RRC ID |
48818
|
Author |
Murakami-Sekimata A, Sato K, Sato K, Takashima A, Nakano A.
|
Title |
O-Mannosylation is required for the solubilization of heterologously expressed human beta-amyloid precursor protein in Saccharomyces cerevisiae.
|
Journal |
Genes Cells
|
Abstract |
In an attempt to express human beta-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 alpha-secretases, members of the Yapsin family, to produce N-terminal (sAPPalpha) and C-terminal (CTFalpha) domains, when its signal sequence was replaced by that of the yeast alpha-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 Deltapmt4 mutant lysate. This caused decreased production of CTFalpha. We conclude that O-mannosylation is required for the solubilization of exogenously expressed human APP.
|
Volume |
14(2)
|
Pages |
205-15
|
Published |
2009-2-1
|
DOI |
10.1111/j.1365-2443.2008.01263.x
|
PII |
GTC1263
|
PMID |
19170767
|
MeSH |
Amyloid beta-Protein Precursor / chemistry*
Amyloid beta-Protein Precursor / genetics
Amyloid beta-Protein Precursor / metabolism*
Animals
COS Cells
Chlorocebus aethiops
Gene Expression Regulation, Fungal
Glycosylation
Humans
Mannose / metabolism*
Protein O-Methyltransferase / genetics
Protein O-Methyltransferase / metabolism
Protein O-Methyltransferase / physiology
Protein Processing, Post-Translational / physiology*
Recombinant Proteins / chemistry
Recombinant Proteins / metabolism
Saccharomyces cerevisiae / genetics
Saccharomyces cerevisiae / metabolism*
Solubility
Transfection
|
IF |
1.655
|
Times Cited |
10
|
WOS Category
|
GENETICS & HEREDITY
CELL BIOLOGY
|
Resource |
DNA material |
pTU1-PalphaAPP (RDB08714) |