The heat of binding the serine protease, porcine
pancreatic elastase, by the inhibitor, turkey ovomucoid
third domain, is dependent on the presence of inorganic
phosphate. This dependence is saturable and can be accurately
modeled as the phosphate binding to a single site on the
protease–inhibitor complex; thus, the elastase–ovomucoid
system provides a unique opportunity to study phosphate–protein
interactions. We have used isothermal titration calorimetry
to investigate this binding, thereby providing one of the
few complete thermodynamic characterizations of phosphate
interacting with proteins. The binding is characterized
by a small favorable ΔG°, a large unfavorable
ΔH°, and a positive ΔCp,
thermodynamics consistent with the release of water being
linked to phosphate binding. These measurements provide
insight into the binding of phosphotyrosine containing
peptides to SH2 domains by suggesting the energetic consequences
of binding phosphate free from other interactions.