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2 Purification of FCP1 and Rap74 3. 2 Calorimetric Parameters 4. 3 Global Analysis of Temperature-Dependent Data Sets 5. Summary Acknowledgments References 24 27 27 28 29 29 29 31 31 34 38 41 43 43 Abstract Broad interest in the thermodynamic driving forces of coupled macromolecular folding and binding is motivated by the prevalence of disorder-to-order transitions observed when intrinsically disordered proteins (IDPs) bind to their partners. Isothermal titration calorimetry (ITC) is one of the few methods available for completely evaluating the thermodynamic parameters describing a protein–ligand binding event.

In this model, TH ¼ 295 K is the temperature at which the enthalpy of hydrophobic interactions vanishes, while TS ¼ 386 is the temperature at which the entropy of hydrophobic interactions vanishes. Within the context of this model, ΔCp is assumed to describe hydrophobic effects. , ΔH0 and ΔS0 are interpreted to describe contributions from electrostatic interactions. When TH ¼ 295 K and TS ¼ 386 K are predetermined by the Baldwin model, the global fit depends upon three parameters. 01 kJ/mol/K (Fig.

This allows discrimination of inhibition mechanism (competitive, noncompetitive, or mixed) and, for competitive inhibitors, determination of KI for each compound from the primary screening data. 4 mL) and long measurement times (10–30 min). Nanocalorimeters can overcome these limitations of conventional ITC. , 2004). Each detector cell on the array consists of two identical thermistor sensing elements connected in a Wheatstone bridge configuration that provides a differential temperature measurement between the sample specimen and reference specimen (Fig.