The catalytic domain of insulin-degrading enzyme forms a denaturant-resistant complex with amyloid β peptide -: Implications for Alzheimer disease pathogenesis
The catalytic domain of insulin-degrading enzyme forms a denaturant-resistant complex with amyloid β peptide -: Implications for Alzheimer disease pathogenesis
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DOI：
10.1074/jbc.m706316200
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发表时间：
2008-06-20
影响因子：
4.8
通讯作者：
Castano, Eduardo M.
中科院分区：
文献类型：
--
作者：
Llovera, Ramiro E.;de Tullio, Matias;Castano, Eduardo M.
在哺乳动物大脑中，胰岛素降解酶（IDE）是胰岛素转化和β淀粉样蛋白（A β）降解的核心。生化和遗传数据支持IDE可能在晚发性阿尔茨海默病（AD）中发挥作用的观点，最近的研究表明AD与2型糖尿病之间存在关联。在这里，我们证明了一个天然折叠的重组IDE能够与a β形成稳定的配合物，在强变性剂处理后抵抗解离。这种相互作用也在大鼠脑IDE中观察到，并在AD皮质组织的sds可溶性部分中检测到。已知在淀粉样蛋白组装中起关键作用的β序列17-27足以与IDE形成稳定的复合物。与聚集的A β相反，单体A β能够在非常缓慢的动力学（t（1/2）类似于45分钟）后与IDE发生不可逆的关联。IDE的部分变性以及10倍摩尔过量胰岛素的预孵育阻止了复合物的形成，这表明a β至少与蛋白酶的部分底物结合位点发生了不可逆的相互作用。有限的蛋白水解表明，A β仍然以sds抗性的方式与IDE的类似25 kda n端片段结合。凝胶消化IDE后的质谱分析。β络合物表明，从包含IDE催化位点的区域衍生的肽被A β回收。综上所述，这些结果提示了一种前所未有的构象依赖性底物结合机制，这种机制可能会干扰A β清除、胰岛素周转，并促进AD的发病。
Insulin-degrading enzyme (IDE) is central to the turnover of insulin and degrades amyloid beta(A beta) in the mammalian brain. Biochemical and genetic data support the notion that IDE may play a role in late onset Alzheimer disease (AD), and recent studies suggest an association between AD and diabetes mellitus type 2. Here we show that a natively folded recombinant IDE was capable of forming a stable complex with A beta that resisted dissociation after treatment with strong denaturants. This interaction was also observed with rat brain IDE and detected in an SDS-soluble fraction from AD cortical tissue. A beta sequence 17-27, known to be crucial in amyloid assembly, was sufficient to form a stable complex with IDE. Monomeric as opposed to aggregated A beta was competent to associate irreversibly with IDE following a very slow kinetics (t(1/2) similar to 45 min). Partial denaturation of IDE as well as preincubation with a 10-fold molar excess of insulin prevented complex formation, suggesting that the irreversible interaction of A beta takes place with at least part of the substrate binding site of the protease. Limited proteolysis showed that A beta remained bound to a similar to 25-kDa N-terminal fragment of IDE in an SDS-resistant manner. Mass spectrometry after in gel digestion of the IDE.A beta complex showed that peptides derived from the region that includes the catalytic site of IDE were recovered with A beta. Taken together, these results are suggestive of an unprecedented mechanism of conformation-dependent substrate binding that may perturb A beta clearance, insulin turnover, and promote AD pathogenesis.