Amyloid (-protein (A() is the major protein constituent found in the senile plaques of Alzheimer's disease (AD) and is produced by proteolytic processing of a large type-I transmembrane protein, (-amyloid precursor protein ((APP). Biochemical studies have demonstrated that genetic mutations in presenilin genes (PS1 and PS2) give rise elevated A(1-42. We have recently reported that A(1-43 can be detected from cultured cell media conditioned by mouse neuroblastoma (N2a) cells stably transfected with human APP695 cDNA and PS1 cDNA carrying exon-9 deletion mutation in addition to other carboxy-terminal heterogeneous A( peptides using immunoprecipitation/mass spectrometry assay. We have further investigated the generation of this A(1-43 species. The results from these studies have revealed several molecular characteristics of this A(1-43 species. Its generation is not correlated to the total A( level and neither the ratio of A(1-42 to A(1-40. It has only being detected from cell culture media conditioned by cells expressing PS1 with exon-9 deletion mutation but not other PS1 mutations including M146L and A246E point mutations. The effect of the exon-9 deletion of PS1 on production of this A(1-43 peptide is same for both wild type APP and mutant APP with Swedish mutation. Intracellular A( peptide profile and subcellular A( peptide distribution were analyzed by subcellular fractionation and cont.... immunoprecipitation/mass spectrometry, which reveals detailed information on the effect of PS1 mutations on intracellular A( peptide production. These ex vivo experimental data will be reported in this presentation and should help us to understand how PS1 mutations function on APP processing and production of A(.
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