5. Ligand Binding

Protein/Ligand Interactions

Ligand binding = many proteins will bind other molecules REVERSIBLY

• bound molecules = ligands

  • ligands can be small molecules (ex. metal ions) or other proteins

• NON-covalent - bc we need it to be reversible

• occurs at the binding site, which is COMPLEMENTARY to the ligand

  • via: H-bonds, ionic interactions, phobic interactions, van Der Waals interactions

ex. (+) ligand, thus expect (-) charge in binding site

• may be cooperative:

  • NON-cooperative: see hyperbolic binding curves

  • Cooperative: see sigmoidal binding curves (allostery), multiple binding sites (4° structure)

Protein Binding site Complementary

→ binding sites will have complementary interactions for the ligand:

• charge

• H-bonding

• non-polar/PHOBIC

• shape/size

ex.

ex. complementary H-bonding

Equation for Describing the Binding of Ligands

→ think of simple dissociation & association equilibrium

Kd = [ L ] needed to have 50% of ligand bound to protein

• dissociation constant

• expresses the strength of interaction, thus reflects affinity

  • increased affinity of ligand to protein = decreased Kd

    • thus, less dissoc. bc protein & ligand have high affinity

note: changing PH/buffer affects Kd/affinity

Fraction of protein bound to ligand: θ “theta”

• generates a hyperbolic relationship b/w [ligand]

• as [L] increases = θincreases

• limit: θ = 0-1

  • θ NEVER = 1, thus fraction of protein bound to ligand NEVER = 1

→ derivation:

→ formula for relationship b/w Kd ([L] at 50% bound) & θ (fraction of protein bound to ligand):

ex. if [ L ] = Kd, meaning 50% of ligand is bound, then θ = 1Kd / 2Kd =0.5

Graphical Representation of Ligand Binding

→ shows that as [ligand] increases, the fraction bound to the protein (θ) increases & that Kd is when 50% of protein is bound to ligand

• fraction of protein bound to ligand NEVER reaches 1 - thus the protein is never fully bound

Q. In the presence of 2.0µM Ni2+, a nickel-binding protein is found to be 25% saturated with nickel ( θ = 0.25 ). What is the value of Kd?

• talking about ligand binding

• only 25% of protein is bound

  • if it was 50% bound it would be equal to Kd → THUS, Kd here ≠ [ L ] = 2.0µM

1. 
   

2. 
   

3. Kd = 6.0µM

• [ L ] needed to have 50% of ligand bound to protein