Allosteric regulation and cooperativity: phosphofructokinase and AMP
Allosteric regulation means a molecule binds an enzyme at a site away from its active site (the pocket where the substrate binds) and changes the enzyme's shape and activity.
Remember
When a cell is short of energy, AMP (adenosine monophosphate, a lower-energy relative of ATP) rises and binds phosphofructokinase, the enzyme that commits glucose to glycolysis, away from its active site, which speeds glycolysis up. High ATP, citrate (a signal of plentiful energy) and an increased hydrogen ion concentration (a lower pH) inhibit the enzyme.
Every enzyme has an active site, the pocket where the reaction happens once the substrate has bound. An allosteric site is a different place on the enzyme (allosteric simply means at another site). A regulator that binds there changes the three-dimensional shape of the whole protein, which is called a conformational change. The new shape alters how well the active site grips the substrate or how fast it works, so the enzyme becomes more active with an allosteric activator and less active with an allosteric inhibitor.
In many allosteric enzymes, an inhibitor favours the T state, a shape with a lower affinity for the substrate, meaning it holds the substrate less readily. An activator favours the R state, which has a higher affinity, so the substrate is held more readily.
Many allosteric enzymes are built from several protein subunits (separate polypeptide chains), and each subunit has its own active site. In positive cooperativity, substrate binding at one active site increases the activity of the other active sites on the same enzyme. The response to rising substrate is then a steeper, S-shaped (sigmoidal) curve, so the enzyme can act like a sensitive switch. A substrate can itself act as an allosteric activator: binding at one active site increases the activity of the other active sites, which are away from that binding site.
Phosphofructokinase is a key regulatory enzyme of glycolysis, the main control point of the pathway. When ATP is spent, AMP rises and binds a regulatory site (separate from the active site), which stabilises the high-affinity form, so glycolysis speeds up and more ATP is made. Fructose-2,6-bisphosphate is a second activator.
Key facts
- Allosteric site
- a regulator binds at a site separate from the active site, and the enzyme changes shape, which alters how active it is.
- R and T states
- in many allosteric enzymes an activator favours the R state, with higher affinity for the substrate, and an inhibitor favours the T state, with lower affinity.
- Positive cooperativity
- binding of substrate at one active site makes the other active sites on the same enzyme more active.
- Curve
- with positive cooperativity, the response to rising substrate is S-shaped (sigmoidal), like a sensitive switch.
- Phosphofructokinase activation
- activators include AMP and fructose-2,6-bisphosphate.
- Phosphofructokinase inhibition
- inhibitors include high ATP, citrate and hydrogen ions (H+).
- Speed of control
- an allosteric regulator binds reversibly to enzyme that is already present, so control works within moments, with no new protein needed.
Exam traps
- ATP binding at a regulatory site does not activate phosphofructokinase, because ATP acts there as an allosteric inhibitor and signals that energy is plentiful, even though it also binds the active site as a substrate.
- Phosphorylation (adding a phosphate group) is not permanent activation, because phosphatases (enzymes that remove the group) can reverse it, and it is a different mechanism from binding at an allosteric site.
- Competition between an inhibitor and the substrate for the same site is competitive inhibition, not allosteric regulation, because an allosteric regulator binds at a different site.
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