Cloning and expression of superoxide dismutase from Sarcoptes scabiei in Escherichia coli

Abstract: Sarcoptes scabiei is a disease-causing parasitic mite of humans and animals that is prevalent worldwide. The parasite lives in burrows in the epidermis of its host. These burrows are formed by a combination of mechanical destruction by the mite and secretion of various factors. The enzyme superoxide dismutase (SOD) catalyzes the dismutation of superoxide into oxygen and hydrogen peroxide. As such, it is an important antioxidant defense in nearly all cells exposed to oxygen. In this project, the enzyme was expressed in transformed Escherichia coli cells. The SOD cDNA from S. scabiei was ligated into two different expression vectors: pPU16 and pET-14b. The S. scabiei SOD open reading frame reported here is 696 nucleotides long and yields a protein with a molecular weight of  69.5 kDa. Only one of the constructs was successfully created, using pPU16. The construct was designated pPU110 and has a sequence coding for a hexahistidine tag downstream of the SOD cDNA and has a sequence coding for the maltose binding protein (MBP) upstream. The expression plasmid pPU110 was verified by DNA-sequencing and the tested in different expression experiments. Analysis using SDS-PAGE showed that recombinant fusion SOD could be readily expressed in E.coli.