Molecular cloning, expression, and characterization of the Cu,Zn superoxide dismutase (SOD1) gene from the entomopathogenic fungus Cordyceps militaris

Abstract

We describe the molecular characterization of the Cu,Zn superoxide dismutase (SOD1) gene of Cordyceps militaris, which is one of the entomopathogenic fungi called a vegetable wasp and plant worm. The SOD1 gene of C. militaris spans 922 bp and consisted of three introns and four exons coding for 154 amino acid residues. The deduced amino acid sequence of the C. militaris SOD1 cDNA showed 88% identity to Claviceps purpurea SOD1, 82% to Neurospora crassa SOD1, and 75-64% to SOD1 sequences from other fungi. The C. militaris SOD1 possesses the typical metal binding ligands of six histidines and one aspartic acid common to fungal SOD1s. The cDNA encoding C. militaris SOD1 was expressed as a 17-kDa polypeptide in the baculovirus-infected insect Sf9 cells. The enzyme activity of the purified recombinant C. militaris SOD1 was approximately 568 U per mg(-1) . Southern blot analysis of the genomic DNA suggested the C. militaris SOD1 was a single gene. Northern and Western blot analysis and enzyme activity assays indicated SOD1 was expressed constitutively. This is the first report of an SOD1 gene from any entomopathogenic fungus.