A yeast genetic assay for caspase cleavage of the amyloid-β precursor protein

被引：0
作者：
Paul L. Gunyuzlu
W. Hunter White
Gary L. Davis
Gregory F. Hollis
Jeremy H. Toyn
机构：
[1] DuPont Pharmaceuticals,
[2] Experimental Station E336/239A,undefined
来源：
Molecular Biotechnology | 2000年 / 15卷
关键词：
Alzheimer’s disease; amyloid-β; caspase; yeast;
D O I：
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学科分类号：
摘要：
A functional assay for proteolytic processing of the amyloid precursor protein (APP) was set up in yeast. This consisted of a membrane-bound chimeric protein containing the β-secretase cleaved C-terminal fragment of APP fused to the Gal4 transcription factor. Using this chimera in a GAL-reporter yeast strain, an expression library of human cDNAs was screened for clones that could activate the GAL-reporter genes by proteolytic processing of the membrane-bound APP-Gal4. Two human proteases, caspase-3 and caspase-8, were identified and confirmed to act by a mechanism that involved proteolysis at the site in the APP-Gal4 chimera that corresponded to the natural caspase cleavage site in APP, thus linking a readily scorable phenotype to proteolytic processing of APP. The activation of caspase-3 involved a mechanism that was independent of aspartic acid residue 175 at the cleavage site normally required for processing of caspase-3.
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页码：29 / 37
页数：8
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