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Ahmed M.H. Ali,Jae Man Lee,Mayumi Yoshida,Kosuke Sakashita,Jumpei Torii,Takahiro Kusakabe,Akinori Hirashima 한국응용곤충학회 2012 Journal of Asia-Pacific Entomology Vol.15 No.4
The insect nervous systemcontains biogenic amines such octopamine (OA),which is synthesized fromtyramine (TA) by catalysis of tyramine-β-hydroxylase (TβH). In this study, the Drosophila 70 kDa tyramine-β-hydroxylase (DmTβH) protein was purified after the recombinant nucleopolyhedrovirus isolated from Bombyx mori (BmNPV) containing the TβH gene was injected into the hemocoel of the fifth instar larvae from the d17B. mori strain. Western blot analysis revealed an immunoreactive band with a molecular mass of 70 kDa. The products formed by incubating the enzyme reaction mixture were separated and detected by reverse phase high-performance liquid chromatography. The optimum pH, temperature, and incubation time for the conversion of TA to OAwere 7.6, 25 °C, and 30 min, respectively. The inhibitory experiments using various concentrations of 1-(2-methoxy-5-methylphenyl) imidazole-2(3H)-thione (MMIT) showed that MMIT inhibited DmTβH dose-dependently and that this method can be applied for screening DmTβH inhibitors.