TcpM: a novel relaxase that mediates transfer of large conjugative plasmids from Clostridium perfringens

Abstract

Conjugative transfer of toxin and antibiotic resistance plasmids in Clostridium perfringens is mediated by the tcp conjugation locus. Surprisingly, neither a relaxase gene nor an origin of transfer (oriT) has been identified on these plasmids, which are typified by the 47 kb tetracycline resistance plasmid pCW3. The tcpM gene (previously called intP) encodes a potential tyrosine recombinase that was postulated to be an atypical relaxase. Mutagenesis and complementation studies showed that TcpM was required for wild-type transfer of pCW3 and that a tyrosine residue, Y259, was essential for TcpM activity, which was consistent with the need for a relaxase-mediated hydrophilic attack at the oriT site. Other catalytic residues conserved in tyrosine recombinases were not required for TcpM activity, suggesting that TcpM was not a site-specific recombinase. Mobilization studies led to the identification of the oriT site, which was located in the 391 bp intergenic region upstream of tcpM. The oriT site was localized to a 150 bp region, and gel mobility shift studies showed that TcpM could bind to this region. Based on these studies we postulate that conjugative transfer of pCW3 involves the atypical relaxase TcpM binding to and processing the oriT site to initiate plasmid transfer.

A novel T4SS‐mediated DNA transfer used by pXO16, a conjugative plasmid from Bacillus thuringiensis serovar israelensis

“…). However, no relaxase genes could be detected through homology searches in this region but described relaxases from the MPF FA group are often atypical (Rocco and Churchward, , Wisniewski et al ., ). Since gene proximity from the putative accessory protein tipO helicase gene is expected, further studies will first focus on the tipP gene as a potential relaxase and on its cognate oriT region.…”

A Glimpse into the World of Integrative and Mobilizable Elements in Streptococci Reveals an Unexpected Diversity and Novel Families of Mobilization Proteins

“…At least some of these could correspond to IMEs. This hypothesis is supported first by the discovery of a novel type of relaxase related to tyrosine recombinases that is encoded by the pCW3 conjugative plasmid from the firmicute C. perfringens (Wisniewski et al, 2016). Second, in proteobacteria, IMEs devoid of relaxase but carrying an oriT are found to be mobilizable (Daccord et al, 2010) and one such IME (IME MTn Sag1 ) have previously been described in S. agalactiae (Achard and Leclercq, 2007).…”

“…In conjugative elements, TcpA proteins are associated with non-canonical relaxases: (i) the relaxase of the conjugative pCW3 plasmid from C. perfringens that is related to tyrosine recombinases (Wisniewski et al, 2016) or (ii) the MobT relaxases from the ICEs belonging to ICE Bs1 /Tn 916 /ICE St3 superfamily that are related to RCR initiators. In this work, we identified many IMEs encoding a TcpA CP: all of them encode a non-canonical relaxase.…”

“…Furthermore, all the plasmids and ICEs that encode FA T4SSs and TcpAs CPs encode non-canonical relaxases. Thus, the pCW3 conjugative plasmid from the Firmicute Clostridium perfringens , which encodes a FA T4SS and a TcpA CP, was recently shown to encode a novel type of relaxase related to tyrosine recombinase (Wisniewski et al, 2016). In the same way, all ICEs, which encode a FA T4SS and a TcpA CP, encode a relaxase belonging to the non-canonical MobT superfamily.…”

Regulation of conjugative transfer of plasmids and integrative conjugative elements

“…In another Gram-positive bacterium, Clostridium perfringens, toxin and antibiotic resistance genes are localized on conjugative plasmids closely related to pCW3, which has an atypical relaxase (Wisniewski and Rood, 2017;Wisniewski et al, 2016). The tcp region of pCW3 comprises 10 genes, and has been identified as the transfer region, showing some similarity to transfer genes of the conjugative transposon Tn916 (Bannam et al, 2006).…”