Integration and global analysis of isothermal titration calorimetry data for studying macromolecular interactions
Isothermal titration calorimetry (ITC) is a powerful and widely used method to measure the energetics of macromolecular interactions by recording a thermogram of differential heating power during a titration. However, traditional ITC analysis is limited by stochastic thermogram noise and by the limited information content of a single titration experiment. Here we present a protocol for bias free thermogram integration based on automated shape analysis of the injection peaks, followed by combination of isotherms from different calorimetric titration experiments into a global analysis, statistical analysis of binding parameters and graphical presentation of the results. This is performed using the integrated public domain software packages NITPIC, SEDPHAT and GUSSI. The recently developed low noise thermogram integration approach and global analysis allow for more precise parameter estimates and more reliable quantification of multisite and multicomponent cooperative and competitive interactions. Titration experiments typically take 1 2.5 h each, and global analysis usually takes 10 20 min.
Nature Protocols