Potentially amyloidogenic fragment of 50 kDa and intracellular processing of amyloid precursor protein in cells cultured under leupeptin.
The principal neuropathological feature of Alzheimer's disease is extracellular deposition of approximately 4-kDa proteinous fragment, designated as beta-amyloid peptides (beta/A4 peptides) derived by proteolytic cleavage from amyloid precursor protein (APP), a large cell-surface receptor-like protein. There has been evidence that APP is proteolytically degraded in the secretory and endosomal/lysosomal pathways. The pathway in which APP is cleaved to generate beta/A4 peptides is still not identified. To clarify the intracellular processing of APP into the generation of beta/A4 peptides, we detected and characterized potentially amyloidogenic or non-amyloidogenic fragments using newly established monoclonal and polyclonal antibodies in the cultured cells with or without leupeptin, potent lysosomal protease inhibitor of lysosome. APP fragments of 50 and 20 kDa containing full-length beta/A4 peptides were identified in the cultured cells. Immunoblot analysis, biochemical study for specific marker enzyme activity of the fractions obtained from subcellular fractionation, sucrose density gradient centrifugation indicated that the 50-kDa APP fragment was produced in the compartment closely related to endosomal/lysosomal system. Our data suggest that the endosomal/lysosomal pathway is involved in the processing and generation of beta/A4 peptides.
Tsuzuki
K
K
Department of Microbiology, Sapporo Medical University School of Medicine, Japan.
Fukatsu
R
R
Takamaru
Y
Y
Fujii
N
N
Takahata
N
N
D000682:Amyloid / Q000737:chemistry / Q000378:metabolism*; D000686:Amyloidosis / Q000139:chemically induced*; D015153:Blotting, Western; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D006801:Humans; D007158:Immunologic Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism*; D000072002:Prion Proteins; D011328:Prions; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism
D000682:Amyloid; D007976:Leupeptins; C512656:PRNP protein, human; D010446:Peptide Fragments; D000072002:Prion Proteins; D011328:Prions; D011498:Protein Precursors; C032854:leupeptin