Compelling evidence suggests that N-terminally truncated and pyroglutamyl-modified amyloid-beta (A beta) peptides play a major role in the development of Alzheimer's disease. Posttranslational formation of pyroglutamic acid (pGlu) at position 3 or 11 of A beta implies cyclization of an N-terminal glutamate residue rendering the modified peptide degradation resistant, more hydrophobic, and prone to aggregation. Previous studies using artificial peptide substrates suggested the potential involvement of the enzyme glutaminyl cyclase in generation of pGlu-A. Here we show that glutaminyl cyclase (QC) catalyzes the formation of A beta(3(pE)-40/42) after amyloidogenic processing of APP in two different cell lines, applying specific ELISAs and Western blotting based on urea-PAGE. Inhibition of QC by the imidazole derivative PBD150 led to a blockage of A beta(3(pE)-42) formation. Apparently, the QC-catalyzed formation of N-terminal pGlu is favored in the acidic environment of secretory compartments, which is also supported by double-immunofluorescence labeling of QC and APP revealing partial colocalization. Finally, initial investigations focusing on the molecular pathway leading to the generation of truncated A beta peptides imply an important role of the amino acid sequence near the beta-secretase cleavage site. Introduction of a single-point mutation, resulting in an amino acid substitution, APP(E599Q), i.e., at position 3 of A beta, resulted in significant formation Of A beta(3(pE)-40/42). Introduction of the APP KM595/596NL Swedish mutation causing overproduction of A beta, however, surprisingly diminished the concentration of A beta(3(pE)-40/42). The study provides new cell-based assays for the profiling of small molecule inhibitors of QC and points to conspicuous differences in processing of APP depending on sequence at the beta-secretase cleavage site.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据