Disk Abalone, Haliotis discus discus, CuZn–Superoxide Dismutase cDNA and its Transcriptional Induction by Aroclor 1254

Abstract

CuZn–superoxide dismutase (CuZn–SOD) is a key antioxidant enzyme playing a first line protective role against reactive oxygen species (ROS) by converting superoxide () into H2O2. The CuZn–SOD gene was isolated from a whole abalone cDNA library and denoted as aCuZn–SOD. The full‐length cDNA of aCuZn–SOD was 1021 bp, which contained 465‐bp open reading frame (ORF) coding 154 amino acids. It contained highly conserved CuZn–SOD signature motif 1 (45GFHVHQFGDNT55) and motif 2 (139GNAGGRQACGVI150). Also, amino acid residues identified as Cu (His47, His49, His64, and His121) and Zn (His64, His72, His81, and ASP84) metal‐binding sites were completely conserved in the aCuZn–SOD. The reverse transcription polymerase chain reaction (RT‐PCR) results showed that aCuZn–SOD mRNA was expressed constitutively in gill, mantle, gonad, abductor muscle, digestive tract, and hemocytes in a tissue‐specific manner. The aCuZn–SOD mRNA was significantly up‐regulated (P < 0.05) in gill and digestive tract tissues after Aroclor 1254 induction compared to untreated and methanol‐injected abalone groups, suggesting that abalone has a potential use in assessing the impact of marine pollutants with the application of gene expression concept. In addition, purified recombinant aCuZn–SOD fusion protein was shown to reduceradical generated by xanthine oxidase assay, showing CuZn–SOD is a functionally active antioxidant enzyme in disk abalone.

Molecular characterization, purification, and antioxidant activity of recombinant superoxide dismutase from the Pacific abalone Haliotis discus hannai Ino

“…The digestive glands, gills, and blood are important immune organs; thus, the high expression of HdhCu/Zn-SOD in these tissues suggests that it may participate in the immune response against infection in mollusks. These results are somewhat similar to those reported by De Zoysa et al ( 2009 ), who showed, using semi-quantitative RT-PCR, that SOD of H. discus discus was highly expressed in the digestive tract and gill. Similarly, for Mg-MnSOD and Mg-Cu/Zn-SOD of Mytilus galloprovincialis and Mm-icCu/Zn-SOD of Meretrix meretrix , transcripts were primarily detected in the hepatopancreas (Gao et al 2012 ; Wang et al 2013 ).…”

“…E. coli ) to express recombinant SOD to determine its activity levels and investigate the role of SOD in marine mollusks. De Zosya et al ( 2009 ) described overexpression of a recombinant Cu/Zn-SOD fusion protein in E. coli K12(TB1) cells, reporting an activity level of 2461 U/mg. In this study, we used a P. pastoris GS115 eukaryotic expression system for in vitro recombinant expression of HdhCu/Zn-SOD from the Pacific abalone.…”