Glycoengineering of Human Cell Lines Using Zinc Finger Nuclease Gene Targeting: SimpleCells with Homogeneous GalNAc O-glycosylation Allow Isolation of the O-glycoproteome by One-Step Lectin Affinity Chromatography
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Glycoengineering of Human Cell Lines Using Zinc Finger Nuclease Gene Targeting : SimpleCells with Homogeneous GalNAc O-glycosylation Allow Isolation of the O-glycoproteome by One-Step Lectin Affinity Chromatography. / Steentoft, Catharina; Bennett, Eric Paul; Clausen, Henrik.
Glycosyltransferases: Methods and Protocols. ed. / Inka Brockhausen . Vol. 1022 Humana Press, 2013. p. 387-402 (Methods in molecular biology (Clifton, N.J.)).Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Communication
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TY - CHAP
T1 - Glycoengineering of Human Cell Lines Using Zinc Finger Nuclease Gene Targeting
T2 - SimpleCells with Homogeneous GalNAc O-glycosylation Allow Isolation of the O-glycoproteome by One-Step Lectin Affinity Chromatography
AU - Steentoft, Catharina
AU - Bennett, Eric Paul
AU - Clausen, Henrik
PY - 2013
Y1 - 2013
N2 - Lectin affinity chromatography is a powerful technique for isolation of glycoproteins carrying a specific glycan structure of interest. However, the enormous diversity of glycans present on the cell surface, as well as on individual proteins, makes it difficult to isolate an entire glycoproteome with one or even a series of lectins. Here we present a technique to generate cell lines with homogenous truncated O-glycans using zinc finger nuclease gene targeting. Because of their simplified O-glycoproteome, the cells have been named SimpleCells. Glycoproteins from SimpleCells can be isolated in a single purification step by lectin chromatography performed on a long lectin column. This protocol describes Zinc finger nuclease gene targeting of human cells to simplify the glycoproteome, as well as lectin chromatography and isolation of glycopeptides from total cell lysates of SimpleCells.
AB - Lectin affinity chromatography is a powerful technique for isolation of glycoproteins carrying a specific glycan structure of interest. However, the enormous diversity of glycans present on the cell surface, as well as on individual proteins, makes it difficult to isolate an entire glycoproteome with one or even a series of lectins. Here we present a technique to generate cell lines with homogenous truncated O-glycans using zinc finger nuclease gene targeting. Because of their simplified O-glycoproteome, the cells have been named SimpleCells. Glycoproteins from SimpleCells can be isolated in a single purification step by lectin chromatography performed on a long lectin column. This protocol describes Zinc finger nuclease gene targeting of human cells to simplify the glycoproteome, as well as lectin chromatography and isolation of glycopeptides from total cell lysates of SimpleCells.
U2 - 10.1007/978-1-62703-465-4_29
DO - 10.1007/978-1-62703-465-4_29
M3 - Book chapter
C2 - 23765677
SN - 978-1-62703-464-7
VL - 1022
T3 - Methods in molecular biology (Clifton, N.J.)
SP - 387
EP - 402
BT - Glycosyltransferases
A2 - Brockhausen , Inka
PB - Humana Press
ER -
ID: 47686302