CHARACTERIZATION OF P-S BOND HYDROLYSIS IN ORGANOPHOSPHOROTHIOATE PESTICIDES BY ORGANOPHOSPHORUS HYDROLASE

The extensive use of organophosphorothioate insecticides in agriculture has resulted in the risk of environmental contamination with a variety of broadly based neurotoxins that inhibit the acetylcholinesterases of many different animal species. Organophosphorus hydrolase (OPH, EC 3.1.8.1) is a broad-spectrum phosphotriesterase that is capable of detoxifying a variety of organophosphorus neurotoxins by hydrolyzing various phosphorus-ester bonds (P-O, P-F, P-CN, and PS) between the phosphorus center and an electrophilic leaving group. OPH is capable of hydrolyzing the P-X bond of various organophosphorus compounds at quite different catalytic rates: P-O bonds (k(cat) = 67-5000 s(-1)), P-F bonds (k(cat) = 0.01-500 s(-1)), and P-S bonds s (k(cat) = 0.0067 to 167 s(-1)). P-S bond cleavage was readily demonstrated and characterized in these studies by quantifying the released free thiol groups using 5,5'-dithio-bis-2-nitrobenzoic acid or by monitoring an upheld shift of approximately 31 ppm by P-31 NMR. A decrease in the toxicity of hydrolyzed products was demonstrated by directly quantifying the loss of inhibition of acetylcholinesterase activity. Phosphorothiolate esters, such as demeton-S, provided noncompetitive inhibition for paraoxon (a P-O triester) hydrolysis, suggesting that the binding of these two different classes of substrates was not identical. (C) 1995 Academic Press, Inc.