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O ‐Mannosylation is required for the solubilization of heterologously expressed human β‐amyloid precursor protein in Saccharomyces cerevisiae
Author(s) -
MurakamiSekimata Akiko,
Sato Ken,
Sato Ken,
Takashima Akihiko,
Nakano Akihiko
Publication year - 2009
Publication title -
genes to cells
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.912
H-Index - 115
eISSN - 1365-2443
pISSN - 1356-9597
DOI - 10.1111/j.1365-2443.2008.01263.x
Subject(s) - yeast , biology , saccharomyces cerevisiae , endoplasmic reticulum , glycosylation , golgi apparatus , biochemistry , mutant , amyloid precursor protein , gene , medicine , disease , pathology , alzheimer's disease
In an attempt to express human β‐amyloid precursor protein (APP) in yeast, we fortuitously found that this protein is only O ‐glycosylated in yeast. APP was effectively expressed in yeast, processed by yeast α‐secretases, members of the Yapsin family, to produce N‐terminal (sAPPα) and C‐terminal (CTFα) domains, when its signal sequence was replaced by that of the yeast α‐mating factor. APP is known to acquire N‐ and O‐ glycosylation through the endoplasmic reticulum (ER) and the Golgi apparatus and is transported to the plasma membrane in mammalian cells. In spite of the presence of canonical N‐ glycosylation consensus sequences, APP was not N ‐glycosylated in the yeast system. Pulse‐chase experiments demonstrated that APP received only O ‐mannosylation in yeast. Examination of yeast pmt mutants, which are defective in the initiation of O ‐mannosylation in the ER, revealed that Pmt4p is most responsible for the oligosaccharide modification of APP. Maturation of APP was slowed down and aggregated forms of APP were observed by sucrose density gradient fractionation of the Δpmt4 mutant lysate. This caused decreased production of CTFα. We conclude that O ‐mannosylation is required for the solubilization of exogenously expressed human APP.