Low Density Lipoprotein Receptor Class A Repeats Are O-Glycosylated in Linker Regions
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Authors: | Nis Borbye Pedersen Shengjun Wang Yoshiki Narimatsu Zhang Yang Adnan Halim Katrine Ter-Borch Gram Schjoldager Thomas Daugbjerg Madsen Nabil G Seidah Eric Paul Bennett Steven B Levery Henrik Clausen |
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Institution: | From the ‡Copenhagen Center for Glycomics, Departments of Cellular and Molecular Medicine and School of Dentistry, Faculty of Health and Medical Sciences, University of Copenhagen, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark and ;the §Clinical Research Institute of Montreal, University of Montreal, Montreal, Quebec H2W 1R7, Canada |
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Abstract: | The low density lipoprotein receptor (LDLR) is crucial for cholesterol homeostasis and deficiency in LDLR functions cause hypercholesterolemia. LDLR is a type I transmembrane protein that requires O-glycosylation for stable expression at the cell surface. It has previously been suggested that LDLR O-glycosylation is found N-terminal to the juxtamembrane region. Recently we identified O-glycosylation sites in the linker regions between the characteristic LDLR class A repeats in several LDLR-related receptors using the “SimpleCell” O-glycoproteome shotgun strategy. Herein, we have systematically characterized O-glycosylation sites on recombinant LDLR shed from HEK293 SimpleCells and CHO wild-type cells. We find that the short linker regions between LDLR class A repeats contain an evolutionarily conserved O-glycosylation site at position −1 of the first cysteine residue of most repeats, which in wild-type CHO cells is glycosylated with the typical sialylated core 1 structure. The glycosites in linker regions of LDLR class A repeats are conserved in LDLR from man to Xenopus and found in other homologous receptors. O-Glycosylation is controlled by a large family of polypeptide GalNAc transferases. Probing into which isoform(s) contributed to glycosylation of the linker regions of the LDLR class A repeats by in vitro enzyme assays suggested a major role of GalNAc-T11. This was supported by expression of LDLR in HEK293 cells, where knock-out of the GalNAc-T11 isoform resulted in the loss of glycosylation of three of four linker regions. |
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Keywords: | Glycosylation Glycosyltransferase Lectin Lipoprotein Receptor-related Protein (LPR) Low Density Lipoprotein (LDL) GALNT O-Glycan Zinc Finger Nuclease |
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