Atypical Processing of Amyloid Precursor Fusion Protein by Proteolytic Activity in Pichia pastoris
| Autor: | Irina Morozova, Adam Markaryan, Bao-shiang Lee, Arnold Kaplan |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Glycosylation
Recombinant Fusion Proteins Blotting Western Molecular Sequence Data Glycine Biophysics Peptide Protein Sorting Signals Horseradish peroxidase Biochemistry Antibodies Pichia Pichia pastoris Amyloid beta-Protein Precursor Epitopes Amyloid precursor protein Serine Humans Amino Acid Sequence Peptide sequence Molecular Biology Horseradish Peroxidase chemistry.chemical_classification biology Cell Biology biology.organism_classification Fusion protein Molecular biology Peptide Fragments Molecular Weight Kinetics chemistry Culture Media Conditioned biology.protein Amyloid precursor protein secretase Dimerization Protein Processing Post-Translational Sequence Analysis Peroxidase |
| Zdroj: | Biochemical and Biophysical Research Communications. 263:596 |
| ISSN: | 0006-291X |
| Popis: | Secretases catalyze the production of important proteolytic products of the amyloid precursor protein. We expressed a fusion protein that contained horseradish peroxidase, fragment 590-695 of amyloid precursor protein, and c-myc and polyhistidine tags in Pichia pastoris. It secreted a 50-kDa N-terminal fragment; a 15-kDa C-terminal fragment accumulated in cells. The N-terminal fragment exhibited peroxidase activity and reacted with antibodies specific for peptides within the sequences -2 to 15 and 21-37 of beta-amyloid peptide. The C-terminal fragment reacted with antibodies that recognize the sequences 649-664 and 676-695 of amyloid precursor protein and the C-terminal c-myc tag. To locate the cut site, the C-terminal fragment was metabolically labeled with either [(35)S]Met or [(3)H]Lys and radiosequenced. A major component, derived from a cleavage at Gly(25)-Ser(26) of beta-amyloid, was detected. Results suggest a predominant atypical cleavage, like that observed in Down Syndrome fibroblasts, occurs between the alpha- and gamma-sites. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect