Characterization of the replication region of the small cryptic plasmid of Campylobacter hyointestinalis

Abstract

The complete nucleotide sequence of a 2.5-kb cryptic plasmid from Campylobacter hyointestinalis was determined. Only one open reading frame (ORF), encoding a polypeptide of M r 39 667, designated RepA, could be identified within the sequence. This was confirmed by minicell analysis. Analysis of the region upstream from the ORF showed an A+T-rich region followed by four 19-bp direct repeats. Together, these features are characteristic of other replication origins ( ori(s)). The promoter sequence of the repA gene was identified by primer extension analysis and both the putative —10 and —35 regions were found to lie within two potential hairpin-loop structures. RepA showed marked amino acid sequence homology to a replication-initiation protein from the Neisseria gonorrhoeae plasmid, pFA3, and with other replication-initiation proteins over two conserved motifs. A putative partitioning ( par) locus was identified upstream from the ori and consisted of a perfect 9-bp inverted repeat and six putative DNA gyrase-binding sites. A putative mobilization origin ( oriT) region was identified. This featured a 19-bp imperfect inverted repeat adjacent to a sequence of 12 bp which showed strong homology to the consensus sequence of the ‘nick regions’ in a variety of oriTs of other plasmids.