title: A Novel Glutamyl (Aspartyl)-Specific Aminopeptidase A from Lactobacillus delbrueckii with Promising Properties for Application.
abstract: Lactic acid bacteria (LAB) are auxotrophic for a number of amino acids. Thus, LAB have one of the strongest proteolytic systems to acquit their amino acid requirements. One of the intracellular exopeptidases present in LAB is the glutamyl (aspartyl) specific aminopeptidase (PepA; EC 3.4.11.7). Most of the PepA enzymes characterized yet, belonged to Lactococcus lactis sp., but no PepA from a Lactobacillus sp. has been characterized so far. In this study, we cloned a putative pepA gene from Lb. delbrueckii ssp. lactis DSM 20072 and characterized it after purification. For comparison, we also cloned, purified and characterized PepA from Lc. lactis ssp. lactis DSM 20481. Due to the low homology between both enzymes (30%), differences between the biochemical characteristics were very likely. This was confirmed, for example, by the more acidic optimum pH value of 6.0 for Lb-PepA compared to pH 8.0 for Lc-PepA. In addition, although the optimum temperature is quite similar for both enzymes (Lb-PepA: 60°C; Lc-PepA: 65°C), the temperature stability after three days, 20°C below the optimum temperature, was higher for Lb-PepA (60% residual activity) than for Lc-PepA (2% residual activity). EDTA inhibited both enzymes and the strongest activation was found for CoCl2, indicating that both enzymes are metallopeptidases. In contrast to Lc-PepA, disulfide bond-reducing agents such as dithiothreitol did not inhibit Lb-PepA. Finally, Lb-PepA was not product-inhibited by L-Glu, whereas Lc-PepA showed an inhibition.
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Timo
authors.initials: T
authors.lastname: Stressler
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Jacob
authors.initials: J
authors.lastname: Ewert
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Merz
authors.affiliation: Department of Food Microbiology and Hygiene, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Joshua
authors.initials: J
authors.lastname: Funk
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Claaßen
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Sabine
authors.initials: S
authors.lastname: Lutz-Wahl
authors.affiliation: Department of Food Microbiology and Hygiene, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Herbert
authors.initials: H
authors.lastname: Schmidt
authors.affiliation: Institute of Microbiology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Kuhn
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Lutz
authors.initials: L
authors.lastname: Fischer
mesh_terms: D003001:Cloning, Molecular / Q000379:methods; D043384:Glutamyl Aminopeptidase / Q000235:genetics*; D006867:Hydrolases / Q000235:genetics; D048668:Lactobacillus delbrueckii / Q000235:genetics*; D013294:Lactococcus lactis / Q000235:genetics; D017386:Sequence Homology, Amino Acid
chemical_list: D006867:Hydrolases; D043384:Glutamyl Aminopeptidase