By Weiming Xia, Huaxi Xu
Within the look for an efficient remedy for Alzheimer's affliction, APP is a different version protein that illustrates the wide variety of easy and complex characterization innovations to be had. Exploring a number of organic suggestions to explain the constitution and serve as of this transmembrane protein, this article provides each one process with specific, step by step protocols to accomplish reproducible effects and supply a framework for learning different membrane proteins.
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Extra resources for Amyloid Precursor Protein A Practical Approach
Brain Res. 779, 262, 1998. 23. Leveugle, B. et al. Heparin oligosaccharides that pass the blood–brain barrier inhibit beta-amyloid precursor protein secretion and heparin binding to beta-amyloid peptide. J. Neurochem. 70, 736, 1998. 24. Rossjohn, J. et al. Crystal structure of the N-terminal, growth factor-like domain of Alzheimer amyloid precursor protein. Nature Struct. Biol. 6, 327, 1999. 25. Hilbich, C. et al. Amyloid-like properties of peptides flanking the epitope of amyloid precursor protein-specific monoclonal antibody 22C11.
APP trafficking has been actively investigated in order to determine the cellular sites where Aβ is generated. 1 APP internalization is mediated via clathrin-coated pits2,3 in the canonical receptor-mediated endocytic pathway. 4–7 Whether raft-associated APP is internalized and, if so, whether this pool subsequently merges with the nonraft pool is unclear. Following internalization, a fraction of APP returns to the cell surface for additional rounds of endocytosis or secretion. The internalization signal for APP resides within its cytoplasmic domain and belongs to the NPXY-type signal, one of several well-recognized motifs.
The DMSO is added to the aliquot of sodium acetate such that the final DMSO concentration is 10% (including the amounts of substrate and inhibitor to be added, which are dissolved in DMSO). 12 Memapsin 25,12 enzyme stock (typically 6 µM, 50 µl aliquot) is added, followed by substrate (20 µl of 300 µM stock FS-2 in DMSO; see Reference 12) to initiate the reaction (2 ml total volume). Fluorescence intensity over a 5-min period was monitored with excitation at 328 nm and emission at 393 nm, using a detector voltage of 700 V on an Aminco Bowman luminescence spectrometer.
Amyloid Precursor Protein A Practical Approach by Weiming Xia, Huaxi Xu