measured total protein concentration as a result of inactive/modified/aggregated fractions, which do not bind the ligand or bind
with much weaker affinity. Similarly, the presence of contaminant
proteins that have been copurified along with the system of interest
will reduce the active concentration. These could be unrelated
contaminants or could be the binding partner of interest or a
related ligand.
3.5 Occam’s Razor
and the Use
of Advanced Data
Fitting
Fitting ITC data using the software provided with instruments is
very quick and simple compared to the measurement itself. These
fitting programs also allow more complex models of binding to be
considered (multiple sites, cooperativity, etc.) but these should only
be employed if there is good evidence from other techniques that
Fig. 4 Effect of errors in the measured concentration of lysozyme in the cell
(upper panel) or tri-acetyl glucosamine in the syringe (lower panel) on the fitted
K d (red), ΔH (blue), and stoichiometry n (green). Fitted values are plotted as a
fraction of the values determined in Fig. 2
146
Christopher M. Johnson