BIOCHEM 4.2 - GLYCOLYSIS

Glycolysis Introduction

• Glycolysis is essential for energy in red blood cells (RBCs) because they operate anaerobically.
• It is crucial for the brain's energy needs
• Glycolysis starts with glucose and results in the generation of pyruvate.

Tracking Carbons

• During glycolysis, a glucose molecule is broken down into two molecules of pyruvate.
• This involves a series of enzymatic reactions where carbon atoms are rearranged, leading to the production of ATP and NADH.

Irreversible Steps of Glycolysis

• Some steps in glycolysis are irreversible.
• Special enzymes are required to reverse these steps in reactions like gluconeogenesis.

Glyceraldehyde 3-Phosphate Dehydrogenase (GAPDH)

• In step 1, glyceraldehyde-3-P (RCHO) interacts with GAPDH's active site sulfhydryl group and forms thiohemiacetal adduct.
• In step 2, the thiohemiacetal is oxidized into a thioester by NAD+, which is then reduced to NADH.
• NAD+ is bound in the active site of the enzyme.
• In step 3, a phosphorylase reaction cleaves the carbon-sulfur bond after a phosphate enters the active site.
• The 3-phosphoglycerate group is displaced, leading to the formation of 1,3-bisphosphoglycerate.
• Lastly, the 1,3-bisphosphoglycerate regenerates the sulfhydryl group.
• Enzyme exchange consists of NADH for NAD+.

Phosphofructokinase-1 Regulation

• Phosphofructokinase-1 (PFK-1) acts as the main regulatory point in glycolysis by controlling the conversion of fructose-6-phosphate (Fru-6-P) to fructose-1,6-bisphosphate (Fru-1,6-BP).
• PFK-1 indirectly affects the levels of glucose-6-phosphate (Glc-6-P) and inhibits hexokinase activity.
• Achieved through the reaction of phosphoglucose isomerase.
• ATP serves as both a substrate and an allosteric inhibitor of PFK-1.
• Providing essential control over its activity, especially in red blood cells (RBCs).
• PFK-1's activity varies based on the energy status of the cell.

Change in Free Energy

• Glycolysis, pyruvate oxidation, and the citric acid cycle are linked by changes in free energy.

Anaerobic Glycolysis

• Anaerobic glycolysis is the process of glucose utilization in red blood cells.
• The result of the process is the production of lactate.

Anaerobic Metabolism in Red Blood Cells

• In red blood cells (RBCs), glucose is broken down into 2 pyruvate molecules through a series of reactions.
• Pyruvate is converted to lactic acid through anaerobic glycolysis.
• This process generates ATP, which is used to maintain electrochemical and ion gradients across the RBC plasma membrane.
• An intermediate of glycolysis, 10-20% of 1,3-bisphosphoglycerate, is diverted to produce 2,3-bisphosphoglycerate (2,3-BPG).
• 2,3-BPG is a regulator of oxygen affinity for hemoglobin.
• 10% of glucose metabolism is shunted towards the pentose phosphate pathway to protect from oxidative stress and provide NADPH.

Pentose Phosphate Pathway

• The irreversible redox stage generates NADPH and pentose phosphates.
• In the reversible interconversion stage excess pentose phosphates are recycled into glycolytic intermediates.

Deficiencies in Glycolysis and Consequences for RBCs

• Errors in glycolysis lead to hemolytic anemias.
• Pyruvate kinase deficiency and G6PDH deficiency (from the pentose phosphate pathway) are common examples that cause hemolytic anemia.
• Diagnosis of these deficiencies relies on evaluating levels of enzymes/metabolites, measuring enzyme activity, and conducting genetic testing.
• Deficiencies lead to a diverse range of consequences, including varying degrees of cellular damage.
• Insufficient ATP leads to the inability to maintain ion gradients and maintain cell shape.
• G6PDH deficiency can cause significant lipid peroxidation and GSH loss because of a failure to recycle it.
• High levels of 2,3-bisphosphoglycerate (2,3-BPG) can decrease oxygen affinity, diminishing the capacity of tissues to receive oxygen.