Complete Nucleotide Sequencing and Molecular Characterization of the pColE9-J Plasmid

Abstract

The ColE9-J plasmid codes for colicin E9 (ColE9) protein, which is an antibacterial protein containing H-N-H endonuclease activity, that degrades chromosomal DNA of sensitive strains of Escherichia coli or closely related bacteria. The ColE9 plasmid has been used as a model system to understand various biological phenomenon such as colicin translocation and protein-protein interactions. In present study we have, for the first time, determined the complete nucleotide sequence of native ColE9-J plasmid. Computer based analysis of the complete nucleotide sequencing data of the pColE9-J plasmid consisting of 7577 bps revealed nine open reading frames (ORFs) located on the plasmid. Of these, seven ORFs including six ORFs coding for ColE9 operon and one ORF for replication region have been previously characterized. We have, thus, identified two more previously unrecognized putative OFRs, in addition to the seven previously characterized ORFs. One of these two ORFs exhibited close homology to the sequence coding for mobA gene of E. coli and it was located on positive strand; however the sequence for putative mobC gene was present on negative strand.