TipB, a novel cell wall hydrolase, is required for efficient conjugative transfer of pXO16 from Bacillus thuringiensis sv. israelensis.

pXO16, a large plasmid from Bacillus thuringiensis serovar israelensis, exhibits unique features. Not only is pXO16 able to transfer at high frequencies within few minutes, but it is also able to transfer among virtually all members of the Bacillus cereus group. Among the proteins encoded in the tip transfer locus of pXO16, TipB displays an atypical organization compared to known conjugative cell wall hydrolases with the large central soluble lytic transglycosylase (SLT) domain missing from the protein. We constructed a tipB deletion mutant which led to significant reduction in transfer efficiencies in both liquid and filter mating. The initial transfer frequencies could be restored when complementing tipB in trans thus showing the TipB implication in pXO16 conjugative transfer. Additionally, truncated versions of TipB were expressed in Escherichia coli to assess the protein lytic activity. When applied exogenously, TipB-2TM, in which the two N-terminal TM domains were removed, yielded a decrease of ca. 40% in optical density of B. thuringiensis sv. israelensis, a lytic activity that could partially be explained by the C-terminal CHAP-like domain. These results confirm TipB conjugative hydrolase function and provide new insights into pXO16 unique conjugative apparatus.