Atrazine Hydrolysis by a Bacterial Enzyme

Lawrence P. Wackett, Michael J. Sadowsky, Mervyn De Souza, Raphi T. Mandelbaum

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Atrazine, 2-chloro-4-(ethylamino)-6-(isopropylamino)-1,3,5-triazine, is metabolized relatively slowly in natural soils and waters by resident microorganisms. Recently, several atrazine-degrading bacterial pure cultures were isolated and the molecular basis of bacterial atrazine metabolism is now beginning to be revealed. Pseudomonas sp. strain ADP was isolated from a herbicide spill site for its ability to use atrazine as the sole source of nitrogen for growth. Atrazine metabolism also liberated the triazine ring carbon atoms as carbon dioxide. Hydroxyatrazine was detected transiently in the growth medium during the course of atrazine metabolism. Previously, hydroxyatrazine was proposed to be derived solely from abiotic hydrolysis catalyzed by soil organic matter and clays. The gene encoding the enzymatic hydrolysis of atrazine by Pseudomonas sp. ADP was cloned and expressed in Escherichia coll. Cell-free atrazine hydrolysis activity in the recombinant E. coli strain was determined by high pressure liquid chromatography. The enzyme, atrazine chlorohydrolase, was purified to homogeneity using ammonium sulfate precipitation and ion exchange chromatography. The purified chlorohydrolase showed a single band on denaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis corresponding to a subunit molecular weight of 60,000. Gene sequencing data yielded a molecular weight of 52,421. Gel filtration chromatography indicated a holoenzyme molecular weight of 240,000 consistent with an α4 or α5 subunit stoichometry. In [18O]-H2O, atrazine chlorohydrolase yielded [18O]-hydroxyatrazine quantitatively. In control experiments incubated and analyzed under the same conditions, [18O] from H2O did not exchange into hydroxyatrazine. These data are consistent with enzymatic hydrolysis of atrazine. Other bacteria were also demonstrated to catalyze atrazine hydrolysis, suggesting this biologically-mediated reaction is widespread in soil and water.

Original languageEnglish (US)
Pages (from-to)82-87
Number of pages6
JournalACS Symposium Series
Volume683
StatePublished - Dec 1 1998

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