Cloning and determination of the nucleotide sequence of the Mn-containing superoxide dismutase gene from Halobacterium halobium.

A group of synthetic 17-mer oligodeoxynucleotides (oligos) was constructed to correspond to a sequence of amino acids situated near the N terminus of the manganese-containing superoxide dismutase (Mn-SOD) purified from the halophilic bacterium, Halobacterium halobium. A cosmid library of a Sau3AI partial digest of halobium DNA, cloned into the BamHI site of pHC79, was probed with the radiolabeled oligos. Cosmid DNA was purified from the clone that showed hybridization at the highest stringency. A 1.8-kb PstI fragment of this DNA which hybridized the probes was subcloned into bacteriophage M13 and transfected into Escherichia coli JM101. The entire insert containing a 600-bp sequence coding for Mn-SOD and its 5'- and 3'-flanking regions was sequenced. The derived amino acid sequence of the structural gene showed a similarity to other manganese and iron-containing SODs in normally conserved regions.