Molecular cloning, characterization and expression analysis of cytoplasmic Cu/Zn-superoxid dismutase (SOD) from pearl oyster Pinctada fucata

Abstract

a b s t r a c tBecause of its capacity to rapidly convert superoxide to hydrogen peroxide, superoxide dismutase (SOD) is crucial in both intracellular signalling and regulation of oxidative stress. In this paper we report the cloning of a Cu/Zn SOD (designated as pfSOD) from the pearl oyster (Pinctada fucata) using rapid amplification of cDNA ends (RACE) PCR. The full-length cDNA of this Cu/Zn SOD contains an open reading frame (ORF) of 471 bp coding for 156 amino acids. No signal peptide was identified at the N-terminal amino acid sequence of Cu/Zn SOD indicating that this pfSOD encodes a cytoplasmic Cu/Zn SOD. This is supported by the presence of conserved amino acids required for binding copper and zinc. Semi-quantitative analysis in adult tissues showed that the pfSOD mRNA was abundantly expressed in haemocytes and gill and scarcely expressed in other tissues tested. After challenge with lipopolysaccharide (LPS), expression of pfSOD mRNA in haemocytes was increased, reaching the highest level at 8 h, then dropping to basal levels at 36 h. These results suggest that Cu/Zn SOD might be used as a bioindicator of the aquatic environmental pollution and cellular stress in pearl oyster.

Identification and analysis of icCu/Zn-SOD, Mn-SOD and ecCu/Zn-SOD in superoxide dismutase multigene family of Pseudosciaena crocea

“…Some signature regions were also found, including two CueZn signature [20] in LycSOD1, one Mn-SOD signature region in LycSOD2 and one CueZn signature region in LycSOD3, and all of them were striking conserved in SOD family. Meanwhile, several relatively conserved aa residues were also revealed in the LycSOD1 and LycSOD3, for instance, two cysteine residues involved in forming one intracellular disulfide bond [11], and four conversed copper and zinc binding sites, which highly influenced on stabilizing the quaternary structure and related to pathogenic mutants [20,21]. The copper binding aa sites mainly made a bridge linking between copper and zinc ions, while zinc binding sites formed the similar structure to Zn finger domains [17,22].…”

Sources of marine superoxide dismutases: Characteristics and applications

“…Semi-quantitative analysis in adult tissues showed that the pfSOD mRNA was abundantly expressed in haemocytes and gill and scarcely expressed in other tissues tested. After challenge with lipopolysaccharide (LPS), expression of pfSOD mRNA in haemocytes was increased, reaching the highest level at 8 h, then dropping to basal levels at 36 h. These results suggest that Cu/Zn-SOD might be used as a bioindicator of the aquatic environmental pollution and cellular stress in pearl oyster [5].…”

“…The pfSOD mRNA was abundantly expressed in haemocytes and gill After challenge with lipopolysaccharide (LPS), expression of pfSOD mRNA in haemocytes was increased [5] Alvinella pompejana ApMn-SOD1 a high thermostable enzyme. Their half-lives were similarly long at 65 • C (<110 min) but exhibited a twofold difference at 80 • C (20.8 vs 9.8 min) [63] Apostichopus japonicus Increase levels of Sod mRNA by thermal and osmotic stresses [64] electrophoresis in reducing conditions; the subunit molecular mass was 18.8 kDa value that is remarkably higher than the one reported for other Cu/Zn-SODs.…”

“…Fe-SOD is found in prokaryotes (sodB), protozoans, and chloroplasts of algae and in plants two families of Fe-SODs have been found: the chloroplastic localized and the plastidial localized [3,5,14]. Fe and Mn-SODs are typically homodimers or homotetramers that probably evolved from a common ancestor.…”

“…Mn-SOD occurs in prokaryotes (sodA) and mitochondria of the eukaryotes (sod2), has the ability to catalyze the toxic superoxide anion into molecular oxygen and hydrogen peroxide [6,15]. Additionally Ni-SOD has recently been purified from several aerobic soil bacteria of Streptomyces [5,16]. Three common isoforms could be distinguished by their differential sensitivities to different chemicals.…”