N-Linked Glycosylation of the Liver Cancer Biomarker GP73
N-Linked Glycosylation of the Liver Cancer Biomarker GP73
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DOI：
10.1002/jcb.21610
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发表时间：
2008-05-01
影响因子：
4
通讯作者：
Block, Timothy M.
中科院分区：
文献类型：
--
作者：
Norton, Pamela A.;Comunale, Mary Ann;Block, Timothy M.
GP 73（特别是岩藻糖基化GP 73）循环水平升高与肝细胞癌之间的相关性表明，有必要对GP 73糖基化程度进行全面分析。对这种低丰度蛋白质的糖基化模式的详细分析受到技术困难的影响。使用常规的凝集素亲和层析，我们已经确定从HCC来源的细胞系分泌的四分之三的GP 73是岩藻糖基化的。使用质谱，我们已经建立了至少两个三个潜在的N-连接的糖基化位点被占领的GP 73从培养的肝癌细胞分泌的大多数分子。此外，添加到重组GP 73中的寡糖类似于大量分泌蛋白中存在的寡糖，主要是具有核心岩藻糖的双触角，具有较小部分的三触角和四触角结构。在重组蛋白上观察到的岩藻糖基化频率与内源性分泌蛋白的凝集素结合模式一致。最后，我们已经开发了一种方法来询问添加到近全长蛋白质或特定序列子的聚糖，提供了嵌入异源环境中的小肽可以保留岩藻糖基化和添加的寡糖的高水平分支的概念证明。J.细胞。104：136-149，2008. (C)威利-利斯公司
The association between elevated circulating levels of GP73 (and fucosylated GP73 in particular) and hepatocellular carcinoma suggests that a thorough analysis of the extent of GP73 glycosylation is warranted. Detailed analysis of the glycosylation patterns of such low abundance proteins are hapered by technical difficulties. Using Conventional lectin affinity chromatography, we have established that three quarters of the GP73 secreted from a cell line derived from HCC is fucosylated. Using mass spectrometry, we have established that at least two of three potential sites of N-linked glycosylation are occupied on most molecules of GP73 secreted from cultured hepatoma cells. Furthermore, the oligosaccharides added to recombinant GP73 resemble those present in the bulk of secreted protein, mostly bi-antennary with core fucose, with a smaller fraction of tri- and tetra-antennary structures. The frequency of fucosylation observed on the recombinant protein agrees well with the pattern of lectin binding of the endogenous secreted protein. Finally, we have developed a method to interrogate the glycans added to either the near full length protein or at a particular sequon, providing proof of concept that a small peptide embedded in a heterologous context can preserve both fucosylation and a high level of branching of oligosaccharides added. J. Cell. Biochem. 104: 136-149, 2008. (C) Wiley-Liss, Inc.