Detection of a soluble form of BACE-1 in human cerebrospinal fluid by a sensitive activity assay
Detection of a soluble form of BACE-1 in human cerebrospinal fluid by a sensitive activity assay
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DOI：
10.1373/clinchem.2006.066720
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发表时间：
2006-06-01
影响因子：
9.3
通讯作者：
Hanemaaijer, Roeland
中科院分区：
文献类型：
--
作者：
Verheijen, Jan H.;Huisman, Linda G. M.;Hanemaaijer, Roeland
背景资料：不溶性淀粉样β-肽沉积物的形成被认为与阿尔茨海默病（AD）中的神经变性有因果关系。β-肽通过蛋白水解酶的作用源自较大的淀粉样前体蛋白（APP）。导致淀粉样蛋白形成的第一个蛋白水解事件是由膜结合的胆固醇基蛋白酶BACE-1（也称为memapsin-2）切割APP。BACE-1的抑制被认为是AD的治疗方法。测定生物样品中BACE-1的活性将有助于阐明AD的发病机制和AD drugs.Methods的开发：我们开发了一种灵敏和特异的BACE-1活性测定法。该检测基于一种基因工程酶原，该酶原可被BACE-1特异性激活。用显色底物测量所得活性酶。使用2耦合反应产生低至0.4 pmol/L的检测限。结果：该测定检测BACE-1活性在人脑组织提取物，以及，出乎意料地，在人脑脊液（CSF）。结论：脑脊液中可溶性BACE-1的检测可用于监测候选药物的体内效应，并可用于诊断或预后判断。(c)2006年美国临床化学协会。
Background: Formation of deposits of the insoluble amyloid beta-peptide is believed to be causally related with neurodegeneration in Alzheimer disease (AD). The beta-peptide originates from a larger amyloid precursor protein (APP) by the action of proteolytic enzymes. The first proteolytic event leading to amyloid formation is the cleavage of APP by the membrane-bound aspartyl protease BACE-1, also known as memapsin-2. Inhibition of BACE-1 is thought to be a therapeutic approach to AD. Measuring BACE-1 activity in biological samples would be useful to elucidate the mechanism of AD and for development of AD drugs.Methods: We developed a sensitive and specific activity assay for BACE-1. The assay is based on a genetically engineered proenzyme that is specifically activated by BACE-1. The resulting active enzyme is measured with a chromogenic substrate. The use of 2 coupled reactions produces a detection limit as low as 0.4 pmol/L.Results: The assay detected BACE-1 activity in extracts of human brain tissue as well as, unexpectedly, in human cerebrospinal fluid (CSF). Gel electrophoresis and Western blotting identified the BACE-1 present in CSF as a truncated soluble form of the originally membrane-bound BACE-1.Conclusion: Detection of the soluble form of BACE-1 in CSF, a relatively easily accessible biological fluid, may be useful for monitoring the effects of drug candidates in vivo and may have diagnostic or prognostic applications. (c) 2006 American Association for Clinical Chemistry.