Expression of functional mammalian P450 2E1 in hairy root cultures.
P450 2E1 is an important mammalian liver enzyme known to metabolize a wide range of compounds including several common environmental pollutants. The medicinal plant, Atropa belladonna, was transformed with Agrobacterium rhizogenes containing a binary vector with rabbit P450 2E1 in either the sense or antisense orientation. The resulting "hairy roots" were isolated and grown in liquid medium. Production of P450 2E1 protein was verified in the roots containing the 2E1 gene in the sense orientation. Transgenic and control root cultures were dosed with the environmental pollutant, trichloroethylene (TCE), and were analyzed for the TCE metabolites, chloral and trichloroethanol. The root cultures expressing the mammalian P450 2E1 had increased levels of the metabolites compared to the levels in the control roots. This method represents a quick way to screen transformants for expression of foreign genes before regeneration of whole plants, and also as a possible source of foreign protein for purification.
Banerjee
Suchitra
S
Department of Biochemistry, Box 357350, University of Washington, Seattle, Washington 98195, USA. sldoty@u.washington.edu
Shang
Tanya Q
TQ
Wilson
Angela M
AM
Moore
Allison L
AL
Strand
Stuart E
SE
Gordon
Milton P
MP
Lafferty Doty
Sharon
S
D000818:Animals; D001532:Atropa belladonna / Q000166:cytology* / Q000235:genetics / Q000378:metabolism; D019392:Cytochrome P-450 CYP2E1 / Q000096:biosynthesis* / Q000235:genetics; D004785:Environmental Pollutants / Q000378:metabolism; D015870:Gene Expression; D018014:Gene Transfer Techniques; D009924:Organ Culture Techniques / Q000379:methods; D018517:Plant Roots / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D011817:Rabbits; D014170:Transformation, Genetic; D019076:Transgenes; D014241:Trichloroethylene / Q000378:metabolism
D004785:Environmental Pollutants; D014241:Trichloroethylene; D019392:Cytochrome P-450 CYP2E1