Novel carboxylated N-glycans contain oligosaccharide-linked glutamic acid

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• 作者：Srikrishna ; Geetha ; Brive ; Lena ; Freeze ; Hudson H.
• 刊名：Biochemical and Biophysical Research Communications
• 出版年：2005
• 出版时间：July 15, 2005
• 年：2005
• 卷：332
• 期：4
• 页码：1020-1027
• 全文大小：533 K

文摘

We previously reported that N-glycans from bovine lung contain novel carboxylate groups. Here, we provide evidence that the carboxylated glycans contain glutamic acid. We labeled HeLa cells with [2,3-³H]glutamate and used a carboxylate-specific monoclonal antibody to enrich for the desired proteins. PNGaseF digestion of these proteins released labeled N-glycans with a free amino group and 1–3 carboxylates. Mild acid hydrolysis had no effect, but strong acid hydrolysis of the glycans released >80 % of the ³H as glutamate. Reducing the carboxylates to alcohols prior to hydrolysis eliminated the [³H]glutamate and generated [³H]4-amino 5-hydroxy pentanoic acid, suggesting that [³H]glutamate was linked to the glycan through its γ-carboxyl. The glutamate-containing N-glycans resisted exoglycosidase digestion and oligosaccharide processing inhibitors greatly reduced [³H]glutamate incorporation. These results demonstrate that mammalian cells synthesize complex-type N-glycans with glutamate linked to their antennae, further expanding their potential for covalent or ionic interactions.