PILOT PROJECT-- IDENTIFICATION OF BETA AMYLOID PROTEIN SECRETASE

鉴于该实验室和其他实验室最近的演示，(I) 淀粉样β蛋白前体(BetaAPP)是由 β-APP分泌酶产生可识别的衍生物，(Ii) 内体-溶酶体系统处理可识别的羧基末端 β-APP衍生物可能是该途径中的重要中间体 产生淀粉样蛋白，以及(Iii)4kD的淀粉样β蛋白(Abeta) 通常是由培养细胞产生和释放的，现在它是现实的 负责确定关键的蛋白水解酶 Abeta的产生量。在这里，我们建议将实验重点放在 β-APP分泌酶的鉴定及其可行性探讨 利用反义cDNA表达快速鉴定这些蛋白水解酶 图书馆。我们计划在这些实验中重点研究β-APP分泌酶。 因为很可能很难分离出这种不寻常的 关键重要的酶传统地使用体外检测，基于 序列特异性和β-APP分泌酶的敲除 反义cDNA几乎肯定会增加表面的水平 达到荧光激活细胞可以检测到的程度 分类。

With the recent demonstration by this laboratory and others (i) that the amyloid beta protein precursor (betaAPP) is constitutively processed by betaAPP secretase to produce identifiable derivatives, (ii) that the endosomal-lysosomal system processes identifiable carboxyl-terminal betaAPP derivatives likely to be important intermediates in the pathway that produces amyloid, and (iii) that 4 kD amyloid beta protein (Abeta) is normally produced and released by cultured cells, it is now realistic to undertake the identification of the key proteases that determine the amount of Abeta produced. Here, we propose experimentation focused on the identification of betaAPP secretase that will explore the feasibility of rapidly identifying these proteases using antisense cDNA expression libraries. We plan to focus on betaAPP secretase in these experiments because it is likely to be difficult to isolate this unusual and critically important enzyme conventionally using in vitro assays based on sequence specificity and because the knockout of betaAPP secretase with antisense cDNAs will almost certainly increase the level of surface betaAPP to a degree that can be detected by fluorescence activated cell sorting.