Assessing Coupled Protein Folding and Binding Through Temperature-Dependent Isothermal Titration Calorimetry

Methods in Enzymology
Debashish SahuScott A Showalter

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. Significantly, when the effective ΔH° for the coupled folding and binding process is determined by ITC in a temperature series, the constant-pressure heat capacity change (ΔCp) associated with these coupled equilibria is experimentally accessible, offering a unique opportunity to investigate the driving forces behind them. Notably, each of these molecular-scale events is often accompanied by strongly temperature-dependent enthalpy changes, even over the narrow temperature range experimentally accessible for biomolecules, making single temperature determinations of ΔH° less informative than typically assumed. Here, we will document the procedures we have adopted in our laboratory for designing, executing, and globally analyzing temperature-dependent ITC studies of coupled folding and binding in IDP interactions. As a biolog...Continue Reading

Citations

Aug 24, 2016·The Journal of Physical Chemistry. B·Eric B Gibbs, Scott A Showalter
Jun 23, 2016·Biophysical Journal·Roderico AcevedoScott A Showalter

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