Biotransformation of atrazine in transgenic tobacco cell culture expressing human P450.
Plant cell cultures in which the appropriate P450 cDNA is introduced are expected to metabolise certain pesticides in large quantities. Two species of human P450 (CYP1A1 and CYP1A2) were introduced into tobacco cells (Nicotiana tabacum L) by Agrobacterium-mediated transformation. The transgenic plant cell cultures were selected by combination of kanamycin-resistance, 7-ethoxycoumarin O-de-ethylase activity, PCR and Western blot analysis. For metabolism studies, 14C-labelled atrazine was used as a model substance. The metabolites de-ethylatrazine and de-isopropylatrazine were found in the control culture as well as in the transgenic culture, whereas the non-phytotoxic metabolite de-ethyl-de-isopropylatrazine was found only in the transgenic cell cultures. The results showed that both foreign enzymes CYP1A1 and CYP1A2 catalyse N-dealkylation of atrazine. However, CYP1A2 exhibited a higher conversion rate than CYP1A1. In a time-course study the enzyme CYP1A2 catalysed predominantly N-de-ethylation followed by de-isopropylation. The extent of metabolism was considerably higher than in non-transformed cell cultures. The transgenic cell cultures can therefore be suitable tools for the production of large quantities of primary oxidised pesticide metabolites.
Bode
Maren
M
Department of Biology V (Ecology/Ecotoxicology/Ecochemistry), Aachen University, 52056 Aachen, Germany.
Stöbe
Petra
P
Thiede
Brigitte
B
Schuphan
Ingolf
I
Schmidt
Burkhard
B
D001280:Atrazine / Q000378:metabolism*; D001711:Biotransformation; D015153:Blotting, Western; D002250:Carbon Radioisotopes / Q000378:metabolism; D002478:Cells, Cultured; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics / Q000378:metabolism*; D019388:Cytochrome P-450 CYP1A2 / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D013997:Time Factors; D014026:Nicotiana / Q000201:enzymology* / Q000235:genetics
D002250:Carbon Radioisotopes; D019363:Cytochrome P-450 CYP1A1; D019388:Cytochrome P-450 CYP1A2; D001280:Atrazine