The Equilibrium Constants of the Adenosine Triphosphate Hydrolysis and the Adenosine Triphosphate-Citrate Lyase Reactions

• Published in: The Journal of biological chemistry 1973-10, Vol.248 (20), p.6966-6972
• Main Authors: Guynn, Robert W., Veech, Richard L.
• Format: Article
• Language: English
• Subjects: Acetyl Coenzyme A; Acetyltransferases - metabolism; Adenosine Triphosphate - metabolism; ATP Citrate (pro-S)-Lyase - metabolism; Citrate (si)-Synthase - metabolism; Clostridium - enzymology; Escherichia coli - enzymology; Hydrolysis; Magnesium; Mathematics; Osmolar Concentration; Oxo-Acid-Lyases - metabolism; Phosphates; Phosphotransferases - metabolism; Protein Binding; Thermodynamics
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)43347-4

The observed standard free energy change (ΔG0obs) for the hydrolysis of the terminal pyrophosphate bond of ATP has been experimentally determined under physiological conditions using an entirely new set of reactions. The observed equilibrium constant (Kobs) for the combined reactions of acetate kinase (EC 2.7.2.1) and phosphate acetyltransferase (EC 2.3.1.8) has been determined at 38°, pH 7.0, ionic strength 0.25, and varying free [Mg2+]. The Kobs of these combined reactions reflects the difference between ΔG0obs for the hydrolysis of acetyl-CoA and the ΔG0obs for the hydrolysis of ATP. Using Σ and square brackets to indicate total concentration, Kobs = [ΣADP][ΣPi]/[ΣATP] x [Acetyl-CoA]/[ΣAcetate][CoA] The observed value of this combined equilibrium constant varies with free [Mg2+], being 0.984 ± 0.009 when [Mg2+] = 0 and 0.218 ± 0.002 when free [Mg2+] = 10-3m. The ΔG0obs for the hydrolysis of acetyl-CoA is virtually unaffected by the free [Mg2+] and has been previously determined to be -8.54 Cal per mole (-35.75 kJ per mole) under the same conditions of temperature, pH, and ionic strength. Therefore at pH 7.0, at ionic strength 0.25, at 38°, and taking the standard state of liquid water to have activity = unity ([H2O] = 1) the ΔG0obs for the reaction [see PDF for equation] can be calculated to be -8.53 Cal per mole (-35.69 kJ per mole) at [Mg2+] = 0 and -7.60 Cal per mole (-31.80 kJ per mole) at [Mg2+] = 10-3m. The corresponding values of Kobs for the ATP hydrolysis reaction are 9.86 x 105m ([Mg2+] = 0) and 2.19 x 105m ([Mg2+] = 10-3m). Equations have been developed for calculating from the experimental data the ΔG0obs of ATP hydrolysis at different free magnesium and hydrogen ion concentrations. The Kobs of ATP hydrolysis has been used in combination with the Kobs of the citrate synthase reaction (EC 4.1.3.7) to calculate the Kobs of the ATP-citrate lyase reaction (EC 4.1.3.8) Kobs = [ΣADP][Acetyl-CoA] [ΣPi] [ΣOxaloacetate]/[ΣATP] [CoA] [ΣCitrate] Under the same near physiological conditions of 38°, pH 7.0, and ionic strength 0.25, the value of Kobs for the ATP-citrate lyase reaction was found to be very sensitive to the free [Mg2+], being 0.975 m at [Mg2+] = 0 and 0.0985 m when free [Mg2+] = 10-3m.