Cloning and sequence analysis of the X-prolyl-dipeptidyl-aminopeptidase gene (pepX) from Lactobacillus delbrückii ssp. lactis DSM7290.
Lactobacillus delbrückii ssp. lactis DSM7290 possesses an X-prolyl-dipeptidyl-aminopeptidase, designated PepX, which catalyses the hydrolytic removal of N-terminal dipeptidyl residues from peptides containing proline in the penultimate position. Using the specific substrate L-Ala-L-Pro-p-nitroanilide, PepX was purified by a four-step procedure including ammonium sulphate fractionation, hydrophobic interaction chromatography, ion exchange chromatography, and affinity chromatography. The N-terminus of the purified protein was sequenced. Screening of a gene library of chromosomal Lactobacillus delbrückii ssp. lactis DSM7290 DNA in the low-copy-number vector pLG339 resulted in the identification of the pepX gene in Escherichia coli using a specific plate assay with Gly-L-Pro-beta-naphthylamide as substrate. Nucleotide sequence analysis revealed an open reading frame of 2376 bp, coding for a protein of 792 amino acids with a molecular mass of 88449 Da.
Meyer-Barton
E C
EC
Universität Kaiserslautern, Abteilung Mikrobiologie, Germany.
Klein
J R
JR
Imam
M
M
Plapp
R
R
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D018819:Dipeptidyl Peptidase 4; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics*; D007778:Lactobacillus / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA
D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D018819:Dipeptidyl Peptidase 4