Characterization of the bleomycin resistance determinant encoded on the transposon Tn5

The transposon Tn5 carries a gene, ble, which confers resistance to bleomycin (Bm) and gives a survival advantage to its host cell. We found that the ble gene product, designated BLMT, is a binding protein with a strong affinity for Bm. BLMT quenched both the antibacterial and DNA-cleaving activities of Bm, when incubated with the antibiotic. An electron spin resonance spin-trapping analysis showed that BLMT inhibits the generation of Bm-induced hydroxyl radical, by trapping Bm but not the hydroxyl radical. Western blot analysis using an anti-BLMT monoclonal antibody revealed that BLMT is immunologically distinct from Bm-binding proteins from Streptomyces verticillus, Staphylococcus aureus and Streptoalloteichus hindustanus. Escherichia coli, transformed with a mutant ble having leucine instead of proline at N-terminal amino acid position 7, lost resistance to Bm, although the cell maintained the survival benefit. This suggests that the Bm resistance mediated by ble is independent of its ability to give a survival advantage to the host bacterium.