Topic 8: Glucose and Glycogen Metabolism

What is the primary energy source for many organisms?

D-glucose

Products & energy released of complete oxidation of glucose

CO₂ and H₂O

~ −2840 kJ/mol (highly energy-rich molecule)

What does glucose serve as for many biomolecules and multiple biosynthetic pathways

a key precursor

Why does glucose occupie a central position in metabolism?

Because of its central role in energy production and biosynthesis

Major Pathways of Glucose Utilization

glycolysis, Pentose Phosphate Pathway (PPP), Storage Pathways

Glycolysis

- Oxidation of glucose to pyruvate producing ATP and NADH

What can pyruvate do after glycolysis?

1. Enter cellular respiration for further ATP production

Or

2. serve as a precursor for biosynthetic reactions

Pentose Phosphate Pathway (PPP)

- Oxidizes glucose-6-phosphate to produce ribose-5-phosphate & NADPH

Ribose-5-phosphate (PPP)

required for nucleotide and nucleic acid synthesis

NADPH (PPP)

provides reducing power for biosynthetic reactions and antioxidant defense

Storage Pathways

- conversion of glucose to glycogen (animals) or starch (plants)

- Acts as a long-term energy reserve

- Can be mobilized when cellular energy demand increases

What was the first metabolic pathway to be fully elucidated (completed around 1940 after decades of research)?

Glycolysis

How long has humans used glycolysis indirectly?

for centuries through fermentation, long before understanding the biochemical mechanism

Glycolysis example

Yeast fermentation: converts glucose → ethanol + CO₂

- results from the anaerobic BD of glucose via glycolysis

What Happens in Glycolysis?

- Glucose (6C) partially oxidized through a series of enzyme-catalyzed rxns

- Produces 2 molecules of pyruvate (3C each)

- E released during oxidation used to synthesize ATP from ADP

- generates reducing equivalents (NADH)

Cellular Location of Glycolysis

In eukaryotic cells, glycolysis occurs in the cytosol

Glycolysis - Conservation Across Organisms

- highly conserved across all forms of life

- the chem rxns are identical in bacteria, plants, animals, and fungi

- Glycolytic enzymes strongly similar in sequence & structure across species

Glycolysis differences between organisms (2)

1. Regulation of the pathway

2. Fate of pyruvate: fermentation (microbes) vs. aerobic respiration (animals)

Number of reactions in glycolysis

10 enzyme-catalyzed reactions

2 phases of glycolysis

1. Preparatory (Investment) Phase - first 5 rxns

2. Payoff Phase - last 5 rxns

Which reactions are irreversible

1, 3, 10

- 7 is reversible, but highly favourable to one side)

Reversible reactions

can proceed in either direction depending on cellular conditions

- the same enzyme catalyzes both directions.

Glycolysis Reaction 1: Phosphorylation of Glucose

- Glucose is phosphorylated at the C-6 hydroxyl group by hexokinase

- ATP is the phosphate donor

Glycolysis Reaction 2: G6P to F6P

- Glucose-6-phosphate (an aldose) is converted to fructose-6-phosphate (a ketose) by phosphohexose isomerase

- an isomerization rxn (rearrangement of atoms w/i the molecule)

Glycolysis Reaction 3: Phosphorylation of Fructose-6-Phosphate

- F-6-P phosphorylated at C-1 position to form fructose-1,6-bisphosphate (ATP & phosphofructokinase-1)

- Called bisphosphate b/c the 2 phosphate groups are attached to different C atoms

diphosphate

molecule with 2 phosphate groups attached to the same atom

Glycolysis Reaction 4: Cleavage of Fructose-1,6-Bisphosphate

- 6-C fructose-1,6-bisphosphate split into two 3-Cmolecules by aldolase

- Products: Dihydroxyacetone phosphate (DHAP) & Glyceraldehyde-3-phosphate (G3P)

- reverse rxn of an aldol condensation → called retro-aldol cleavage.

Glycolysis Reaction 5: Isomerization of Triose Phosphates

- Only glyceraldehyde-3-phosphate (G3P) can proceed through

- Dihydroxyacetone phosphate (DHAP) produced in Reaction 4 is converted to G3P by triose phosphate isomerase

- ensures both 3-C molecules produced from glucose continue in glycolysis

Preparatory (Investment) Phase Summary

- rxns 1-5 which prepare glucose for E extraction

- 2 ATP molecules consumed (Rxns 1 & 3)

- Fructose-1,6-bisphosphate split into 2 triose phosphates (Rxn 4)

- DHAP → G3P so both molecules enter next phase (Rxn 5).

Resulting in 2 molecules of G3P produced from 1 glucose

- each step of the payoff phase has to occur twice per glucose molecule because two G3Ps present

Glycolysis Reaction 6: Oxidation of Glyceraldehyde-3-Phosphate

- Glyceraldehyde-3-phosphate (G3P) oxidized to 1,3-bisphosphoglycerate (1,3-BPG) by glyceraldehyde-3-phosphate dehydrogenase

- Inorganic phosphate (Pi) added to the molecule

Glycolysis Reaction 7: ATP Formation (Substrate-Level Phosphorylation)

- Phosphate group at C-1 of 1,3-bisphosphoglycerate transferred to ADP, producing ATP by phosphoglycerate kinase

- Occurs through transfer of a high-energy acyl phosphate group

- reversible rxn but strongly favored in the forward dirn

Coupling of Reactions 6 & 7

- Oxidation of glyceraldehyde-3-phosphate (rxn 6) generates high-E intermediate (1,3-BPG)

- the stored E is used to synthesize ATP in Rxn 7 (Substrate-Level Phosphorylation)

- Because 2 G3P molecules are formed from 1 glucose, this rxn produces 2 ATP per glucose molecule in glycolysis

Types of ATP Production in Glycolysis (3)

1. Substrate-Level Phosphorylation

2. Oxidative phosphorylation

3. Photophosphorylation

Substrate-Level Phosphorylation

ATP formed by direct transfer of a phosphate group from a phosphorylated substrate to ADP

Oxidative phosphorylation

ATP synthesis coupled to electron transfer to oxygen

Photophosphorylation

ATP synthesis driven by light energy during photosynthesis

Glycolysis Reaction 8: Conversion of 3-Phosphoglycerate to 2- Phosphoglycerate

- rearrangement rxn where phosphate group shifts from C-3 to C-2

- Mutases catalyze the transfer of functional groups within the same molecule (phosphoglycerate mutase)

Glycolysis Reaction 9: Formation of Phosphoenolpyruvate (PEP)

- rxn occurs through dehydration (removal of a water molecule)

- 1 H₂O released by enolase

Standard free energy of hydrolysis of 2-phosphoglycerate

−17.6 kJ/mol

Standard free energy of hydrolysis of phosphoenolpyruvate (PEP)

−61.9 kJ/mol

What does the removal of water cause within the molecule?

-does not greatly change the total E of the molecule

- causes E redistribution within the molecule resulting in formation of a very high-E phosphate bond in PEP

Important Concept of Reaction 9

- phosphoenolpyruvate (PEP) is the second high-E intermediate in glycolysis

- its high-energy phosphate bond will be used in the next step to generate ATP through substrate-level phosphorylation

Glycolysis Reaction 10: Formation of Pyruvate and ATP

- Phosphate group of PEP transferred to ADP, producing ATP (ex. of substrate-level phosphorylation) by pyruvate kinase

- initial product is enol-pyruvate, which quickly tautomerizes to the more stable keto form of pyruvate

- Remaining E released, making the rxn strongly favorable.

ATP Yield in Glycolysis

- ATP Investment Phase: 2 ATP consumed (rxn 1 & 3)

- ATP Generation Phase: 4 ATP produced (rxn 7 & 10)

- Net ATP gain of 2 ATP per glucose molecule

Net NADH Production in Glycolysis

- 2 NADH molecules

- produced during oxidation of glyceraldehyde-3-phosphate (Rxn 6).

Overall & Net Reaction Glycolysis

Overall Reaction:

Glucose + 2 ATP + 2 NAD⁺ + 4 ADP + 4 Pi → 2 Pyruvate + 2 ADP + 2 NADH + 2 H⁺ + 4 ATP + 2 H₂O

Net Reaction: (just cancel extras out, dont memorize)

Glucose + 2 ADP + 2 Pi + 2 NAD⁺ → 2 Pyruvate + 2 ATP + 2 NADH + 2 H⁺ + 2 H₂O

Glycolysis Under Aerobic Conditions (with O2)

- NADH transfers electrons to O2 through the ETC which regenerates NAD⁺, allowing glycolysis to continue

- Additional ATP is produced via oxidative phosphorylation

General Principles of Metabolic Regulation

- Cells regulate metabolic pathways to use E & metabolites efficiently

- Regulation usually occurs at key enzymatic steps in a pathway

Most important control points of Metabolic/glycolysis Regulation (3)

- First committed steps

- Exergonic and irreversible rxns

- Enzyme-limited steps

Major Factors Controlling Glycolysis (5)

1. ATP consumption (cellular energy demand)

2. Regeneration of NAD⁺

3. Allosteric regulation of key enzymes

4. Hormonal regulation (long-term control)

5. Expression levels of glycolytic enzymes

Key Regulatory Enzymes of Glycolysis (3)

1. Hexokinase

2. Phosphofructokinase-1 (PFK-1) - major control point

3. Pyruvate kinase

Regulation of Hexokinase

- Inhibited by glucose-6-phosphate

- Isoenzymes (Hexokinase I, II, III, IV)

Muscle Hexokinase (Hexokinase I)

- High affinity for glucose

- Half-saturation at ~0.1 mM glucose

- Maximally active at normal blood glucose (4-5 mM)

Glucokinase (Hexokinase IV) - Liver

- Low affinity for glucose

- Half-saturation at ~10 mM glucose

- Active only when glucose is high (after meals)

What specialized isoenzyme do liver cells contain?

glucokinase (Hexokinase IV or Hexokinase D)

Glucokinase (Hexokinase IV) - Characteristics

- Highly specific for D-glucose

- Lower affinity for glucose compared with hexokinase

Glucokinase (Hexokinase IV) - Physiological Role

- Becomes active when blood glucose levels are high (after a meal)

- Helps liver cells remove excess glucose from blood by converting it to glucose-6-phosphate

Glucokinase (Hexokinase IV) - Functional Significance

- Hexokinase: active at normal glucose levels

- Glucokinase: active at high glucose concentrations

- allows liver to regulate blood glucose levels after meals by inc glucose utilization

Why is Phosphofructokinase-1 (PFK-1) the main regulatory step/control point?

- Exergonic, Irreversible & 1st committed step of glycolysis

Committed step

produces a product that has to keep proceeding through the rest of glycolysis

- Glucose → Glucose-6-phosphate is not committed b/c G6P can enter the pentose phosphate pathway or glycogen synthesis

- Fructose-1,6-bisphosphate must proceed through glycolysis

Regulatory Importance

PFK-1's activity reflects the energy status of the cell

Activators (low energy signal) (3)

- AMP

- ADP

- Fructose-2,6-bisphosphate (in some tissues/organisms)

Inhibitor (high-energy signal) (1)

ATP

What happens to glycolysis when ATP levels are high?

glycolysis slows

What happens to glycolysis when AMP/ADP levels increase?

glycolysis is stimulated to produce more ATP

Key Hormones in Regulation of Glycolysis (Liver) (3)

1. Insulin

2. Glucagon

3. Epinephrine

Insulin (High Blood Glucose - Fed State)

- Stimulates glycolysis

- Inhibits gluconeogenesis

- Promotes glucose utilization

Glucagon (Low Blood Glucose - Fasting)

- Inhibits glycolysis

- Stimulates gluconeogenesis

Glycolysis in Cancer Cells (Warburg Effect)

- Cancer cells rely heavily on glycolysis for E

- Often experience hypoxia (low O2 ) due to poor blood supply

- Increased glucose uptake

Adaptations in Glycolysis in Cancer Cells

1. High expression of GLUT1 & GLUT3 glucose transporters

2. Hexokinase II

Hexokinase II Properties (Cancer Cell Adaptation)

- Binds mitochondrial outer MB

- Interacts w voltage dependent anion channel (VDAC)

- Can use mitochondrial ATP directly

- Unlike hexokinase I, not inhibited by glucose-6-phosphate

NAD⁺ Regeneration (Warburg Effect/Cancer Cells)

- Cell has limited amount of NAD+

- Pyruvate is converted to lactate

- NADH → NAD⁺ (regenerate) to allows glycolysis to continue

Regulation using Pyruvate Kinase

- last enzyme of glycolysis

- several isozymes encoded by deferent genes

- allosterically regulated

Regulation using Pyruvate Kinase in Muscle & Liver

In muscles: activated by E signals (AMP & ATP)

- In liver: 2 forms (Active/phosphorylated & inactive/dephosphorylated); when high sugar --> insulin activates; when low sugar --> glucagon inactivates

How is energy produced in red blood cells

- no mitochondria, so rely on glycolysis

pyruvate kinase deficiency/hemolytic anemia

- use ATP to pump sodium-potassium pumps in RBCs which maintain biconcave shape

- with PK deficiency, there is no ATP power pump --> cell shape becomes spiculated

Fate of Pyruvate

1. low O2: Fermentation to ethanol in yeast or Fermentation to lactate in muscle cells

2. Aerobic: Pyruvate converted to 2-Acetyl-CoA with loss of 2CO2, then through citric acid cycle become 4 CO2 + 4H2O

Location of Lactic Acid Fermentation

the cytoplasm of cells in mainly in animal muscle cells

Lactic Acid Fermentation Physiological Effects

Lactate accumulation can cause muscle fatigue and cramps during intense exercise

Cori Cycle (Lactic Acid Fermentation)

- Lactate produced in muscle is transported to the liver

- Liver converts lactate back to glucose

Industrial Importance of Lactic Acid Fermentation

- Lactobacillus species convert lactose sugar to lactic acid (yogurt, cheese, curd)

- Lactic acid lowers pH, causing denaturation and precipitation of milk proteins

Can other sugars enter glycolysis besides glucose?

Yes

Mannose Entry in Glycolysis

- mannose (derived from polysaccharides & glycoproteins)

- mannose to mannose-6-phosphate by hexokinase, then to fructose 6-phosphate by phosphomannose isomerase

Fructose Entry in Glycolysis (Most Tissues & Liver)

- In most tissues: fructose from fruits or produced by hydrolysis of sucrose. Converted to fructose-6-phosphate by hexokinase

- In liver tissues: fructose to fructose-1-phosphate by fructokinase, then into dihydroxyacetone phosphate + glyceraldehyde by fructose-1-phosphate aldolase

Galactose Entry in Glycolysis

- originates from lactose (milk sugar)

Galactose → Galactose-1-phosphate (Galactokinase) → UDP-galactose + Glucose-1-phosphate (UDP-glucose:galactose-1-phosphate uridylyltransferase (GALT)) → UDP-glucose- recycled (UDP-galactose 4-epimerase). Glucose-1-phosphate → Glucose-6-phosphate

Galactosemia

- caused by deficiency of enzymes in galactose metabolism (galactokinase, UDP-glucose galactose-1-phosphate urdiyltransferase, UDP-glucose-4-epiemrase)

Effects of Galactosemia

- accumulation of galactose & toxic metabolites in blood

- lead to cellular & tissue damage

Lactose Intolerance

- Caused by loss of lactase activity in intestinal cells during adulthood.

- Most humans produce lactase during childhood

- lactase production declines after childhood in many pops.

Consequences of Lactose Intolerance

- Undigested lactose reaches the intestine

- Gut microorganisms ferment lactose producing toxic metabolites and gas

Lactose Intolerance Symptoms

- Abdominal pain, bloating, diarrhea

Gluconeogenesis

synthesis of glucose from noncarbohydrate precursors

- reverse of glycolysis

Why is gluconeogenesis essential?

because some tissues rely heavily on glucose as their primary energy source

Major glucose-dependent tissues

- Brain and nervous system

- Erythrocytes (RBCs)

- Renal medulla

- Testes

- Embryonic tissues

Why is gluconeogenesis necessary?

- Glycogen stores are limited in the body

- brain alone requires more than half of total glycogen stores during fasting

- during fasting or intense exercise, glucose must be synthesized to maintain blood glucose levels

Occurrence of Gluconeogenesis

in animals, plants, and microorganisms

Similarity and difference of gluconeogenesis in different organisms

- core pathway is similar in all organisms

- regulation differs

Main site of gluconeogenesis in animals

- Liver (primary)

- Adrenal cortex (smaller extent)

What can animals synthesize glucose from? (Precursors for Gluconeogenesis)

- Pyruvate, Lactate, Glycerol, Glucogenic amino acids

- cannot synthesize glucose from acetyl-CoA produced from fatty acid oxidation

What can plants synthesize glucose from? (Precursors for Gluconeogenesis)

from acetyl-CoA via the glyoxylate cycle

What can microorganisms synthesize glucose from? (Precursors for Gluconeogenesis)

can use many different carbon sources to synthesize glucose