MicroCal VP-ITC - 3.1 Designing ITC Experiments

VP-ITC MicroCalorimeter User’s Manual
3.1 Designing ITC Experiments
For a ligand X binding to a single set of n identical sites on a macromolecule M, i.e.,
M + X = MX
MX + X = MX
2
. .
. .
. .
MX
n-1
+ X = MX
n
the single-site binding constant is
[filled sites]
K =
___________________
[empty sites][X]
and
∆G
o
= —R T lnK = ∆H
o
-T ∆S
o
Where ∆G
o
, ∆ H
o
and ∆S
o
are the free energy, enthalpy, and entropy change for single site
binding.
By non-linear least squares fit of calorimetric titration data, the parameters K, ∆H
o
, and n are
determined directly in a single experiment and ∆G
o
and ∆S
o
may then be calculated. Titration
calorimetry is the only technique capable of defining all of these parameters in a single
experiment resulting in nearly complete thermodynamic characterization of the interaction.
Measuring the binding isotherm at a second temperature allows additional determination of the
change in heat capacity of binding through the relation:
∆H
o
T2
- ∆H
o
T1
∆C
p
=
___________________________
T2
-
T1
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