Phosphotriesterases active site, structure

A phosphotriesterase isolated from the soil bacterium Pseudomonas diminuta is the best characterized enzyme of this type. There is evidence for the presence of two active site Zn2+ ions in vivo. A crystal structure of the dinuclear Cd2+ form is available in which the metal ions are bridged by a carbamylated Lys-amino group with a metal-metal distance of 3.8 A [ 18]. Substrate hydrolysis follows a SN2 type reaction and nucleophilic attack of M-OH is likely, but mechanistic details are not yet clear. 

Fig. 32 The structure of the active site of the phosphotriesterase with the competitive inhibitor bound as determined by X-ray crystallography.

An array of 16 enantiomeric pairs of chiral phosphate, phosphonate and phosphinate esters was used to establish the breadth of stereoselective discrimination inherent within the bacterial phosphotriesterase and 15 mutant enzymes. The results demonstrate that the catal5hic properties of the wide-type of phosphotriesterases can be exploited for the kinetic resolution of a wide-range of compounds and that the active site of this enzyme is remarkably amenable to structural perturbations via amino acid substitution. The first use of dialkyl phosphates as stereodirecting... 

The proposed mechanism for phosphotriester cleavage by PONl and DFPase is shown in Fig. 3.16. According to Bigley and Raushel, these enzymes present two calcium ions located in the core of the structure, however, only one participate in the active site. This is the only mechanism of phosphotriesterases where a single active site metal ion is employed in the catalysis. The pH profiles for both enzymes indi-... 

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