Characterization of S-triazine herbicide metabolism by a Nocardioides sp isolated from agricultural soils

Atrazine, a herbicide widely used in corn production, is a frequently detec ted groundwater contaminant. Nine gram-positive bacterial strains able to u se this herbicide as a sole source of nitrogen were isolated from four farm s in central Canada. The strains were divided into two groups based on repe titive extragenic palindromic (rep)-PCR genomic fingerprinting with ERIC an d BOXAIR primers, Based on 16S ribosomal DNA sequence analysis, both groups were identified as Nocardioides sp. strains. None of the isolates minerali zed [ring-U-C-14] atrazine. There was no hybridization to genomic DNA from these strains using atzABC cloned from Pseudomonas sp, strain ADP or trzA c loned from Rhodococcus corallinus. S-Triazine degradation was studied in de tail in Nocardioides sp, strain C190. Oxygen was not required for atrazine degradation by,whole cells or cell extracts. Based on high-pressure liquid chromatography and mass spectrometric analyses of products formed from atra zine in incubations of whole cells with (H2O)-O-18, sequential hydrolytic r eactions converted atrazine to hydroxyatrazine and then to the end product N-ethylammelide. Isopropylamine, the putative product of the second hydroly tic reaction, supported growth as the sole carbon and nitrogen source. The triazine hydrolase from strain C190 was isolated and purified and found to have a K-m for atrazine of 25 mu M and a V-max of 31 mu mol/min/mg of prote in. The subunit molecular mass of the protein was 52 kDa. Atrazine hydrolys is was not inhibited by 500 mu M EDTA but mas inhibited by 100 mu M Mg, Cu, Co, or Zn, Whole cells and purified triazine hydrolase converted a range o f chlorine or methylthio-substituted herbicides to the corresponding hydrox y derivatives. In summary, an atrazine-metabolizing Nocardioides sp, widely distributed in agricultural soils degrades a range of s-triazine herbicide s by means of a novel s-triazine hydrolase.