Phosphotriesterase mechanism

Binding of a designed substrate analogue to diiso-propyl fluorophosphatase implications for the phosphotriesterase mechanism. J. Am. Chem. Soc. 128 12750-7. 

The mechanism resembles that proposed for a phosphotriesterase (Fig. 12-24). The triesterase catalyzes detoxification of organophosphorus toxins such as parathion (Box 12-E) and seems to have evolved rapidly from a homologous protein of unknown function.721 The phosphotriesterase contains two Zn2+ ions in a dimetal center. An unusual structural feature is a carbamate group, formed from Lys 169 and C02, which provides a bridging ligand for the metal pair.721-725 A carbamylated lysine also functions in ribulose bisphos-phate carboxylase (Fig. 13-11). 

The methanolyses of a series of <9,0-diethyl 0-aryl phosphates (101 X = O) and 0,0-diethyl 5-aryl phosphorothioates (101 X = S) promoted by methoxide and two metal ion systems, (La3+)2(MeO )2 and a complex of Zn2+ (MeO-) with 1,5,9-triazacyclododecane (79 M = Zn), has been studied in methanol at 25 °C. The kinetic data for the metal-catalysed reactions were analysed in terms of a common mechanism where there is extensive cleavage of the P-XAr bond in the transition state. The relevance of these findings to the mechanism of action of a phosphotriesterase enzyme present in a soil bacterium that hydrolyses paraoxon was discussed.100... 

Chen, S-L., Fang, W-H., Himo, F. (2007). Theoretical study of the phosphotriesterase reaction mechanism. J. Phys. Chem. B 111 1253-5. 

Wong, K.-Y., Gao, J. (2007). The reaction mechanism of para-oxon hydrolysis by phosphotriesterase Jrom combined QM/ MM simulations. Biochemistry 46 13352-69. 

Scheme 5 A model for the mechanism of the phosphotriesterase reaction. The data do not clearly distinguish between a concerted mechanism of phosphoryl transfer, or a phosphorane intermediate in the model above the reaction is shown as a concerted process. In either case, the P-OLg bond is largely broken in the transition state, requiring that breakdown of any putative intermediate is rate-limiting for less activated substrates. For more labile substrates, such as paraoxon, a step involving proton transfer that occurs after phosphoryl transfer is rate-limiting.

Selected Biological Aspects. - Phosphonate and phosphinic acids and their derivatives have been widely used in haptens. Examples additional to those already discussed in other sections include the phosphonic acid anion 363 which was an alternative to the preferred but synthetically inaccessible structure 364. The mechanism-based probe 365 has been synthesised and shown to modify a bacterial phosphotriesterase. This strategy for generating a probe is general and should allow the isolation of a host of unique catalysts. ... 

Arvlester Hydrolase and Parathion Hydrolase. Arylester hydrolase in mammalian serum (42.431 catalyzes very efficient hydrolysis of the organophosphorus oxons such as paraoxon (Table III). This enzyme does not hydrolyze the parent phosphorothioate insecticides such as methyl parathion, but only the oxon metabolites such as methyl paraoxon (441. This enzyme is has been referred to as phosphotriesterase however, a triester is not required as seen in the rapid hydrolysis of 10 organophosphinates by rabbit serum arylester hydrolase (44). This mechanism appears only rarely in birds... 

The enzyme phosphotriesterase hydrolyzes many different organophosphorus triesters, including several acetylcholinesterase inhibitors. The chemical mechanism involves an activated water molecule that directly attacks the phosphorus center with a resultant inversion of configuration [708]. Thus the overall reaction mechanism does not involve the formation of a phosphorylated-enzyme intermediate, but it does involve a Schiff base-type interaction between carbon dioxide and a lysine residue giving a carbamate group in the active site. 

The phosphotriesterase-catalyzed reaction involves an SN2-type mechanism, with inversion of configuration at the phosphorus atom [708] activated water is presumed to attack the electrophilic phosphorus of the substrate, shown as an attack on diethyl 4-nitrophenyl phosphate (a non-biological substrate) in Figure 26(b). Both zinc ions work to activate hydrolytic water and polarize the P-0 bond [77i, 114]. 

Figure 26. (a) The active site of phosphotriesterase. (b) the proposed reaction mechanism of the enzyme. 

Wong, K.Y, Gao, J. The reaction mechanism of paraoxon hydrolysis by phosphotriesterase from combined QM/MM simulations. Biochemistry 2007,46(46), 13352-69. 

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-... 

Bigley, A.N., Raushel, F.M., 2013. Catalytic mechanisms for phosphotriesterases. Biochim. Biophys. Acta 1834 (1), 443-453. 

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