title: Characterization, cloning, sequencing, and expression of an aminopeptidase N from Streptomyces sp. TH-4.
abstract: The aminopeptidase N (TH-4AP) of Streptomyces sp. TH-4 was purified from a culture supernatant. The purified enzyme had a molecular mass of 95 kDa. The gene encoding TH-4AP was cloned and sequenced. The primary structure of the protein possessed the PepN-conserved motif GxMEN and the zinc-binding motif HExxHx18E, and showed 88% identity with that of PepN from Streptomyces lividans strain 66. We succeeded in overproducing a His-tagged recombinant enzyme using Escherichia coli. The enzyme had a 1.5-fold higher activity in the presence of cobalt ions than in their absence. To evaluate the possible application of TH-4AP to decrease the content of bitter peptides, we investigated the ability of Streptomyces aminopeptidases to hydrolyze synthetic peptides by a coupling method using L-amino acid oxidase and peroxidase. The substrate specificity of TH-4AP toward synthetic peptides was significantly different from that toward aminoacyl-p-nitroanilide derivatives.
authors.affiliation: Research Institute for Biological Sciences, Okayama (RIBS, Okayama), 7549-1 Kibichuo-cho, Kaga-gun, Okayama 716-1241, Japan. hatanaka@bio-ribs.com
authors.forename: Tadashi
authors.initials: T
authors.lastname: Hatanaka
authors.forename: Jiro
authors.initials: J
authors.lastname: Arima
authors.forename: Misugi
authors.initials: M
authors.lastname: Uraji
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Uesugi
authors.forename: Masaki
authors.initials: M
authors.lastname: Iwabuchi
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001709:Biotechnology; D018826:CD13 Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D018826:CD13 Antigens