Calorimetric Methods to Characterize the Forces Driving Macromolecular Association and Folding Processes

Global stability of an α-ketoglutarate-dependent dioxygenase (TauD) and its related complexes

“…More specifically, iron(II) confers approximately 2.2 °C in stability for the enzyme, as well as increasing the unfolding energy, Δ H cal , by approximately 12.7 kcal/mol (Table 2, Figure S9 reactions A and B1). Previous reports have shown there is no significant structural rearrangement when iron(II) binds to the 2-His-1-carboxylate facial triad [16,32,33], which suggests the overall increase in stability for the enzyme is primarily the change in enthalpy for binding iron(II). These data are supported by our CD data, which show no differences in structure between TauD and Fe-TauD; and by recently reported thermodynamic properties of metal binding to TauD, which indicate an overall binding enthalpy of −11.5 kcal/mol [21].…”

“…The taurine-bound enzyme is stabilized by substrate interactions with several amino acid residues within the active site pocket (Fig. 1 B , blue residues): the sulfonate moiety H-bonds with His70, Val102, and Arg270; the amine group forms H-bonds through two local water molecules; taurine’s C2 has hydrophobic interactions with Tyr73; and both the C1 and C2 atoms have hydrophobic interactions with Phe159 and Phe206 to aid in positioning the taurine molecule for efficient hydroxylation [32–36]. Thus, a conformational change induced by taurine supports a H-bonding network that holds the active site closed and positions hydrophobic residues away from solvent [35].…”

Biophysics of Nucleic Acids Celebrating the 75th Birthday of Professor Kenneth J. Breslauer

“…Conversely, the binding entropy includes favorable (e.g., desolvation and release of water molecules to bulk solvent) and unfavorable (e.g., conformational and/or motion restrictions) contributions arising from both the ligand and target (as reviewed in [8]). Acquisition of the requisite thermodynamic binding profiles as a function of temperature affords evaluation of heat capacity changes (∆C p ) accompanying the binding process [131,132]. This extra-thermodynamic information is essential for elucidating binding processes that are coupled with desolvation and/or folding.…”

“…In this respect, ITC protocols must be designed to Life 2022, 12,1438 derive the requisite intrinsic thermodynamic data by conducting measurements in an array of buffers with distinct ionization enthalpies at various pH. The calorimetric experiments should be conducted over a sufficiently broad temperature range to account for heat capacity changes involved in the association process (as reviewed in [131]).…”

Microcalorimetry of Macromolecules: The Physical Basis of Biological Structures

“…A combination of calorimetric and spectroscopic approaches led to the resolution of nearest neighbor contributions [22], which culminated in the development of predictive capabilities [25] that find numerous applications in the fields of biophysics, biochemistry, and molecular biology. Recent progress in the development of ultra-sensitive differential scanning calorimeters (DSC) (for reviews refer to [26,27]), coupled with advances in oligonucleotide synthetic capabilities, have facilitated measurements of DNA duplex energetics with improved accuracy and reliability. Considering the abundance of empirical data and vast array of applications emerging from the analysis of nucleic acid energetics, there are a number of fundamental questions that remain unresolved.…”