Cooperative Ligand Binding

Cooperative Ligand Binding (Definition)

cooperatively refers to a situation where the presence of one affects others

in the context of ligand binding, cooperatively refers to a scenario when ligand binding at one binding site affects binding of a ligand at another binding site.

Cooperative ligand binding depends on the presence of more than one binding site in a molecule or complex of molecules.

Cooperatively is very commonly observed in proteins that form complexes containing multiple protein chains, which each have a ligand binding site.

cooperative ligand binding refers to

cooperatively refers to a situation where the presence of one affects others

in the context of ligand binding, cooperatively refers to a scenario when ligand binding at one binding site affects binding of a ligand at another binding site.

Cooperative Ligand Binding depends on

Cooperative ligand binding depends on the presence of more than one binding site in a molecule or complex of molecules.

Cooperative Ligand Binding commonly observed in

Cooperatively is very commonly observed in proteins that form complexes containing multiple protein chains, which each have a ligand binding site.

Positive Cooperative Ligand Binding

Positive cooperativity in ligand binding occurs when the affinity of a ligand binding at a second binding site is higher than the affinity of a ligand binding at the first binding site.

This means that the Kd or dissociation constant of ligand binding at the second binding site Kd2 is lower than the Kd1 (ligand binding at first binding site).

Kd2<Kd1

Positive Cooperative Ligand Binding graph

shape of graph

affinity for T vs R

low, medium, high ligand concentration

observed binding for ligand binding to a ligand with positive cooperatively is sigmoidal or S- shaped

observed binding modeled by a low affinity T state and high affinity R state

In this model at a low ligand concentration

• most molecules in population in T state

• observed binding curve and apparent ligand affinity would most closely resemble the binding of the low affinity T state.

At high ligand concentrations

• Most molecules in population would be in the R state

• observed binding curve and apparent ligand affinity would most closely resemble the binding of the high affinity R state.

At intermediate ligand concentrations

• there would be a mix of molecules in the R state and molecules in the T state,

• observed binding curve and apparent ligand affinity would be in between the low affinity T state and the high affinity R state

At intermediate ligand concentrations the binding is highly sensitive to ligand concentration. Small changes in lag in concentration can result in relatively large changes in fractional saturation.

Positive Cooperative Ligand Binding graph shape

observed binding for ligand binding to a ligand with positive cooperatively is sigmoidal or S- shaped

observed binding modeled by a low affinity T state and high affinity R state

Positive Cooperative Ligand Binding low

In this model at a low ligand concentration

• most molecules in population in T state

• observed binding curve and apparent ligand affinity would most closely resemble the binding of the low affinity T state.

Positive Cooperative Ligand Binding intermediate

At intermediate ligand concentrations

• there would be a mix of molecules in the R state and molecules in the T state,

• observed binding curve and apparent ligand affinity would be in between the low affinity T state and the high affinity R state

Positive Cooperative Ligand Binding high

At high ligand concentrations

• Most molecules in population would be in the R state

• observed binding curve and apparent ligand affinity would most closely resemble the binding of the high affinity R state.

At intermediate ligand concentrations

the binding is highly sensitive to ligand concentration. Small changes in lag in concentration can result in relatively large changes in fractional saturation.

Positive Cooperative Ligand Binding graph axis

X: [Ligand]

Y: fractional saturation

Protein Dimer Example

As an example, let's consider a protein dimer consisting of two subunits. The dimer can exist in a low affinity T state represented by the red squares or a high affinity R state represented by blue circles.

A dimer can have subunits both in the R state or both in the T state. A dimer can switch between the T state and R states, and there is an equilibrium between the two states. Each subunit has one binding site for the ligand L.

Stabilization of the R State by Ligand Binding

The key to understanding positive cooperative binding is that binding of the ligand lowers the free energy of the higher affinity R state, which leads to stabilization of the R state.

This means that the R state becomes more populated at equilibrium, at increased concentrations of ligand

in the absence of ligand or in the presence of low concentrations of lag in the low affinity T state will be favored. Favourability of the T state over the R state can be indicated by an arrow pointing towards the T state that is longer than the arrow pointing towards the R state. The fractional saturation of the ligand binding sites will be relatively low.

At high ligand concentrations the high affinity R state will be favored. Favourability of the R state over T state can be indicated by an arrow pointing towards the R state that is longer than the arrow pointing towards the T state. The fractional saturation of the ligand binding sites will be relatively high

at intermediate lag in concentrations there will be a mix of molecules in the T state and R state. The observed binding affinity and fractional saturation of a population of molecules at an intermediate concentration of ligand will be intermediate between the low affinity T state and the high affinity R state.

Stabilization of the R State by Ligand Binding 1

The key to understanding positive cooperative binding is that binding of the ligand lowers the free energy of the higher affinity R state, which leads to stabilization of the R state.

This means that the R state becomes more populated at equilibrium, at increased concentrations of ligand

Stabilization of the R State by Ligand Binding low

in the absence of ligand or in the presence of low concentrations of lag in the low affinity T state will be favored.

Favourability of the T state over the R state can be indicated by an arrow pointing towards the T state that is longer than the arrow pointing towards the R state.

The fractional saturation of the ligand binding sites will be relatively low.

Stabilization of the R State by Ligand Binding intermediate

at intermediate lag in concentrations there will be a mix of molecules in the T state and R state.

The observed binding affinity and fractional saturation of a population of molecules at an intermediate

concentration of ligand will be intermediate between the low affinity T state and the high affinity R state.

Stabilization of the R State by Ligand Binding high

At high ligand concentrations the high affinity R state will be favored.

Favourability of the R state over T state can be indicated by an arrow pointing towards the R state that is longer than the arrow pointing towards the T state.

The fractional saturation of the ligand binding sites will be relatively high

Visualizing Cooperative Binding with Panels

Panels are useful to visualize the ligand binding and conformational state of molecules in a population at different ligand concentrations

at low levels of ligand most of the molecules are in the T state. Most of the ligand binding sites do not have ligand bound, as shown in panel one

at intermediate levels of ligand concentration there is a mix of molecules in the R state and T state. In panel two half of the molecules are in the R state and half are in the T state, and exactly half of the potential ligand binding sites are bound by ligand.

At high levels of ligand most of the molecules are in R state and most of the ligand binding sites have a ligand bound, as shown in panel three.

Summary of Cooperative Ligand Binding

• Cooperative ligand binding involves two ligand binding sites

• Ligand binding with positive cooperativity is when affinity at the second binding site is higher than at the first site (Kd2<Kd1)

• Ligand binding stabilizes the high affinity R-state

• At low ligand concentrations, the low affinity T-state is more populated

• At high ligand concentrations, the high affinity R-state is more populated

• At intermediate ligand concentrations, there is a mix of molecules in the R-state and T-state and an intermediate affinity

Cooperative Ligand Binding

• The ligand binding at one binding site affects binding of a ligand at another binding site

• Depends on more than one binding site in a molecule or molecular complex

• Common when multiple protein chains each with binding sites for a ligand associated to form complexes

Ligand binding with positive cooperativity

affinity at the second binding site is higher than at the first site (Kd2<Kd1)

Cooperative ligand binding can be modeled as two states

Dimer with two binding sites with positive cooperative ligand binding

Ligand binding stabilizes the high affinity R-state