Two structural forms of the active site in Cd2+-containing phosphotriesterases (PTE), i.e., Cd2+/Cd2+ -PTE and Zn2+/Cd2+-PTE, were determined by performing ab initio and density functional theory calculations on active site models. The calculated results indicate that the second bridging ligand in the active site of the recently reported high-resolution X-ray crystal structures of Cd2+/Cd2+ -PTE and Zn2+/Cd2+ -PTE should be a hydroxide ion, rather than a water molecule. The protonated active site structures of the Cd2+-containing enzymes are all significantly different from that of Zn2+/Zn2+ -PTE. In Cd2+/Cd2+-PTE and Zn2+/Cd2+- PTE, the second bridging ligand always simultaneously coordinates to the two metal ions, whether it is a hydroxide ion or a water molecule. The pK(a) values calculated for the second bridging ligand are in good agreement with the observed kinetic pK(a) values for both the Cd2+- and Zn2+-substituted enzymes. The calculated results strongly support the conclusion that the observed kinetic pK(a) is actually the pK(a) of the second bridging ligand, and strongly support the previously postulated catalytic mechanism of PTE involving a bridging hydroxide that attacks the phosphorus center of the substrate.