Models of the metal ion binding sites of native ZnZn and of cadmium-substituted ZnCd and CdCd phosphotriesterase, including full amino acid side chains, were geometry optimized with quantum mechanical methods, with effective fragment potentials (EFP) representing the protein environment surrounding the active site. One to three water molecules were included in the active site in addition to the bridging hydroxide. Comparison with recent X-ray diffraction results Benning, M. M.; Shim, H.; Raushel, F. M.; Holden, H. M. Biochemistry 2001, 40, 2712-22 is hindered by the presence of ethylene glycol molecules in the active site. We suggest that the ethylene glycol required for crystallization distorts the structure of the water network in the active site and that the theoretical structures provide a better description of the system in aqueous solution. Cd-113 NMR isotropic shielding calculations were performed to analyze the experimentally determined chemical shifts at 212 and 116 ppm, respectively, for the CdCd enzyme. The calculated isotropic shieldings correlate with the coordination number of the metal ions, indicating that the CdCd enzyme has one more ligand at the high shift site than at the low shift site. Theoretically, a number of energetically close structures are found for the CdCd structure. Formally, one of these agrees with the X-ray structure and is supported by the NMR assignment. For the hybrid ZnCd enzyme, the most stable theoretical structure is Cd1Zn2, with the metal bound to the Od1 of the carboxylate of the first-shell aspartate designated M 1, but the energy difference between Cd1Zn2 and the lowest energy Zn1Cd2 structure is only about 2 kcal/mol and decreasing with the addition of water molecules. The Zn1Cd2 arrangement is found experimentally.