Beschreibung:
<jats:p>In cell extracts of <jats:italic>Lactobacillus delbrückii</jats:italic> ssp. <jats:italic>lactis</jats:italic> DSM7290 a peptidase with the ability to hydrolyse Phe‐β‐naphthylamide (Phe‐β‐NA) and His‐β‐NA could be detected. <jats:italic>Escherichia coli</jats:italic> lacking the enzyme activity in an enzymic plate assay was used to screen high‐copy‐number and low‐copy‐number plasmid libraries of size‐fractionated <jats:italic>Lactobacillus</jats:italic> DNA. Clones with the desired phenotype were detected, and the gene, designated <jats:italic>pepN</jats:italic>, was further subcloned and sequenced. A large open reading frame of 2529 nucleotides is predicted to encode a protein of 843 amino acids (95358 Da). Comparison of the <jats:italic>pepN</jats:italic> gene from <jats:italic>Lb. delbrückii</jats:italic> ssp. <jats:italic>lactis</jats:italic> DSM7290 indicates that it is homologous to genes of the family of Zn<jats:sup>2+</jats:sup>‐metallohydrolases and PepN shows identity with the active centre Zn<jats:sup>2+</jats:sup>‐binding motif of these enzymes. The substrate Lys‐β‐NA is more effectively cleaved than Phe‐β‐NA or His‐β‐NA which were used for screening in <jats:italic>E. coli</jats:italic>. The cloned <jats:italic>pepN</jats:italic> gene was efficiently overexpressed in <jats:italic>E. coli</jats:italic> and subcloning of the gene in <jats:italic>Lactobacillus casei</jats:italic> resulted in a moderate overexpression of approximately 20‐fold. The <jats:italic>pepN</jats:italic> gene product was purified from the <jats:italic>pepN</jats:italic>‐deficient <jats:italic>E. coli</jats:italic> strain CM89, using the substrate Lys‐<jats:italic>p</jats:italic>‐nitroanilide (Lys‐NH‐Ph) in the assay procedure. In a four‐step procedure including streptomycin sulfate precipitation, anion‐exchange chromatography and gel filtration the peptidase was purified to electrophoretic homogeneity.</jats:p>