Characterization of BctA, a mating apparatus protein required for transfer of the Bacteroides fragilis conjugal element BTF-37

Abstract

We have previously described the identification of BTF-37, an autonomously transferable chromosomal element isolated from Bacteroides fragilis clinical isolate LV23. In this study, we determined that BTF-37 harbors a 16kb conjugal transfer-encoding region that contains an almost identical copy of a previously identified Bacteroides sp. conjugation-specific gene bctA. BctA has been shown to be required for conjugation in other Bacteroides sp. strains, but no information is available regarding its function. We now report strain distribution and gene expression profiles of bctA. The bctA gene was present in conjugative transposon-harboring B. fragilis strains, but not on a non-transferable B. fragilis plasmid. We also showed that recombinant BctA predominantly localized to the bacterial membrane, and that its N-terminal 32 amino acids were cleaved in an Escherichia coli protein expression system, indicating the presence of a signal sequence. Expression of bctA consistently increased ~3-fold upon pre-exposure of conjugating B. fragilis LV23 to subinhibitory concentrations of tetracycline. Maximum expression occurred 60min post-tetracycline induction, which also coincided with the time at which highest conjugation frequencies were seen for strain LV23. Based on localization, signal sequence and tetracycline inducibility, our results indicate that BctA is indeed an important member of the Bacteroides conjugal apparatus, since its gene is regulated by conditions that specifically control conjugation.