Characterization of the lep operon of Escherichia coli. Identification of the promoter and the gene upstream of the signal peptidase I gene

Abstract

The DNA sequence flanking the gene (lep) encoding signal peptidase I of Escherichia coli has been determined. The upstream flanking sequence contains a gene (lepA) that encodes a polypeptide of 598 amino acid residues and terminates 18 base pairs from the initiation codon of the lep gene. The position of the lep promoter was determined by both gene fusion with the lacZ gene as well as by S1 nuclease mapping of the lep mRNA to be 73 base pairs upstream from the initiation codon of the lepA gene. The lepA gene was cloned into a high expression vector (pIN-III), and its gene product was identified to be a protein of apparent molecular weight of 76,000. This gene product was preferentially localized in the cytoplasmic membrane and periplasmic fractions upon subcellular fractionation. The DNA sequence immediately downstream of the lep gene contains features consistent with a rho factor-independent transcriptional termination site, indicating that the lep operon encodes only two proteins (lepA and lep).