Fischer Frauke, Molinari Maurizio, Bodendorf Ursula, Paganetti Paolo
Nervous System, Novartis Pharma AG, Basel, Switzerland. Institute for Research in Biomedicine, Bellinzona, Switzerland.
J Neurochem. 2002 Mar;80(6):1079-88. doi: 10.1046/j.0022-3042.2002.00806.x.
beta-Site APP-cleaving enzyme (BACE) initiates the processing of the amyloid precursor protein (APP) leading to the generation of beta-amyloid, the main component of Alzheimer's disease senile plaques. BACE (Asp2, memapsin 2) is a type I transmembrane aspartic protease responsible for the beta-secretase cleavage of APP producing a soluble form of the ectodomain (sAPPbeta) and the membrane-bound, carboxy-terminal intermediates C99 and C89. BACE maturation involves cysteine bridge formation, N -glycosylation and propeptide removal. We investigated variants of BACE in which the disulphide bonds of the catalytic domain spanning between Cys216/Cys420, Cys278/Cys443 and Cys330/Cys380 were removed by mutagenesis. When transfected in cultured cells, these mutants showed impaired maturation. Nevertheless, a fraction of mutated protein retained both the competence to mature as well as the activity to process APP. For the generation of a functional enzyme the conserved Cys330/Cys380 bond was the most critical, whereas the two bonds between Cys216/Cys420 and Cys278/Cys443, which are typical for the membrane-bound BACE, appeared to be less important.
β-位点淀粉样前体蛋白裂解酶（BACE）启动淀粉样前体蛋白（APP）的加工过程，导致β-淀粉样蛋白的产生，β-淀粉样蛋白是阿尔茨海默病老年斑的主要成分。BACE（Asp2，膜天冬氨酸蛋白酶2）是一种I型跨膜天冬氨酸蛋白酶，负责APP的β-分泌酶裂解，产生细胞外结构域的可溶性形式（sAPPβ）以及膜结合的羧基末端中间体C99和C89。BACE的成熟涉及半胱氨酸桥的形成、N-糖基化和前肽的去除。我们研究了BACE的变体，其中通过诱变去除了催化结构域中位于Cys216/Cys420、Cys278/Cys443和Cys330/Cys380之间的二硫键。当在培养细胞中进行转染时，这些突变体显示出成熟受损。然而，一部分突变蛋白既保留了成熟的能力，也保留了加工APP的活性。对于功能性酶的产生，保守的Cys330/Cys380键最为关键，而Cys216/Cys420和Cys278/Cys443之间的两个键（这是膜结合BACE的典型键）似乎不太重要。
