Molecular cloning and characterization of the copper/zinc and manganese superoxide dismutase genes from the human parasiteClonorchis sinensis

Abstract

A copper/zinc superoxide dismutase (Cu/ZnSOD) gene and a manganese superoxide dismutase (MnSOD) gene of the human parasiteClonorchis sinensishave been cloned and their gene products functionally characterized. GenesCu/ZnSODandMnSODencode proteins of 16 kDa and 25·4 kDa, respectively. The deduced amino acid sequences of the two genes contained highly conserved residues required for activity and secondary structure formation of Cu/ZnSOD and MnSOD, respectively, and show up to 73·7% and 75·4% identities with their counterparts in other animals. The genomic DNA sequence analysis of Cu/ZnSOD gene revealed this as an intronless gene. Inhibitor studies with purified recombinant Cu/ZnSOD and MnSOD, both of which were functionally expressed inEscherichia coli, confirmed that they are copper/zinc and manganese-containing SOD, respectively. Immunoblots showed that bothC. sinensisCu/ZnSOD and MnSOD should be antigenic for humans, and both, especially theC. sinensisMnSOD, exhibit extensive cross-reactions with sera of patients infected by other trematodes or cestodes. RT-PCR and SOD activity staining of parasite lysates indicate that there are no significant differences in mRNA level or SOD activity for both species of SOD, indicating cytosolic Cu/ZnSOD and MnSOD might play a comparatively important role in theC. sinensisantioxidant system.