Cell and Matrix Biology, Johannes Gutenberg-University, 55128 Mainz, Germany.
J Biol Chem. 2011 Aug 5;286(31):27741-50. doi: 10.1074/jbc.M111.252718. Epub 2011 Jun 6.
Identification of physiologically relevant substrates is still the most challenging part in protease research for understanding the biological activity of these enzymes. The zinc-dependent metalloprotease meprin β is known to be expressed in many tissues with functions in health and disease. Here, we demonstrate unique interactions between meprin β and the amyloid precursor protein (APP). Although APP is intensively studied as a ubiquitously expressed cell surface protein, which is involved in Alzheimer disease, its precise physiological role and relevance remain elusive. Based on a novel proteomics technique termed terminal amine isotopic labeling of substrates (TAILS), APP was identified as a substrate for meprin β. Processing of APP by meprin β was subsequently validated using in vitro and in vivo approaches. N-terminal APP fragments of about 11 and 20 kDa were found in human and mouse brain lysates but not in meprin β(-/-) mouse brain lysates. Although these APP fragments were in the range of those responsible for caspase-induced neurodegeneration, we did not detect cytotoxicity to primary neurons treated by these fragments. Our data demonstrate that meprin β is a physiologically relevant enzyme in APP processing.
鉴定生理相关的底物仍然是蛋白酶研究中最具挑战性的部分,这有助于理解这些酶的生物学活性。锌依赖性金属蛋白酶 meprin β 已知在许多组织中表达,在健康和疾病中发挥作用。在这里,我们证明了 meprin β 和淀粉样前体蛋白(APP)之间的独特相互作用。尽管 APP 作为一种广泛表达的细胞表面蛋白,被深入研究与阿尔茨海默病有关,但它的确切生理作用和相关性仍然难以捉摸。基于一种称为末端胺同位素标记的底物(TAILS)的新型蛋白质组学技术,鉴定到 APP 是 meprin β 的底物。使用体外和体内方法随后验证了 meprin β 对 APP 的加工。在人脑中发现了约 11 和 20 kDa 的 APP N 端片段,但在 meprin β(-/-) 小鼠脑裂解物中未发现。尽管这些 APP 片段在负责半胱天冬酶诱导的神经退行性变的范围内,但我们没有检测到用这些片段处理的原代神经元的细胞毒性。我们的数据表明,meprin β 是 APP 加工中一种生理相关的酶。