Binding Properties of Myoglobin and Hemoglobin

Dissociation constant (Kd)

The ligand concentration at which half of the protein's receptor binding sites are occupied.

Formula for Kd

Kd = [P][L] / [PL]

Calculation of Kd

From a binding isotherm.

Saturation in binding studies

Specific binding.

[L] when f = 0.5

[L] = Kd

Assumption in Scatchard analysis

[L] ≈ [L]total when [L] is much greater than [P].

Total ligand concentration formula

[L]total = [L]free + [L]bound

[L]bound without non-specific binding

[L]bound = [PL]

[PL] in Scatchard analysis

[PL] is negligible when [PL] is very low relative to total [L].

Myoglobin

A monomeric protein found in muscle, primary oxygen-carrying pigment.

Red color of meat

Bound Fe²⁺ complexed in a haem group in myoglobin.

Structure of myoglobin

Eight helices + haem group.

Fe²⁺ in myoglobin

Complexed into protoporphyrin and chelated by four nitrogen atoms of the tetrapyrrole ring.

Haem in myoglobin

Held in place by hydrophobic interactions.

Myoglobin's O2 binding curve

Rectangular hyperbolic curve (one binding site per protein).

Fraction of occupied binding sites (f)

f = [PL] / ([P] + [PL])

Haemoglobin

An oxygen-transport metalloprotein in red blood cells.

Red color of blood

Haemoglobin.

Subunits of haemoglobin

4 (2 alpha and 2 beta subunits).

Haem groups in haemoglobin

4, one per subunit.

States of haemoglobin

Oxy-haemoglobin (4 O2 bound), Deoxy-haemoglobin (0 O2 bound).

Haemoglobin's O2 binding curve

Sigmoidal, indicating cooperative binding.

Curves of haemoglobin's binding curve

High affinity and low affinity binding curves.

O2 release at tissue pO2 levels

Cooperative binding enhances O2 unloading in tissues.

Equation for sigmoidal O2 binding curve

The Hill equation.

Hill equation

Describes the fraction of a macromolecule saturated by ligand as a function of ligand concentration.

Hill coefficient n = 1

Indicates no cooperativity.

Hill coefficient n > 1

Indicates positive cooperativity.

Hill coefficient n < 1

Indicates negative cooperativity.

Lower pH in tissues

Due to metabolic production of CO2, which releases protons upon hydration.

Bohr effect

The phenomenon where lower pH stabilizes deoxy-Hb and promotes O2 release to tissues.

Bohr effect and O2 delivery

Binding of protons stabilizes the low-affinity form of haemoglobin, increasing O2 unloading.