MOLECULAR-CLONING AND SEQUENCING OF A NONHEME BROMOPEROXIDASE GENE FROM STREPTOMYCES-AUREOFACIENS ATCC-10762

A bromoperoxidase gene (bpoT), recently cloned form Streptomyces aureofaciens Tu24, was used as a probe in Southern blot hybridization of total DNA from S. aureofaciens ATCC 10762. A single SstI fragment of 5.4 kb was detected, which was cloned via an enriched gene library into Escherichia coli. The functional bromoperoxidase gene was located on a 2.1 kb BamHI-HindIII fragment by subcloning into S. lividans TK64, using the multicopy plasmid pIJ486. The enzyme was overproduced in S. lividans TK64 (up to 30 000 times compared to S. aureofaciens ATCC 10762) and showed the same electrophoretic and immunological properties as the bromoperoxidase BPOA2 purified from S. aureofaciens ATCC 10762. DNA sequence analysis revealed an open reading frame encoding a predicted polypeptide with the same M(r) and N-terminal amino acid sequence as the purified subunit of BPO-A2.