Gas‐solution microcalorimeter for determining heat binding curves

A gas‐solution reaction microcalorimeter has been developed for measuring the heat produced from a macromolecule in solution when the partial pressure of a reacting ligand is changed. The accumulated result of such stepwise heat processes enables the construction of a heat binding curve: namely the enthalpy per mole of reaction site versus the logarithm of the ligand activity [Gill, Richey, Bishop, and Wyman, Biophys. Chem. 21, 1 (1985)]. The high sensitivity (1 μcal) of the calorimeter is achieved by careful twinning and closed‐loop proportional control achieved through continuous computer monitoring. A 15‐μl liquid sample, for example 5‐mM heme hemoglobin, is held in the form of a thin layer by means of a filter paper membrane. This configuration facilitates rapid equilibrium with exposed gases and allows for the completion of a given heat process within the range of 10 min. Gas partial pressures are accurately produced by means of a precision dilution valve incorporated into the calorimeter. The automa...