Calorimetry

Abstract

In biology, particularly in studies relating the structure of biopolymers to their functions, two of the most important questions are: (1) how tightly does a small molecule bind to a specific interaction site? and (2) if the small molecule is a substrate and is converted to a product, how fast does the reaction take place? Because almost any chemical reaction or physical change is accompanied by a change in heat or enthalpy, an isothermal titration calorimeter is an ideal instrument to measure either how much of a reaction has taken place or the rate at which a reaction is occurring. In contrast to optical methods, calorimetric measurements can be done with reactants that are spectroscopically silent (a chromophore or fluorophore tag is not required), can be done on opaque, turbid or heterogeneous solutions (e.g. cell suspensions) and can be done over a range of biologically relevant conditions (temperature, salt, pH, etc ). Key Concepts: Almost all chemical reactions are accompanied by a change in heat or enthalpy. Calorimetry can determine the complete set of thermodynamic parameters that characterise binding reactions, for example K, Δ G °, Δ H °, Δ S ° and Δ C p . Binding reactions are typically pH, salt and temperature dependant.

Revisiting Zinc Coordination in Human Carbonic Anhydrase II

“…Data collected in ACES can be simply fit using a one-site binding model (Figure ); however, serious metal dilution issues are observed in other buffers, which require a more sophisticated analysis. Initially we described these curves as multiple binding events . However, these isotherms can be better described as single-site binding events with a number of complicating dilution equilibria.…”

“…An illustrative case is the data collected in 100 mM PIPES at pH 7.4, shown in Figure . Initially this unique curve shape was assigned to two Zn 2+ binding sites associated with CA, where the second binding site was described as an adventitious metal binding site . The curvature of this endothermic portion of the raw data was actually the result of metal-dilution heat, although the origin of this endothermic dilution process is still unclear.…”

“…Initially we described these curves as multiple binding events. 29 However, these isotherms can be better described as single-site binding events with a number of complicating dilution equilibria. An illustrative case is the data collected in 100 mM PIPES at pH 7.4, shown in Figure 4.…”

“…Initially this unique curve shape was assigned to two Zn 2+ binding sites associated with CA, where the second binding site was described as an adventitious metal binding site. 29 The curvature of this endothermic portion of the raw data was actually the result of metal-dilution heat, although the origin of this endothermic dilution process is still unclear. The shape of the integrated raw isotherm (Figure 4, blue trace) past the equivalence point can be corrected using heats associated with the background titration of Zn(NO 3 ) 2 into PIPES buffer under the same condition (Figure 4, red trace).…”