Isolation and characterization of mammalian D-aspartyl endopeptidase.
The accumulation of D-isomers of aspartic acid (D-Asp) in proteins during aging has been implicated in the pathogenesis of Alzheimer's disease (AD), cataracts and arteriosclerosis. Here, we identified a specific lactacystin-sensitive endopeptidase that cleaves the D-Asp-containing protein and named it D-aspartyl endopeptidase (DAEP). DAEP has a multi-complex structure (MW: 600 kDa) and is localized in the inner mitochondrial membrane. However, DAEP activity was not detected in E. coli, S. cerevisiae, and C. elegans. A specific inhibitor for DAEP, i-DAEP: (benzoyl-L-Arg-L-His-[D-Asp]-CH(2)Cl; MW: 563.01), was newly synthesized and inhibited DAEP activity (IC(50), 3 microM), a factor of ten greater than lactacystin on DAEP. On the other hand, i-DAEP did not inhibit either the 20S or 26S proteasome. And we identified succinate dehydrogenase and glutamate dehydrogenase 1 as components of DAEP by affinity label using biotinylated i-DAEP. In the long life span of mammals, DAEP may serve as a scavenger against accumulation of racemized proteins in aging. Insights into DAEP will provide the foundation for developing treatments of diseases, such as AD, in which accumulation of D-Asp-containing proteins are implicated.
Kinouchi
T
T
Department of Radiation Life Science and Radiation Medical Science, Research Reactor Institute, Kyoto University, Osaka, Japan. kinouchi@HL.rri.kyoto-u.ac.jp
Nishio
H
H
Nishiuchi
Y
Y
Tsunemi
M
M
Takada
K
K
Hamamoto
T
T
Kagawa
Y
Y
Fujii
N
N
D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000737:chemistry; D000375:Aging / Q000378:metabolism; D000544:Alzheimer Disease / Q000188:drug therapy / Q000201:enzymology*; D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D017173:Caenorhabditis elegans / Q000201:enzymology; D026603:D-Aspartic Acid / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D005969:Glutamate Dehydrogenase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification; D051379:Mice; D051336:Mitochondrial Membranes / Q000201:enzymology*; D024101:Mitochondrial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification; D009842:Oligopeptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000627:therapeutic use; D011480:Protease Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000627:therapeutic use; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry; D061988:Proteasome Inhibitors; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013385:Succinate Dehydrogenase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification
D024101:Mitochondrial Proteins; D009097:Multienzyme Complexes; D009842:Oligopeptides; D011480:Protease Inhibitors; D061988:Proteasome Inhibitors; C524547:benzoyl-L-Arg-L-His-(D-Asp)-CH(2)Cl; C067713:lactacystin; D026603:D-Aspartic Acid; D013385:Succinate Dehydrogenase; D005969:Glutamate Dehydrogenase; D016282:Aspartic Acid Endopeptidases; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine