Search

Microcalorimetry and high-performance liquid chromatography (h.p.l.c.) have been used to conduct a thermodynamic investigation of the biochemical reaction {phospho enolpyruvate(aq) +d-erythrose 4-phosphate(aq) +H2O(l) =2-dehydro-3-deoxy-d- arabino-heptonate 7-phosphate(aq) +phosphate(aq)}, the first step in the metabolic pathway leading to the biosynthesis of chorismate. DAHP synthase, the enzyme used to carry out this reaction, was prepared by using molecular biology techniques and then isolated in a highly purified form. The h.p.l.c. was used to establish that this reaction went to completion and that the apparent equilibrium constant K′( T=298.15 K, pH ≈8.2) is >1.4 · 10 3. The calorimetrically determined molar enthalpy for this biochemical reaction isΔrHm(cal) =−(67.7 ±1.5)kJ · mol−1atT=298.15 K, pH =8.18, and ionic strengthIm=0.090 mol · kg−1. Use of this result together with a chemical equilibrium model led to the standard molar enthalpy of reactionΔrHmo=−(70.0 ±3.0)kJ · mol−1atT=298.15 K and Im=0 for the chemical reference reaction {phospho enolpyruvate 3−(aq) +d-erythrose 4-phosphate 2−(aq) +H2O(l) =2-dehydro-3-deoxy-d- arabino-heptonate 7-phosphate 3−(aq) +HPO42−(aq)}. Use of an estimated value of the standard molar entropy changeΔrSmofor this reaction together with the measured value of ΔrHmoyielded the standard molar Gibbs free energy change ΔrGmo≈−39 kJ · mol−1and K≈7 · 106for the chemical reference reaction at T=298.15 K andIm=0. Some insight into the thermochemistry of this reaction has been obtained by assessing the contributions to ΔrHmothat correspond to the loss of a phosphate from phospho enolpyruvate and to the addition of a pyruvate group tod-erythrose 4-phosphate.