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Strategies for assessing proton linkage to bimolecular interactions by global analysis of isothermal titration calorimetry data

Tuesday, October 09, 2012 — Poster Session I

1:00 p.m. – 3:00 p.m

Natcher Conference Center, Building 45




  • H Zhao
  • N.P. Coussens
  • P Schuck


Isothermal titration calorimetry (ITC) is a traditional and powerful method for studying the linkage of ligand binding to proton uptake or release. The theoretical framework has been developed previously and numerous applications have appeared. In the current work, we explored strategic aspects of experimental design and data interpretation. We simulated families of ITC data sets that embed different strategies with regard to the number of experiments, and experimental conditions (pH, buffer ionization enthalpy, temperature). We then re-analyzed the families of data sets in the context of global analysis, employing a proton linkage binding model implemented in the global data analysis platform SEDPHAT, and examined the information content of all data sets by a detailed statistical error analysis of the parameter estimates. In particular, we studied the impact of different assumptions about the knowledge of the exact macromolecular concentrations, which in practice presents an experimental limitation for many systems. Our results show that the global analysis can yield reliable estimates of the thermodynamic parameters for intrinsic binding and protonation, and that in the context of the global analysis the exact molecular component concentrations may not be required. Additionally, the benefit of conducting experiments at a range of temperatures has been illustrated.

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