Mechanism of Thermal Decomposition of Carbamoyl Phosphate and Its Stabilization by Aspartate and Ornithine Transcarbamoylases

• Published in: Proceedings of the National Academy of Sciences - PNAS 2008-11, Vol.105 (44), p.16918-16923
• Main Authors: Wang, Qin, Xia, Jiarong, Guallar, Victor, Krilov, Goran, Kantrowitz, Evan R.
• Format: Article
• Language: English
• Subjects: Active sites; Aspartate Carbamoyltransferase - chemistry; Aspartate Carbamoyltransferase - metabolism; Biochemistry; Biological Sciences; Carbamyl Phosphate - chemistry; Carbamyl Phosphate - metabolism; Catalytic Domain; Chemical compounds; Chemical reactions; Computer Simulation; Crystallography, X-Ray; Dihedral angle; Enzymes; Escherichia coli - enzymology; Hydrogen bonds; Kinetics; Models, Molecular; Molecules; Organisms; Ornithine Carbamoyltransferase - chemistry; Ornithine Carbamoyltransferase - metabolism; Phosphates; Potential energy; Protons; Quantum mechanics; Substrate Specificity; Thermal decomposition; Thermodynamics
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0809631105

Carbamoyl phosphate (CP) has a half-life for thermal decomposition of 5,000. Both of these transcarbamoylases use an ordered-binding mechanism in which CP binds first, allowing the formation of an enzyme·CP complex. To understand how the enzyme·CP complex is able to stabilize CP we investigated the mechanism of the thermal decomposition of CP in aqueous solution in the absence and presence of enzyme. By quantum mechanics/molecular mechanics calculations we show that the critical step in the thermal decomposition of CP in aqueous solution, in the absence of enzyme, involves the breaking of the C-O bond facilitated by intramolecular proton transfer from the amine to the phosphate. Furthermore, we demonstrate that the binding of CP to the active sites of these enzymes significantly inhibits this process by restricting the accessible conformations of the bound ligand to those disfavoring the reactive geometry. These results not only provide insight into the reaction pathways for the thermal decomposition of free CP in an aqueous solution but also show why these reaction pathways are not accessible when the metabolite is bound to the active sites of these transcarbamoylases.