Functional energetic landscape in the allosteric regulation of muscle pyruvate kinase. 1. Calorimetric study
Abstract
The energetic landscape of the allosteric regulatory mechanism of RMPK was characterized by isothermal titration calorimetry (ITC). ITC data for RMPK binding to substrates PEP and ADP, for the allosteric inhibitor Phe, and for combination of ADP and Phe were globally analyzed. The thermodynamic parameters characterizing the linked-multiple-equilibrium system were extracted. Four novel insights were uncovered. (1) The binding preference of ADP for either the T or R state is temperature-dependent, namely, more favorable to the T and R states at high and low temperatures, respectively. Depending on the temperature, binding of ADP can regulate RMPK activity by favoring the enzyme to either the R or T state. (2) The binding of Phe is negatively coupled to that of ADP (3) The release or absorption of protons linked to the various equilibria is specific to the particular reaction. (4) The R {--} T equilibrium is accompanied by a significant DeltaC(p).