A Pseudomonas isolate (GLC11) capable of growth in the presence of up to 125 mM glyphosate [N-phosphonomethyl glycine (PMG)] has been isolated. Unlike the previously isolated Pseudomonas PG2982 and other bacterial strains, isolate GLC11 grows equally well in commercial formulation and analytical grade PMG. Utilisation of PMG as a phosphorus source is repressed by inorganic phosphate (Pi) in both isolates. Enzymatic activity responsible for carbon-phosphorus bond cleavage (C-P lyase) was detected in cell-free extracts of both isolates and was partially purified. Resolution on DE-52 anion exchange chromatography yielded a single peak of C-P lyase activity. The molecular mass of C-P lyase as analysed by gel permeation chromatography is approximately 200 kDa. The enzyme activity was localised in the periplasmic space of bacteria. The specific activity of C-P lyase was different for different phosphonates when used as substrates.