Overexpression of methyl parathion hydrolase and its application in detoxification of organophosphates.
The coding region of mpd gene corresponding to mature methyl parathion hydrolase (MPH) was heterologously overexpressed in Escherichia coli BL21 (DE3) by using pET expression system. The lactose-induced expression yield of MPH is increased 2-fold compared with IPTG as inducer. Furthermore, it was found that specific activity of MPH increased 48% by reducing the induction temperature to 22 degrees C. The addition of 25 mM lactose at 22 degrees C, the MPH activity of fermentation broth had a specific activity of 1.4 x 10(4) U/mg protein. Plasmid was no significant decrease in the modified medium. The optimal pH and temperature of MPH were 8.0 and 30 degrees C, respectively. Over a period of 5 months, the dried cells showed no significant decrease in the activity of the detoxifying enzymes. The crude enzymes in 50 mM citrate-phosphate buffer (pH 8.0) were able to degrade about 98% of the organophosphate pesticides sprayed on cabbage. The detoxification efficiency was superior to that of the treatments of water, detergent, and a commercially available enzyme product. Additionally, the products of pesticide hydrolysis generated by treatment with the enzyme extract were determined to be virtually nontoxic.
Yang
Jijian
J
State Key Laboratory of Integrated Management of Pest Insects & Rodents, Institute of Zoology, Chinese Academy of Sciences, Da Tun Lu, Beijing 100101, P.R. China.
Yang
Chao
C
Jiang
Hong
H
Qiao
Chuanling
C
D001483:Base Sequence; D001673:Biodegradation, Environmental; D001937:Brassica; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005285:Fermentation; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism*; D008658:Inactivation, Metabolic; D008743:Methyl Parathion / Q000378:metabolism* / Q000633:toxicity; D009943:Organophosphorus Compounds / Q000378:metabolism* / Q000633:toxicity; D010575:Pesticides / Q000378:metabolism / Q000633:toxicity; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013696:Temperature
D017931:DNA Primers; D009943:Organophosphorus Compounds; D010575:Pesticides; D011994:Recombinant Proteins; D008743:Methyl Parathion; D006867:Hydrolases