Purification and biochemical characterization of glutathione S-transferase from common pistachio psyllid, Agonoscena pistaciae Burckhardt and Lauterer (Hemiptera: Psyllidae)
نویسندگان
1 Department of Plant Protection, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
2 Department of Plant Protection, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran.
3 Department of Pesticides Researches, Iranian Research Institute of Plant Protection, Agricultural Re-search, Education and Extension Organization (AREEO), Tehran, Iran
4 Department of Crop Protection, Faculty of Agriculture,Vali-e-Asr University
5 Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
doi
10.22117/jesi.2019.124035.1277چکیده
Glutathione S-transferase (GST) was purified and isolated from adults of common pistachio psyllid, Agonoscena pistaciae Burckhardt and Lauterer, by using ammonium sulfate precipitation and affinity chromatography using Glutathione Sepharose 4B column. The purified enzyme appeared as a single band on SDS-PAGE with an apparent molecular weight of 28.4 kDa. GST was purified 39.27-fold with a yield of 12.37% and a specific activity of 30.08 U mg-1 protein from A. pistaciae. The optimum temperature and pH of the enzyme activity were 30 °C and 9.0, respectively. The Km and Kcatvalues for GSH (Glutathione) substrate were also determined to be 0.44 mM and 152.9 s−1 and for CDNB (1-chloro-2,4-dinitrobenzene) substrate to be 0.33 mM and 207.7 s−1, respectively. GST’s activity was completely inhibited by the addition of EDTA, ZnCl2, andSDS; however, partially inhibited by CaCl2, BaCl2, CoCl2, KCl, MnCl2, Urea, MgCl2 and Hg2Cl2. The in vitro inhibition studies indicated that all kinds of conventional insecticides (i.e. imidacloprid, acetamiprid, phosalone, and amitraz) possessed inhibitory effects on the activity of purified GST. Our study broadens the biochemical information on A. pistaciae’s GST and this information will help us to understand the mechanisms of insecticide resistance in this key pest.
