Glycolysis Overview and Stages

Glycolysis: Common Pathway

• Occurs in the cytoplasm of eukaryotic cells
• Glucose is converted to 2 pyruvate and 2 ATP
• 4 ATP are made, but 2 are consumed
• Enzymes are organized to allow for substrate channeling, ensuring the movement of substrates and products through enzymes and preventing the release of toxic intermediates

Two Stages of Glycolysis

Stage 1: Trapping and Preparation Phase

• No ATP is generated, but some is invested
• Conversion of glucose to fructose 1,6-bisphosphate
• Three steps:
  • Phosphorylation (uses ATP) produces glucose 6-phosphate, trapping glucose inside the cell
  • Isomerization
  • Second phosphorylation (uses ATP) produces a compound that can be cleaved into two phosphorylated 3-carbon molecules
• Completed with the cleavage of fructose 1,6-bisphosphate into:
  • Glyceraldehyde 3-phosphate
  • Dihydroxyacetone phosphate (converted to glyceraldehyde 3-phosphate)

Key Enzymes in Glycolysis

Hexokinase

• Transfers phosphoryl group from ATP to glucose, forming glucose 6-phosphate
• Added to the hydroxyl group on carbon 6
• Allows for two things:
  • Prevents glucose from leaving the cell through the transporter it came in through (no longer the same substrate)
  • Enables the cell to utilize glucose for energy production

Regulation of Glycolysis

In Muscle Cells

• Energy charge of the cell is the primary control of glycolysis
• Ratio of ATP to AMP
• Phosphofructokinase is the most important control site
• Allosteric inhibition by ATP
• AMP reverses the inhibitory action
• Inhibited by a decrease in pH (caused by the buildup of lactic acid)

In Liver Cells

• Corresponds to the biochemical versatility of the liver
• Phosphofructokinase is regulated by ATP, but drops in ATP are not typically experienced in the liver
• Citrate inhibits phosphofructokinase
• Fructose 2,6-bisphosphate is an activator of phosphofructokinase
• Form of feedforward stimulation

Gluconeogenesis

• Synthesis of glucose from non-carbohydrate precursors
• Critical to maintain blood glucose levels
• Glucose is the primary fuel for the brain
• Glucose is the only fuel for red blood cells
• Glycogen reserves can supply glucose for about a day
• Primarily occurs in the liver and, to a lesser degree, in the kidney
• Glucose produced by gluconeogenesis helps supply the brain, skeletal muscle, or heart muscle

Regulation of Gluconeogenesis

• Not a complete reversal of glycolysis
• Must bypass the three irreversible reactions of glycolysis
• The conversion of pyruvate into phosphoenolpyruvate begins with the formation of oxaloacetate
• Oxaloacetate is shuttled into the cytoplasm and converted into phosphoenolpyruvate
• The conversion of fructose 1,6-bisphosphate into fructose 6-phosphate and orthophosphate is an irreversible step
• The generation of free glucose is an important control point
• Fructose 6-phosphate is readily converted into glucose 6-phosphate
• Glucose 6-phosphate is transported into the lumen of the endoplasmic reticulum
• Glucose 6-phosphate is hydrolyzed into glucose by glucose 6-phosphatase
• Glucose and Pi are shuttled back to the cytoplasm by a pair of transporters### Galactose and Glucose Metabolism
• Galactose 1-phosphate acquires a uridyl group from UDP-glucose, producing UDP-galactose and glucose 1-phosphate, catalyzed by galactose 1-phosphate uridyl transferase.
• UDP-galactose is converted to UDP-glucose by UDP-galactose 4-epimerase, which inverts the hydroxyl group at carbon 4.

Regulation of Glycolysis

• Glycolysis has two main roles: breaking down glucose to form ATP and providing building blocks for biosynthetic reactions.
• The pathway is regulated by allosteric effectors or covalent modification, with enzymes that catalyze irreversible reactions being sites of control.
• Key regulatory enzymes in glycolysis include:
  • Hexokinase: inhibited by its own product, glucose 6-phosphate.
  • Phosphofructokinase: most important control site in mammals, inhibited by ATP and activated by AMP.
  • Pyruvate kinase: regulated by ATP, fructose 1,6-bisphosphate, and alanine.

Glycolysis in Muscle and Liver

• In muscle, glycolysis is regulated by feedback inhibition to meet the need for ATP, with phosphofructokinase and pyruvate kinase being key regulatory enzymes.
• In liver, glycolysis is regulated by the biochemical versatility of the liver, with phosphofructokinase, hexokinase, and pyruvate kinase being regulated differently than in muscle.

Glucose Transport

• Glucose transporters allow for diffusion of glucose across the plasma membranes of animal cells.
• Different types of glucose transporters are found in different tissues, including:
  • GLUT1 and GLUT3: found in nearly all mammalian cells.
  • GLUT2: found in liver and pancreatic beta cells.
  • GLUT4: found in muscle and fat cells.
  • GLUT5: found in the small intestine.

Gluconeogenesis

• Gluconeogenesis is the synthesis of glucose from non-carbohydrate precursors, such as pyruvate, lactate, and amino acids.
• The pathway is critical for maintaining blood glucose levels, particularly in the brain and red blood cells.
• Gluconeogenesis occurs primarily in the liver and kidney, and involves the reversal of glycolysis, with some key differences:
  • The conversion of pyruvate to phosphoenolpyruvate involves the formation of oxaloacetate and the use of biotin as a prosthetic group.
  • The conversion of fructose 1,6-bisphosphate to fructose 6-phosphate involves the hydrolysis of fructose 1,6-bisphosphate.
  • The generation of free glucose is an important control point, with glucose 6-phosphate being transported into the lumen of the endoplasmic reticulum and then hydrolyzed into glucose.

Energy Charge and Regulation

• The energy charge of the cell determines whether glycolysis or gluconeogenesis will be more active.
• The energy charge is reflected in the levels of ATP, ADP, and AMP, with high energy charge favoring gluconeogenesis and low energy charge favoring glycolysis.
• The regulation of glycolysis and gluconeogenesis is also influenced by the levels of fructose 2,6-bisphosphate, which is an allosteric regulator of phosphofructokinase.

Balance Between Glycolysis and Gluconeogenesis

• The balance between glycolysis and gluconeogenesis is sensitive to blood glucose concentration, with high blood glucose levels favoring glycolysis and low blood glucose levels favoring gluconeogenesis.
• The balance is regulated by the phosphorylation of a single serine residue within a protein, which determines the activity of phosphofructokinase 2 and fructose bisphosphatase 2.