some amino acids.

  • TCA Cycle Intermediates: Can also be used to make glucose.

  • Any compound that can be converted to either pyruvate or oxaloacetate can serve as a starting material for gluconeogenesis.

• Glucogenic Amino Acids Grouped by Site of Entry:

  • Pyruvate: Alanine, Cysteine, Glycine, Serine, Threonine, Tryptophan*.

  • alpha-Ketoglutarate: Arginine, Glutamate, Glutamine, Histidine, Proline.

  • Succinyl-CoA: Isoleucine*, Methionine, Threonine, Valine.

  • Fumarate: Phenylalanine*, Tyrosine*.

  • Oxaloacetate: Asparagine, Aspartate.

  • Note: All these amino acids are precursors of blood glucose or liver glycogen because they can be converted to pyruvate or citric acid cycle intermediates. Of the 20 common amino acids, only leucine and lysine are unable to furnish carbon for net glucose synthesis (* denotes amino acids that are also ketogenic).

Glycolysis Vs Gluconeogenesis (Differences, irreversible steps)

1. Key differences

Q: Where do glycolysis and gluconeogenesis mainly occur?

• Glycolysis: Mainly in muscle and brain.

• Gluconeogenesis: Mainly in the liver.

Q: Are glycolysis and gluconeogenesis exact opposites?

No. 7 glycolysis steps are reversible and shared, while 3 irreversible steps must be bypassed.

Q: What happens to glucose-6-phosphate (G6P) in muscle vs liver?

• Muscle: Enters glycolysis to produce energy.

• Liver: Glucose-6-phosphatase converts G6P into free glucose in the ER, allowing glucose to be released into the blood.

2. Bypass reactions

Q: Which 3 irreversible glycolysis steps are bypassed in gluconeogenesis?

Glycolysis enzyme	Gluconeogenesis bypass
1. Hexokinase	Glucose-6-phosphatase
2. Phosphofructokinase-1 (PFK-1)	Fructose-1,6-bisphosphatase
3. Pyruvate kinase	Pyruvate carboxylase + PEP carboxykinase

Q: Why are bypass reactions needed?

The 3 irreversible glycolysis reactions cannot simply run backwards, so gluconeogenesis uses different enzymes.

Glycolysis/Gluconeogenesis Regulation

Q: How are glycolysis and gluconeogenesis regulated?

They are reciprocally regulated: when one pathway is active, the other is generally inhibited to prevent futile cycling.

Q: Where do the 3 bypasses occur?

• Bypass 1: Mitochondria and cytoplasm.

• Bypass 2: Cytoplasm.

• Bypass 3: Endoplasmic reticulum (ER).

3. Bypass 1 — Pyruvate to PEP

Q: What are the two enzymes that convert pyruvate to PEP?

1. Pyruvate carboxylase: Pyruvate (3C) → oxaloacetate (4C).

2. PEP carboxykinase (PEPCK): Oxaloacetate → PEP (3C), releasing CO₂.

Q: What does pyruvate carboxylase require, and what activates it?

• Requires ATP and biotin.

• Activated by acetyl-CoA, signalling that gluconeogenesis should be favoured over further oxidation of acetyl-CoA in the citric acid cycle.

Pyruvate Vs Lactate Derived Routes

4. Pyruvate-derived vs lactate-derived routes

Q: What is the pyruvate-derived route from mitochondria to cytoplasm?

1. Pyruvate → oxaloacetate in the mitochondria.

2. Oxaloacetate → malate so it can cross the mitochondrial membrane.

3. Malate exits into the cytoplasm and converts back to oxaloacetate.

4. Cytosolic PEPCK converts oxaloacetate → PEP, releasing CO₂.

Q: What is the lactate-derived route to PEP?

1. Lactate → pyruvate in the cytoplasm via lactate dehydrogenase, producing NADH.

2. Pyruvate enters the mitochondria.

3. Pyruvate → oxaloacetate via pyruvate carboxylase.

4. Mitochondrial PEPCK converts oxaloacetate → PEP, releasing CO₂.

5. PEP exits to the cytoplasm.

Q: What is the key difference between the two routes?

• Pyruvate-derived: Oxaloacetate leaves as malate; PEP is made in the cytoplasm.

• Lactate-derived: PEP is made inside the mitochondria and then transported out.

Memory aid: Pyruvate route = malate shuttle; lactate route = mitochondrial PEP.

Bypass 2 + 3

• Bypass 2 – Formation of Fructose 6-Phosphate:

  • Fructose-1,6-bisphosphatase (FBPase-1) converts Fructose 1,6-bisphosphate o Fructose 6-phosphate.

  • Requires Mg²⁺ as a cofactor.

  • Facilitates the formation of ATP.

  • Subject to regulation by Fructose 2,6-bisphosphate and dual regulation by AMP.

• Bypass 3 – Formation of Glucose:

  • Glucose-6-phosphatase converts glucose 6-phosphate to glucose.

  • Under substrate-level control.

  • Performs "un-trapping" of glucose, permitting its release into the bloodstream.

  • Enzyme is found in the liver and kidney, but not in skeletal muscle and brain.

  • Allows liver glycogen to be converted to glucose and released into the bloodstream.

Gluconeogenesis Net Equation

 2Pyruvate + 4ATP + 2GTP + 2NADH + 2H+ + 4H₂O ---> Glucose + 4ADP + 2GDP + 6P_i + 2NAD+

Gluconeogenesis Regulation

• Reciprocal Regulation: When glycolysis is turned on, gluconeogenesis is turned off.

• Cellular Energy Status:

  • Cell energy is HIGH: Glycolysis should be off; pyruvate channels towards the synthesis and storage of glucose (or fatty acids).

  • Cell energy is LOW: Glycolysis should be on to provide energy.

• Regulators:

  • The fate of pyruvate depends on Acetyl-CoA levels.

  • Mediated through the 3 bypass steps.

  • Fructose 2,6-bisphosphate (F26BP) activates PFK-1 (glycolysis) and inhibits FBPase-1 (gluconeogenesis).

Cori Cycle

• Mechanism:

  • Vigorous exercise can lead to a buildup of lactate and NAD⁺ in contracting muscle due to O₂ shortage and the need for elevated glycolysis.

  • Lactate is returned to the liver via the blood, where it is reoxidized to pyruvate by liver lactate dehydrogenase (LDH).

  • The liver provides glucose (via gluconeogenesis, costing 6 ATP) to muscle for exercise and reprocesses lactate into new glucose.

Defects in Gluconeogenesis

• General Effect: A deficiency of any of the four gluconeogenic enzymes results in severe hypoglycaemia and lactic acidosis:

  • Bypass 1: Pyruvate carboxylase (PC) & PEP carboxykinase (PEPCK).

  • Bypass 2: Fructose-1,6-bisphosphatase (FBPase-1).

  • Bypass 3: Glucose-6-phosphatase (G6Pase).

• Pyruvate Carboxylase (PC) Deficiency: Causes failure to thrive, developmental delay, recurrent seizures, and metabolic acidosis, leading to tissue and nervous system damage.

Glycogen as Energy Reservoir

• Glycogen Function: Serves as an important energy reservoir whose breakdown is carefully controlled.

• Chemical Structure: Branched homopolymer of (alpha 1,4)-linked glucose chains cross-linked approximately every 10 units by (alpha 1,6)-glycosidic linkages.

• Major Stores: Located in two major tissues in the body: liver and muscle tissue.

Glycogen and Osmotic Balance (Relationship between Glycogen and Glucose Metabolism)

• Polymerization Necessity: Storing glucose as glycogen polymers prevents osmotic stress in the cytosol, which would otherwise be detrimental to the cell.

• Cytosolic Storage Granules:

  • Beta-granules: Cytosolic granules that vary in size, structure, and subcellular location.

  • alpha-granules: Protein-rich granules composed of 20–40 clustered (beta)-granules.

  • Visible in well-fed animals, but absent after a 24-hour fast.

• Anaerobic Fuel: Unlike fatty acids, released glucose can supply energy in the absence of O₂ for anaerobic activity.

Glycogen in Metabolism

• Muscle Glycogen: Immediate source of energy for both aerobic and anaerobic activity.

• Liver Glycogen: Reservoir of glucose for other tissues when dietary glucose is unavailable between meals (Note: The brain cannot use fatty acids as an energy source).

• Two Main Processes:

  1. Glycogenesis: Glycogen synthesis.

  2. Glycogenolysis: Glycogen breakdown.

Direct Vs Alternative Paths of Glucose Uptake

• Direct Path:

  • Uses the same first step as glycolysis, where glucose is converted to Glucose 6-phosphate (G-6-P) by hexokinase.

  • Operates in all tissues, with muscle and liver being the prominent sites.

• Alternative Path:

  • Glucose is taken up by erythrocytes and converted to lactate.

  • Blood lactate is subsequently taken up by the liver and converted to glucose via gluconeogenesis.

  • Erythrocytes contain no mitochondria, and thus no Citric Acid Cycle.

Formation of Glucose-1-Phosphate

• Reaction: Phosphoglucomutase catalyses the reversible conversion of Glucose 6-phosphate to Glucose 1-phosphate.

• Mechanism: Involves rotation of the phosphate group resulting in relocation of the phosphoryl group from C-6 to C-1.

• Note on Erythrocytes: NADPH is essential for regenerating reduced glutathione, protecting erythrocytes from oxidative damage.

Formation of UDP-Glucose

• Reaction: UDP-glucose pyrophosphorylase adds UTP to Glucose 1-phosphate to form UDP-glucose.

• Thermodynamics: Irreversible and strongly exergonic

• Activation: The anomeric carbon of glucose is activated by attachment to a nucleotide via a phosphate ester linkage.

• Substrate: Sugar nucleotides (such as UDP-glucose) serve as the substrates for polymerization.

Glycogen Formation + Branching

• Glycogen Synthase: Transfers glucosyl units from UDP-glucose to the non-reducing end of a glycogen chain via an (alpha 1--> 4) linkage.

• Glycogenin Core & Priming:

  • Glycogenin protein forms the core of a glycogen particle.

  • Catalyses autocatalytic formation of a glycosidic bond between the glucose of UDP-glucose and Tyr194 of glycogenin.

  • Adds seven more glucose residues from UDP-glucose to build a primer that can be acted upon by glycogen synthase.

• Branching Enzyme: Amylo-(alpha 1-4  --> alpha 1- 6) transglycosylase forms a new branch point via an (alpha 1-6) linkage.

Branching Purpose + Energy Cost

• Advantages of Branching:

  • Increases solubility of the glycogen molecule.

  • Increases the number of non-reducing ends, allowing for more rapid mobilization of glucose when needed.

  • Increases the number of accessible sites for regulatory enzymes.

• Energy Cost: Incorporating 1 molecule of glucose into glycogen costs 2 ATP equivalents:

  • 1 ATP to convert Glucose to Glucose 6-phosphate.

  • 1 UTP to convert Glucose 1-phosphate to UDP-glucose.

Glycogenolysis - Breaking down Glycogen

• Three Major Enzymes in Glycogen Degradation:

  1. Glycogen phosphorylase: Cleaves glucose from the non-reducing ends of glycogen.

  2. Debranching enzyme: Cleaves "limit dextrins" in the branched structure of glycogen.

  3. alpha-Amylase: Cleaves glycogen into maltose, maltotriose, and other small oligosaccharides in a random manner.

Glycogen Phosphorylase Mechanism

• Function: Catalyses phosphorolytic cleavage at the non-reducing ends of glycogen chains.

• Cofactor: Requires pyridoxal phosphate (Vitamin B6).

• Mechanism: Inorganic phosphate attacks the \(\alpha 1\rightarrow 4\)-glycosidic bond at the non-reducing end, yielding Glucose 1-phosphate.

• Limit: Acts repetitively until it reaches a point four residues away from an (\(\alpha 1\rightarrow 6\)) branch point.

Importance of Cellular Compartmentalization

• Compartmental Separation: Pathways are separated into distinct cellular compartments (e.g., Glucose 6-phosphatase is located inside the ER lumen, whereas hexokinase is in the cytosol).

• Prevention of Futile Cycling: Compartmentalizing enzymes prevents simultaneous synthesis and breakdown of glucose, avoiding wasteful energy expenditure.

• Precise Regulation: Enables more precise control over metabolic pathways.

• Efficiency: Ensures glucose generated by gluconeogenesis or glycogenolysis is efficiently released into the bloodstream rather than immediately re-phosphorylated and consumed in the cell.

Hormonal Control of Glucose Metabolism

• Insulin: Stimulates glycogen synthesis and inhibits glycogen breakdown in liver and muscle tissue during the fed state.

• Glucagon: Stimulates glycogen breakdown specifically in liver tissue during the fasting state.

• Adrenaline (Epinephrine): Stimulates glycogen breakdown in both liver and muscle tissue during fight-or-flight responses.

• [Image/Diagram Note: Comprehensive hormonal signal transduction cascade diagram comparing High Blood Glucose (Insulin) vs Low Blood Glucose (Glucagon) pathways is located on Slide 17].

Insulin Action

• Trigger: Elevated blood glucose levels after a meal trigger insulin release from pancreatic \(\beta\) cells.

• Metabolic Effects in Liver:

  1. Stimulates glycogenesis.

  2. Stimulates glycolysis.

  3. Inhibits glycogenolysis.

• Muscle Effects: Produces similar metabolic effects in muscle tissue.

Glucagon Action

• Trigger: Reduced blood glucose levels between meals trigger glucagon release from pancreatic (alpha) cells.

• Signaling Mechanism: Acts through G-protein coupled receptors using cAMP as an intracellular second messenger.

• Metabolic Effects in Liver:

  1. Inhibits glycogenesis.

  2. Inhibits glycolysis.

  3. Stimulates glycogenolysis.

• Muscle Effects: Exerts little effect on muscle tissue.

Adrenaline Action

• Trigger: Fight-or-flight or stressful situations trigger adrenaline release from the adrenal glands.

• Signaling Mechanism: Binds G-protein coupled receptors using cAMP as an intracellular messenger.

• Metabolic Effects in Liver:

  1. Stimulates glycogenolysis.

  2. Inhibits glycolysis.

  3. Stimulates gluconeogenesis.

• Metabolic Effects in Muscle:

  1. Stimulates glycogenolysis.

  2. Stimulates glycolysis.

Glycogen Storage Diseases

Etiology: Genetic defects in either Glucose 6-phosphatase or the Glucose 6-phosphate transporter T1 cause Type Ia glycogen storage disease.

Type Ia (von Gierke): Affected Enzyme = Glucose 6-phosphatase | Primary Organ = Liver | Symptoms = Enlarged liver, kidney failure.

Type IIIa (Cori or Forbes): Affected Enzyme = Debranching enzyme | Primary Organ = Liver, skeletal, and cardiac muscle | Symptoms = Enlarged liver in infants; myopathy.

Type IIIb: Affected Enzyme = Liver debranching enzyme (muscle enzyme normal) | Primary Organ = Liver | Symptoms = Enlarged liver in infants.

Type XI (Fanconi-Bickel): Affected Enzyme = Glucose transporter (GLUT2) | Primary Organ = Liver | Symptoms = Failure to thrive, enlarged liver, rickets, kidney dysfunction.