Characterization of iron superoxide dismutase cDNAs from plants obtained by genetic complementation in Escherichia coli

Abstract

The inability of superoxide dismutase (SOD; superoxide:superoxide oxidoreductase, EC 1.15.1.1)-deficient mutents of Escherichia coli to grow aerobically on minimal medium can be restored by functional complementation with a heterotogous SOD-encoding sequence. Based upon this property, a phenotypic selection system has been developed for the isoloation of clones containing eukaryotic SOD cDNAs. cDNA expression libraries from both Nicotiana plumbaginifolia and Arabidopsis thaliana were transformed into a SOD-deficient E. coli strain by electroporation, and clones containing functional SODs were selected by growth on minimal medium. Analysis of these clones revealed the identity of cDNAs encoding the iron Im of superoxide dismutase (FeSOD) - the first SODs of this type to be cloned from eukaryotes. The presence of this enzyme in these two divergent plant species challenges previous ideas that FeSOD is found in only a few plant families. In addition, these results show the potential for shotgun cloning of eukarytics genes by complementation of bacterial mutants, particularly when it is combined with a highly efficient transformation method, such as electroporation. (.