Expression of human cytochromes P450 1A1 and P450 1A2 as fused enzymes with yeast NADPH-cytochrome P450 oxidoreductase in transgenic tobacco plants.
Among 11 isoforms of the human cytochrome P450 enzymes metabolizing xenobiotics, CYP 1A1 and CYP 1A2 were major P450 species in the metabolism of the herbicides chlortoluron and atrazine in a yeast expression system. CYP1A2 was more active in the metabolism of both herbicides than CYP1A1. The fused enzymes of CYP1A1 and CYP1A2 with yeast NADPH-cytochrome P450 oxidoreductase were functionally active in the microsomal fraction of the yeast Saccharomyces cerevisiae and showed increased specific activity towards 7-ethoxyresorufin as compared to CYP1A1 and CYP1A2 alone. Then, both fused enzymes were each expressed in the microsomes of tobacco (Nicotiana tabacum cv. Samsun NN) plants. The transgenic plants expressing the CYP1A2 fusion enzyme had higher resistance to the herbicide chlortoluron than the plants expressing the CYP1A1 fusion enzyme did. The transgenic plants expressing the CYP1A2 fused enzyme metabolized chlortoluron to a larger extent to its non-phytotoxic metabolites through N-demethylation and ring-methyl hydroxylation as compared to the plants expressing the CYP1A1 fused enzyme. Thus, the possibility of increasing the herbicide resistance in the transgenic plants by the selection of P450 species and the fusion with P450 reductase is discussed.
Shiota
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N
Department of Biological and Environmental Science, Faculty of Agriculture, Kobe University, Nada, Japan.
Kodama
S
S
Inui
H
H
Ohkawa
H
H
D001280:Atrazine / Q000378:metabolism; D001483:Base Sequence; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics / Q000378:metabolism*; D019388:Cytochrome P-450 CYP1A2 / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D006540:Herbicides / Q000378:metabolism; D006801:Humans; D009251:NADPH-Ferrihemoprotein Reductase / Q000235:genetics / Q000378:metabolism*; D010671:Phenylurea Compounds / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000201:enzymology*; D010947:Plants, Toxic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000378:metabolism; D014026:Nicotiana / Q000201:enzymology
D017931:DNA Primers; D006540:Herbicides; D010671:Phenylurea Compounds; D011993:Recombinant Fusion Proteins; D019363:Cytochrome P-450 CYP1A1; D019388:Cytochrome P-450 CYP1A2; D009251:NADPH-Ferrihemoprotein Reductase; D001280:Atrazine; C007033:chlortoluron