title: Purification and characterization of a major kyotorphin-hydrolyzing peptidase of rat brain.
abstract: We purified a major kyotorphin (L-Tyr-L-Arg)-hydrolyzing peptidase (KTPase) from the rat brain, to electrophoretic homogeneity using conventional chromatographic techniques. KTPase was purified 1,660-fold with a specific activity of 161 mumol/min/mg protein and 6.8% recovery. The purified enzyme was composed of a single polypeptide with a molecular mass of 67 kDa and an isoelectric point (pI) of 5.5. KTPase has the ability to hydrolyze a variety of natural dipeptides. It also liberated NH2-terminal tyrosine from Tyr-Gly-Gly and Tyr-Tyr-Leu. Bestatin and arphamenine B were potent inhibitors of this enzyme, while amastatin and puromycin had little effect. An excess of anti-KTPase antibody raised in a white rabbit precipitated approximately 80% of the kyotorphin-hydrolyzing activity in the cytosol of rat brain. These data suggested that 67 kDa KTPase has a role in the degradation of kyotorphin within neuronal cells of the rat brain.
authors.affiliation: Faculty of Pharmacy & Pharmaceutical Sciences, Fukuyama University, Hiroshima.
authors.forename: K
authors.initials: K
authors.lastname: Akasaki
authors.forename: H
authors.initials: H
authors.lastname: Yoshimoto
authors.forename: A
authors.initials: A
authors.lastname: Nakamura
authors.forename: H
authors.initials: H
authors.lastname: Shiomi
authors.forename: H
authors.initials: H
authors.lastname: Tsuji
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology*; D004150:Dipeptidases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D007700:Kinetics; D008297:Male; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012967:Sodium Dodecyl Sulfate; D013379:Substrate Specificity
chemical_list: D011480:Protease Inhibitors; D012967:Sodium Dodecyl Sulfate; D004150:Dipeptidases; C073383:kyotorphin I dipeptidase