CLONING AND DNA-SEQUENCE ANALYSIS OF PEPQ, A PROLIDASE GENE FROM LACTOBACILLUS-DELBRUECKII SUBSP LACTIS DSM7290 AND PARTIAL CHARACTERIZATION OF ITS PRODUCT

From a genomic library of Lactobacillus delbrueckii subsp. lactis (DSM 7290) DNA, in the low-copy-number vector pLG339, a recombinant clone w as selected, which complemented a mutation in the prolidase gene (pepQ ) of Escherichia coli UK173. Nucleotide sequence analysis revealed an open reading frame of 1104 nucleotides corresponding to a protein of 3 68 amino acids with a calculated pI of 4.64 and a molecular mass of 41 087 Da. The start site of pepQ transcription was determined by primer extension analysis with mRNA prepared from L. delbrueckii. Based on h omology of the gene product to various peptidases and on the substrate specificity determined, the peptidase was designated PepQ. The influe nce of Various protease inhibitors and cations on peptidase activity i ndicated that PepQ is a metalloprotease. The absence of a membrane-spa nning domain and a signal peptide sequence argues for a cytoplasmic lo calization of the enzyme.