Key Laboratory of Monitoring and Management of Plant Diseases and Insects, Ministry of Agriculture, Nanjing Agricutural University,Nanjing 210095, China 2. Department of Biological Chemistry, Rothamsted Research, Harpenden, Herts, AL5 2JQ, United Kingdom
Using the Bac-to-Bac baculovirus expression system, the acetylcholinesterase 1 gene (ace1) of Aphis gossypii was successfully expressed in Sf9 cells. The enzyme was secreted into the cell culture medium and tested. The affinity (Km) of the expressed AChE1 for the substrate analog ATChI was 144.5±28.6 μM. The sensitivity index (indicated by Ki value) of expressed AChE1 to 2 carbamate insecticides (pirimicarb and triazamate) and 2 organophosphate insecticides (omethoate and demeton-S-methyl) were 513, 1 883, 5 and 23 (mM·min) -1, respectively, which is in agreement with previously reported values obtained with the natural enzyme. The expressed enzyme showed highest activity when harvested at 2-3 d after infection of the insect cells. The expressed AChE1 was very stable without obvious loss in enzyme activity after 90 d when stored at -20 ℃, although 70% of the enzyme activity was lost after 7 d when stored at room temperature.