Inhibitor Report for: Sulfotep

The mechanism by which the binuclear metallophosphotriesterases (PTEs, E.C. 3.1.8.1) catalyse substrate hydrolysis has been extensively studied. The mu-hydroxo bridge between the metal ions has been proposed to be the initiating nucleophile in the hydrolytic reaction. In contrast, analysis of some biomimetic systems has indicated that mu-hydroxo bridges are often not themselves nucleophiles, but act as general bases for freely exchangeable nucleophilic water molecules. Herein, we present crystallographic analyses of a bacterial PTE from Agrobacterium radiobacter, OpdA, capturing the enzyme-substrate complex during hydrolysis. This model of the Michaelis complex suggests the alignment of the substrate will favour attack from a solvent molecule terminally coordinated to the alpha-metal ion. The bridging of both metal ions by the product, without disruption of the mu-hydroxo bridge, is also consistent with nucleophilic attack occurring from the terminal position. When phosphodiesters are soaked into crystals of OpdA, they coordinate bidentately to the beta-metal ion, displacing the mu-hydroxo bridge. Thus, alternative product-binding modes exist for the PTEs, and it is the bridging mode that appears to result from phosphotriester hydrolysis. Kinetic analysis of the PTE and promiscuous phosphodiesterase activities confirms that the presence of a mu-hydroxo bridge during phosphotriester hydrolysis is correlated with a lower pK(a) for the nucleophile, consistent with a general base function during catalysis.

A novel method for immobilization of acetylcholinesterase (AChE) by binding covalently to a cross-linked chitosan-multiwall carbon nanotube (MWNT) composite is described. In addition a sensitive, fast, cheap and automatizable flow injection detection of an organophosphorous insecticide was developed. The MWNTs were homogeneously distributed in the chitosan membrane which showed a homogeneous porous structure. The immobilized AChE could catalyze the hydrolysis of acetylthiocholine with a K(M)app value of 177 microM to form thiocholine, which was then oxidized to produce detectable signal in a linear range of 1.0-500 microM and fast response. MWNTs could catalyze the electrooxidation of thiocholine, thus increasing detection sensitivity. Based on the inhibition of an organophosphorous insecticide on the enzymatic activity of AChE, using Sulfotep as a model compound, the conditions for the flow-injection detection of the insecticide were optimized. Both biocompatibility of chitosan and inherent conductive properties of MWNTs favored the detection of the insecticide from 1.5 to 80 microM along with good stability and reproducibility. 95 % reactivation from inhibited AChE could be regenerated by using 2-pyridinealdoxime methiodide within 15 min for 15 times. The detection of Sulfotep samples exhibited satisfactory results. The proposed flow-injection analysis device can be applied to automated determination and characterization of enzyme inhibitors.

Acid hydrolysis is a means of disposal of diazinon, O,O-diethyl-O-(2-isopropyl-4-methyl-6-pyrimidinyl)-phosphorothioate. Procedures were developed for the hydrolysis of three military standard diazinon formulations (47.5% emulsifiable concentrate, 2% dust and 0.5% oil solution). The kinetics of hydrolysis at four temperatures were obtained for each formulation in order to predict the half-life of diazinon between 5 degrees and 50 degrees C. The toxicities of the reaction mixtures were evaluated by aquatic bioassay with bluegill sunfish (Lepomis macrochirus). When the hydrolysis of a formulation had reduced the diazinon concentration by greater than 99.9%, there was less than 50% reduction in the toxicity of the reaction mixture to bluegill sunfish. Analysis of reaction mixtures showed that the toxic component was O,O,O,O-tetraethyldithiopyrophosphate (Sulfotepp), which was present in all diazinon formulations.

The mechanism by which the binuclear metallophosphotriesterases (PTEs, E.C. 3.1.8.1) catalyse substrate hydrolysis has been extensively studied. The mu-hydroxo bridge between the metal ions has been proposed to be the initiating nucleophile in the hydrolytic reaction. In contrast, analysis of some biomimetic systems has indicated that mu-hydroxo bridges are often not themselves nucleophiles, but act as general bases for freely exchangeable nucleophilic water molecules. Herein, we present crystallographic analyses of a bacterial PTE from Agrobacterium radiobacter, OpdA, capturing the enzyme-substrate complex during hydrolysis. This model of the Michaelis complex suggests the alignment of the substrate will favour attack from a solvent molecule terminally coordinated to the alpha-metal ion. The bridging of both metal ions by the product, without disruption of the mu-hydroxo bridge, is also consistent with nucleophilic attack occurring from the terminal position. When phosphodiesters are soaked into crystals of OpdA, they coordinate bidentately to the beta-metal ion, displacing the mu-hydroxo bridge. Thus, alternative product-binding modes exist for the PTEs, and it is the bridging mode that appears to result from phosphotriester hydrolysis. Kinetic analysis of the PTE and promiscuous phosphodiesterase activities confirms that the presence of a mu-hydroxo bridge during phosphotriester hydrolysis is correlated with a lower pK(a) for the nucleophile, consistent with a general base function during catalysis.

A novel method for immobilization of acetylcholinesterase (AChE) by binding covalently to a cross-linked chitosan-multiwall carbon nanotube (MWNT) composite is described. In addition a sensitive, fast, cheap and automatizable flow injection detection of an organophosphorous insecticide was developed. The MWNTs were homogeneously distributed in the chitosan membrane which showed a homogeneous porous structure. The immobilized AChE could catalyze the hydrolysis of acetylthiocholine with a K(M)app value of 177 microM to form thiocholine, which was then oxidized to produce detectable signal in a linear range of 1.0-500 microM and fast response. MWNTs could catalyze the electrooxidation of thiocholine, thus increasing detection sensitivity. Based on the inhibition of an organophosphorous insecticide on the enzymatic activity of AChE, using Sulfotep as a model compound, the conditions for the flow-injection detection of the insecticide were optimized. Both biocompatibility of chitosan and inherent conductive properties of MWNTs favored the detection of the insecticide from 1.5 to 80 microM along with good stability and reproducibility. 95 % reactivation from inhibited AChE could be regenerated by using 2-pyridinealdoxime methiodide within 15 min for 15 times. The detection of Sulfotep samples exhibited satisfactory results. The proposed flow-injection analysis device can be applied to automated determination and characterization of enzyme inhibitors.

Proctolin-induced, dose-dependent (10-8-2 10-6 M) contraction of the isolated foregut of Schistocerca gregaria was antagonised non-competitively by sulfotep (2 10-6-10-5 M). A higher dose of sulfotep (5 10-5 M) caused restoration of the proctolin dose-response curve to its control value. Neostigmine (10-5 M) caused non-competitive inhibition of proctolininduced tissue contraction. Increasing the dose of neostigmine to 10-4 M restored the proctolin response to control values. Sulfotep (10-5 M) and neostigmine (10-4 M) caused inhibition of acetylcholinesterase (AChE) activity in tissue homogenates obtained from guts pretreated with either drug for 20 min. The stimulatory effect of sulfotep (5 10-5 M) on proctolin-induced gut contraction was abolished by pretreatment of tissues with atropine (10-6 M). Under these conditions, 5 10-5 M sulfotep caused further antagonism of the action of proctolin. The results suggest that sulfotep is a proctolin receptor antagonist in the locust foregut. However, higher concentrations inhibit tissue AChE activity, thereby allowing endogenous acetylcholine to activate muscarinic receptors. This leads to enhanced tissue contractility which masks the antagonistic effect of sulfotep on proctolin-induced contraction.

Acid hydrolysis is a means of disposal of diazinon, O,O-diethyl-O-(2-isopropyl-4-methyl-6-pyrimidinyl)-phosphorothioate. Procedures were developed for the hydrolysis of three military standard diazinon formulations (47.5% emulsifiable concentrate, 2% dust and 0.5% oil solution). The kinetics of hydrolysis at four temperatures were obtained for each formulation in order to predict the half-life of diazinon between 5 degrees and 50 degrees C. The toxicities of the reaction mixtures were evaluated by aquatic bioassay with bluegill sunfish (Lepomis macrochirus). When the hydrolysis of a formulation had reduced the diazinon concentration by greater than 99.9%, there was less than 50% reduction in the toxicity of the reaction mixture to bluegill sunfish. Analysis of reaction mixtures showed that the toxic component was O,O,O,O-tetraethyldithiopyrophosphate (Sulfotepp), which was present in all diazinon formulations.