In fact, no conserved motifs or domains were detected in any of the LAB MobB proteins with interproscan
and only two transmembrane regions were found that were similarly positioned among them (Fig. 2c). In our opinion, the actual annotation of Orf6 and its related proteins deserves further investigation. It should be mentioned that fgenesb assigned the four mob genes to a single operon. However, this was not supported because all genes were preceded by individual promoters and RBS sequences, apart from mobA2, which Cetuximab lacked both cis-acting elements, further supporting our hypothesis for the disruption of an ancestral mobA gene into mobA1 and mobA2. In addition, a typical oriT detected upstream of mobC (position 1261–1355 nt) was almost identical to that of the pLJ42 plasmid (100% query coverage with 97% identity). Among the top blastp hits for RepA of pREN was the homologous protein of plasmid pUCL287 isolated from Tetragenococcus halophilus (formerly Pediococcus halophilus) (99% query coverage, 70% identity with e-value 8e−121). pUCL287 is the prototype for a family of theta-type replicons (Benachour et al., 1995, 1997). One of the main features of the pUCL287 family – apart from the actual sequence
Selleck DAPT of its Rep protein – is the distinct structure of the ori (Benachour et al., 1997). In pUCL287, ori spans 187 bp and is located upstream of the repA gene. The sequence is arranged in three different regions (Fig. 3a). The first is an AT-rich region, necessary for the melting of the two strands during plasmid replication, containing four 11-mer direct repeats, while the third region, which is the binding site of the Rep protein, consists of a 22-mer iteron tandemly repeated 4.5 times. These two regions are separated by a 37-bp variable sequence. Indeed, an ori sequence, carrying essentially all of the afore-mentioned features,
was detected in pREN, although a certain degree of deviation from the expected Montelukast Sodium sequence repeats (i.e. perfect 11-mer or 22-mer repeats) was observed. The same situation was evident for the ori sequences of other plasmids of the family. For this reason, we investigated whether a consensus could be determined for this region. Multiple sequence alignment of the oris of the pUCL287 family, found either predeposited in the GenBank files or manually determined by us (data not shown), was performed (Fig. 3a). According to our findings, the AT-rich stretch was highly conserved over its full length, and showed the presence of the consensus 9-mer sequence CCTCTTTT(A/T), tandemly repeated four times. In the 37-bp variable region, no repeats could be identified, although a significant number of conserved positions were observed, indicating that the region may not be entirely variable after all.