Regulation of pyruvate kinase

Pyruvate kinase enzyme regulation

  • Pyruvate kinase (PK) catalyzes the final step in glycolysis, producing the second of two ATP molecules generated in the glycolytic pathway.
  • The enzyme converts phosphoenolpyruvate (PEP) and ADP to pyruvate and ATP
  • This reaction is a committed step leading to either anaerobic fermentation or oxidative phosphorylation of pyruvate.
  • In most cells the reaction is essentially irreversible and is one of the major control points of glycolysis.
  • The regulation of PK is important for controlling the levels of ATP, GTP and glycolytic intermediates in the cell.
  • Pyruvate kinase also serves as a switch between the glycolytic and gluconeogenic pathways in certain tissues.

Pyruvate kinase is a tetrameric protein. There are four isozymes of pyruvate kinase in vertebrates:

L (liver), R (erythrocytes), M1 (muscles, hearts and brain) and M2 (only form detectable in early fetal tissue and present in most adult tissues).

Pyruvate kinase activity is most broadly regulated by allosteric effectors, covalent modifiers and hormonal control.

Figure 1: Regulation of pyruvate kinase activity

Allosteric effectors

Fructose-1, 6-bisphosphate (FBP): Allosteric activator of PK (Feedforward activation)

FBP binds to the allosteric binding site on domain C of pyruvate kinase and changes the conformation of the enzyme, causing the activation of pyruvate kinase activity. As an intermediate present within the glycolytic pathway, FBP provides feedforward stimulation because the higher the concentration of FBP, the greater the allosteric activation and magnitude of pyruvate kinase activity.

ATP: Allosteric inhibitor of PK (Feedback inhibition)

Acetyl-CoA and long-chain fatty acids (signs of abundant energy supply):  allosterically inhibit all isozymes of pyruvate kinase

Accumulation of alanine (which can be synthesized from pyruvate by transamination) allosterically inhibits pyruvate kinase, slowing the production of pyruvate by glycolysis.

Covalent modification

  • The liver isozyme (L form), but not the muscle isozyme (M form), is subject to further regulation by phosphorylation.
  • Covalent modifiers serve as indirect regulators by controlling the phosphorylation and dephosphorylation of enzymes, resulting in the activation and inhibition of enzymatic activity.
  • In the liver glucagon and epinephrine serve as covalent modifiers by activating protein kinase A which in turn phosphorylates, and deactivates pyruvate kinase.
  • In contrast, the secretion of insulin in response to blood sugar elevation activates phosphoprotein phosphatase I, causing the dephosphorylation and activation of pyruvate kinase.
  • ¬†This regulation system is responsible for the avoidance of a futile cycle through the prevention of simultaneous activation of pyruvate kinase and enzymes that catalyze gluconeogenesis.

Futile cycle: A futile cycle occurs when two metabolic pathways run simultaneously in opposite directions and have no overall effect other than to dissipate energy in the form of heat.

High glucose: dephosphorylated form of PK (active)

Low glucose: phosphorylated form of PK (Inactive)

Figure 2: Activation and inactivation of Pyruvate kinase by covalent modification

Hormonal control

  • In order to prevent a futile cycle, glycolysis and gluconeogenesis are heavily regulated in order to ensure that they are never operating in the cell at the same time.
  • As a result, the inhibition of pyruvate kinase by glucagon, cyclic AMP and epinephrine, not only shuts down glycolysis, but also stimulates gluconeogenesis.
  • Alternatively, insulin interferes with the effect of glucagon, cyclic AMP and epinephrine, causing pyruvate kinase to function normally and gluconeogenesis to be shut down.

When blood glucose level is low: Glucagon-triggered cyclic AMP cascade leads to the phosphorylation of pyruvate kinase, which diminishes its activity.

(cAMP activates protein kinase A which inturn phosphorylate Pyruvate kinase L isoform)

This mechanism prevents the liver from consuming glucose by glycolysis when the blood glucose concentration is low; instead, liver sparing it for export to the brain and other organs.

When blood glucose level is high: Insulin secretion activates a protein phosphatase (PP) which inturn dephosphorylates pyruvate kinase and activates it. As result glycolysis continues.

Figure 3: Regulation of PK activity by glucagon and insulin

Point to remember: The muscle isozyme (M form) is not affected by this phosphorylation mechanism.

  • In muscle, the effect of increased [cAMP] is quite different.
  • In response to epinephrine, cAMP activates glycogen breakdown and glycolysis and provides the fuel needed for the fight-or-flight response.

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