Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines

BACKGROUND: Glycoproteins play a critical role in the cellular activities of eukaryotes. Sialic acid is typically the outermost monosaccharide of glycolipids and glycoproteins, and is necessary for normal development. RESULTS: A strategy based on avidin-biotin affinity was established to enrich sial...

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Autores principales:	Liang,Yangui, Hua,Qiang, Pan,Pengwei, Yang,Jie, Zhang,Qi
Lenguaje:	English
Publicado:	Sociedad de Biología de Chile 2015
Materias:	Click chemistry Sialic acid gp96 Sialyltransferase
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602015000100052
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spelling	oai:scielo:S0716-976020150001000522016-02-16Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell linesLiang,YanguiHua,QiangPan,PengweiYang,JieZhang,Qi Click chemistry Sialic acid gp96 Sialyltransferase BACKGROUND: Glycoproteins play a critical role in the cellular activities of eukaryotes. Sialic acid is typically the outermost monosaccharide of glycolipids and glycoproteins, and is necessary for normal development. RESULTS: A strategy based on avidin-biotin affinity was established to enrich sialylated glycoproteins from HeLa cervical carcinoma, SW1990 pancreatic adenocarcinoma, and A549 lung adenocarcinoma cells. Using HPLC-MS/MS, western blot, real-time PCR, and enzyme-linked immunosorbent assay, gp96 was identified in all three cell lines. No significant difference in the protein expression of gp96 was detected at the whole cell level, but the amount of bioti-nylated gp96 in SW1990 cells was 30-40 % lower than that in A549 and HeLa cells, and the amount of sialylated gp96 in SW1990 cells was 30 % lower than that in A549 and HeLa cells. Immunoblotting results showed that the expression of sialyltransferase proteins in the total cell lysates from HeLa and A549 cells were higher than that in SW1990 cells. CONCLUSIONS: We established a new method for investigating the expression and sialylation of glycoproteins using metabolic labeling, click chemistry, and avidin-biotin affinity. We successfully used this method to purify sialylated glycoproteins from cancer cell lines. Our results showed that the levels of gp96 sialylation varied across different cancer cell lines, and this may be because of differences in sialyltransferase expression.info:eu-repo/semantics/openAccessSociedad de Biología de ChileBiological Research v.48 20152015-01-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602015000100052en10.1186/S40659-015-0041-8
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	Click chemistry Sialic acid gp96 Sialyltransferase
spellingShingle	Click chemistry Sialic acid gp96 Sialyltransferase Liang,Yangui Hua,Qiang Pan,Pengwei Yang,Jie Zhang,Qi Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines
description	BACKGROUND: Glycoproteins play a critical role in the cellular activities of eukaryotes. Sialic acid is typically the outermost monosaccharide of glycolipids and glycoproteins, and is necessary for normal development. RESULTS: A strategy based on avidin-biotin affinity was established to enrich sialylated glycoproteins from HeLa cervical carcinoma, SW1990 pancreatic adenocarcinoma, and A549 lung adenocarcinoma cells. Using HPLC-MS/MS, western blot, real-time PCR, and enzyme-linked immunosorbent assay, gp96 was identified in all three cell lines. No significant difference in the protein expression of gp96 was detected at the whole cell level, but the amount of bioti-nylated gp96 in SW1990 cells was 30-40 % lower than that in A549 and HeLa cells, and the amount of sialylated gp96 in SW1990 cells was 30 % lower than that in A549 and HeLa cells. Immunoblotting results showed that the expression of sialyltransferase proteins in the total cell lysates from HeLa and A549 cells were higher than that in SW1990 cells. CONCLUSIONS: We established a new method for investigating the expression and sialylation of glycoproteins using metabolic labeling, click chemistry, and avidin-biotin affinity. We successfully used this method to purify sialylated glycoproteins from cancer cell lines. Our results showed that the levels of gp96 sialylation varied across different cancer cell lines, and this may be because of differences in sialyltransferase expression.
author	Liang,Yangui Hua,Qiang Pan,Pengwei Yang,Jie Zhang,Qi
author_facet	Liang,Yangui Hua,Qiang Pan,Pengwei Yang,Jie Zhang,Qi
author_sort	Liang,Yangui
title	Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines
title_short	Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines
title_full	Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines
title_fullStr	Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines
title_full_unstemmed	Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines
title_sort	development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in hela, sw1990 and a549 cell lines
publisher	Sociedad de Biología de Chile
publishDate	2015
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602015000100052
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Development of a novel method to evaluate sialylation of glycoproteins and analysis of gp96 sialylation in Hela, SW1990 and A549 cell lines

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