biochemistry: non-enzymatic proteins

3 areas where binding occur

between enzymes and substrates, receptors and signal molecules, and transporters and small molecules (for transport)

ligand

small molecule that binds to a protein

ligand binding

reversible process that involves noncovalent (weak) interactions; allows for protein and ligand to be reused

association constant (K_a)

equilibrium constant for the association (binding) of the protein

equation for K_a

[PL]/[P][L]

dissociation constant

the equilibrium constant for the dissociation of ligand from protein-ligand complex

formula for K_d

[P][L]/[PL]

a larger K_d indicates

more of the [P][L] and low binding affinity

fractional saturation (𝛳)

the fraction of occupied protein binding sites

fractional saturation equation

occupied binding sites/total binding sites

a lower K_d indicates

higher affinity of protein for ligand and a higher binding affinity

myoglobin (Mb)

concentrated in muscle tissue, functions as a storage depot for O₂

hemoglobin (Hb)

major protein in red blood cells, transports O₂ in blood from the lungs to the tissues

globin fold

8 𝛼-helix fold up into a globular structure; myoglobin and hemoglobin

heme

prosthetic group, Fe²⁺ porphyrin complex, O₂ will only bind to its reduced state

heme

prosthetic group, Fe²⁺ porphyrin complex, O₂ only binds to its reduced state (Fe²⁺)

myoglobin structure

consists of a single polypeptide chain, monomer (1 subunit), 8 𝛼-helices, 1 heme

hemoglobin structure

dimer of 𝛼𝛽 dimers (4 subunits), 32 𝛼-helices, 4 hemes

myoglobin O₂ binding curve

hyperbolic

hemoglobin O₂ binding curve

sigmoidal

hemoglobin’s 2 major conformational states

tense and relaxed state

deoxyhemoglobin (T state)

O₂ unbound form, low O₂ affinity, Fe²⁺ doesn’t fit well in the heme, puckered heme

oxyhemoglobin (R state)

O₂ bound form, high O₂ affinity, electrons are redistributed, changes the size of the ion, planar heme

collaborative binding

binding of a ligand to a macromolecule increases the affinity of remaining binding sites for subsequent ligands

concerted model

the tetramer exists in either T state or R state, ligand binding shifts the entire tetramer simultaneously from T to R (or R to T)

sequential model

the tetramer can contain both T and R states, affected by L binding, ligand binding to one subunit induces a conformational change of adjacent subunit to the R state (has different binding affinity than original state)

hill plots

quantitative assessment of cooperativity