title: Nardilysin cleaves peptides at monobasic sites.
abstract: Nardilysin (N-arginine dibasic convertase, EC 3.4.24.61) was first identified on the basis of its ability to cleave peptides containing an arginine dibasic pair, i.e., Arg-Arg or Arg-Lys. However, it was observed that an aromatic residue adjacent to the dibasic pair (i.e., Phe-Arg-Lys) could alter the cleavage site. In this study we determined whether nardilysin can cleave peptides at a single basic residue. Nardilysin cleaves beta-endorphin at the monobasic site, Phe(17)-Lys(18), with a k(cat)/K(m) of 2 x 10(8) M(-)(1) min(-)(1). This can be compared to a k(cat)/K(m) of 8.5 x 10(8) M(-)(1) min(-)(1) for cleavage between a dibasic pair in dynorphin B-13. Nardilysin also cleaves calcitonin at His-Arg and somatostatin-14 at Cys-Lys. We examined the hydrolysis of fluorogenic peptides based on the beta-endorphin 12-24 sequence, Abz-T-P-L-V-T-L-X(1)-X(2)-N-A-I-I-K-Q-EDDnp. Nardilysin hydrolyzes the peptides when X(1)-X(2) = F-K, F-R, W-K, M-K, Y-K, and L-K. The kinetics of cleavage at F-K and F-R are similar; however, K-F is not hydrolyzed. Nardilysin cleaves at two monobasic sites M-K and F-R of the kallidin model peptide Abz-MISLMKRPPGFSPFRSSRI-NH(2), releasing desArg(10) kallidin (KRPPGFSPF). However, nardilysin does not release desArg(10) kallidin from the physiological precursor low molecular weight kininogen. These studies extend the range of potential substrates for nardilysin and further substantiate that nardilysin is a true peptidase.
authors.affiliation: Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, Kentucky 40536-0298, USA.
authors.forename: K Martin
authors.initials: KM
authors.lastname: Chow
authors.forename: Oliver
authors.initials: O
authors.lastname: Oakley
authors.forename: Jack
authors.initials: J
authors.lastname: Goodman
authors.forename: Zhangliang
authors.initials: Z
authors.lastname: Ma
authors.forename: Maria Aparecida
authors.initials: MA
authors.lastname: Juliano
authors.forename: Luiz
authors.initials: L
authors.lastname: Juliano
authors.forename: Louis B
authors.initials: LB
authors.lastname: Hersh
mesh_terms: D000595:Amino Acid Sequence; D024322:Amino Acids, Aromatic / Q000737:chemistry; D024361:Amino Acids, Basic / Q000737:chemistry*; D000599:Amino Acids, Diamino / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D007704:Kininogens / Q000737:chemistry; D008666:Metalloendopeptidases / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000138:chemical synthesis / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D013004:Somatostatin / Q000737:chemistry; D013552:Swine; D001615:beta-Endorphin / Q000737:chemistry
chemical_list: D024322:Amino Acids, Aromatic; D024361:Amino Acids, Basic; D000599:Amino Acids, Diamino; D007704:Kininogens; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D013004:Somatostatin; D001615:beta-Endorphin; D008666:Metalloendopeptidases; C087976:nardilysin