Isolation and characterization of aminopeptidase (Jc-peptidase) from Japanese cedar pollen (Cryptomeria japonica).
An aminopeptidase, Jc-peptidase, was purified from Japanese cedar pollen by seven steps, including precipitation with ammonium sulfate, ion-exchange chromatography, gel filtration, hydrophobic interaction chromatography on phenyl-agarose, and high-performance liquid chromatography. Purified Jc-peptidease has a molecular weight of 42 kDa and hydrolyzes the synthetic substrates of L-phenylalanyl-4-methylcoumaryl-7-amide (Phe-MCA) with Km = 5 x 10(-5) M, Tyr-MCA with Km = 7 x 10(-4) M, Leu-MCA with Km = 1 x 10(-3) M, and Met-MCA with Km = 1 x 10(-3) M. Other MCA analogues such as Arg-MCA or Glu-MCA failed to serve as its substrates. The activity was inhibited in the presence of phebestin, [(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl-L-valyl]-L-phenylalanine, with Ki = 4.7 x 10(-5) M, or bestatin, [(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoyl]-L-leucine, with Ki = 1.1 x 10(-4) M. According to amino acid sequence analysis, the N-terminal amino group seems to be blocked. The physiological function of the aminopeptidase (Jc-peptidase) has not been clarified in vivo.
Noguchi
Yukinori
Y
Toin Human Science and Technology Center and Department of Biomedical Engineering, Toin University of Yokohama, 1614, Kurogane-cho, Aoba-ku, Yokohama 225-8502, Japan.
Nagata
Hiroyuki
H
Koganei
Hajime
H
Kodera
Yoh
Y
Hiroto
Misao
M
Nishimura
Hiroyuki
H
Inada
Yuji
Y
Matsushima
Ayako
A
D000626:Aminopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D000645:Ammonium Sulfate; D011232:Chemical Precipitation; D002845:Chromatography; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D029779:Cupressaceae / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008970:Molecular Weight; D011058:Pollen / Q000201:enzymology*; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D000626:Aminopeptidases; D000645:Ammonium Sulfate