Kinouchi T, Nishio H, Nishiuchi Y, Tsunemi M, Takada K, Hamamoto T, Kagawa Y, Fujii N
Department of Radiation Life Science and Radiation Medical Science, Research Reactor Institute, Kyoto University, Osaka, Japan.
Amino Acids. 2007 Jan;32(1):79-85. doi: 10.1007/s00726-006-0348-4. Epub 2006 Oct 6.
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.
衰老过程中蛋白质中天冬氨酸D-异构体（D-Asp）的积累与阿尔茨海默病（AD）、白内障和动脉硬化的发病机制有关。在此，我们鉴定出一种特异性的对乳胞素敏感的内肽酶，它能切割含D-Asp的蛋白质，并将其命名为D-天冬氨酰内肽酶（DAEP）。DAEP具有多聚体结构（分子量：600 kDa），定位于线粒体内膜。然而，在大肠杆菌、酿酒酵母和秀丽隐杆线虫中未检测到DAEP活性。一种新合成的DAEP特异性抑制剂i-DAEP：（苯甲酰-L-精氨酸-L-组氨酸-[D-天冬氨酸]-CH₂Cl；分子量：563.01）可抑制DAEP活性（IC₅₀，3 μM），其对DAEP的抑制作用比乳胞素强10倍。另一方面，i-DAEP对20S或26S蛋白酶体均无抑制作用。我们通过使用生物素化的i-DAEP进行亲和标记，鉴定出琥珀酸脱氢酶和谷氨酸脱氢酶1是DAEP的组成成分。在哺乳动物的长寿过程中，DAEP可能作为一种清除剂，对抗衰老过程中消旋化蛋白质的积累。对DAEP的深入了解将为开发治疗AD等与含D-Asp蛋白质积累有关疾病的方法奠定基础。
