| Summary: | 博士 === 國防醫學院 === 生命科學研究所 === 90 === Cu/Zn-superoxide dismutase (Cu/Zn-SOD) and catalase cDNA were cloned from zebrafish using RACE-PCR technique; both of the cDNA coding regions were successfully expressed in Escherichia coli. The wild type Cu/Zn-SOD（ZSOD1） was purified by His-tagged technique and was found labile than plant Cu/Zn-SOD. In an attempt to enhance enzyme stability, a mutant enzyme (ZSODC7A, Cys-7 to Ala) was obtained by site-specific mutagenesis and showed higher thermo-stability than wild-type enzyme. In addition, we investigated the modulation of the activation of microglial cell by transfecting a Cu/Zn-SOD cDNA to microglia cells. Parental and transfected BV-2 cells were then subjected to LPS stimulation. The results showed that in Cu/Zn-SOD-transfected microglial cells, the expression and activity of Cu/Zn-SOD increased. On the other hand, these cells upon activation by LPS, had lower production of NO than the parental microglial cells. This finding suggests that the transfected Cu/Zn-SOD may provide a neuroprotective function via suppression of microglial activation. Finally, the recombinant ZSOD1 and ZSODC7A were used to protect fish from 100 ppm of paraquat induced oxidative injury by soaking fish larva with SOD enzyme. The results were significant.
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