Chemistry Reference
In-Depth Information
protein and the ligand is the saturation transfer difference technique. With galec-
tin - 1 and its growth -regulatory interaction with ganglioside GM1 as an example
(for details on ganglioside and tumor biology of this interaction, please see Chap-
ters 25 and 30), saturation transfer from the protein to the carbohydrate ligand
was picked up for the terminal Gal and GalNAc units [13]. This technique has
been adapted to use the fl uorine atom (
19
F) as sensor [14]. Close contact between
lectin and ligand underlies this transfer process, allowing us to map spatial param-
eters. However, the only technique that directly determines binding enthalpies is
isothermal titration calorimetry (ITC) (for techniques to measure carbohydrate-
binding specifi cities, please see Chapter 14 ).
ITC measures the heat evolved after addition of a ligand to a receptor as a func-
tion of ligand concentration [15]. A single titration experiment can yield a complete
binding isotherm from which the enthalpy of binding (
H
° ), the association con-
stant (
K
a
) and the stoichiometry (
n
) can be directly determined. Then, the entropy
of the binding (
Δ
Δ
S °
) can be calculated from the expression:
ΔΔ Δ
GHTS
°=
°−
°=−
T
ln
,
a
where
R
is the universal gas constant and
T
is the absolute temperature. By way
of illustration, Figure 13.8 shows the calorimetric titration of human galectin-1
Figure 13.8
Calorimetric titration of human
galectin - 1 with
N
- acetyllactosamine. The upper
panel shows the experimental measurement of
the heat released after sequential addition of
5 -
298.08 K. The lower panel shows the plot of this
heat as a function of the sugar : protein molar
ratio. The continuous line corresponds to the
best fi t of the experimental data in a one-set-of-
sites model. The thermodynamic parameters
derived from the analysis are given.
l aliquots of 10 mM
N
- acetyllactosamine to
a 0.1 mM solution of human galectin-1 at
μ
