Cloning and orientation of the gene encoding aminopeptidase N in Escherichia coli

The pepN gene, that encodes aminopeptidase N in Escherichia coli, has been cloned into the multicopy plasmid pBR322. Expression of the cloned pepN gene results in overproduction of the enzyme. The restriction map of the 6.7 Kb insert was established and the gene was further localized by analysis of the in vitro constructed delection plasmid and mutant plasmids generated by Tn5 insertions. Chromosome mobilization experiments, using pep-N-lac fusion strains allowed us to infer a clockwise direction of transcription for the pepN gene.