Glycoproteomics of Trypanosoma cruzi trypomastigotes using subcellular fractionation, lectin affinity, and stable isotope labeling
Glycoproteomics of Trypanosoma cruzi trypomastigotes using subcellular fractionation, lectin affinity, and stable isotope labeling
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DOI：
10.1021/pr060364b
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发表时间：
2006-12-01
影响因子：
4.4
通讯作者：
Orlando, Ron
中科院分区：
文献类型：
--
作者：
Atwood, James A., III;Minning, Todd;Orlando, Ron
在这里，我们详细介绍了第一个人类病原体的糖蛋白质组学分析。我们描述了一种方法，使细胞器和细胞表面的N-连接的糖蛋白从锥虫克氏锥虫，南美锥虫病的病原体的鉴定。该方法基于亚细胞分级分离方案，以产生富含细胞器或质膜/细胞质蛋白的组分。通过凝集素亲和捕获的糖肽从每个亚细胞级分和稳定的同位素标记的聚糖附着位点（H2O）-O-18，我们明确地确定了36个糖基化位点的35糖肽映射到29个糖蛋白。我们还首次提出了11 T的表达证据。cruzi特异性糖蛋白的研究，并提供了表明粘蛋白相关表面蛋白家族（MASP）和分散基因家族（DGF-1）被N-连接聚糖后修饰的实验数据。
Herein we detail the first glycoproteomic analysis of a human pathogen. We describe an approach that enables the identification of organelle and cell surface N-linked glycoproteins from Trypanosoma cruzi, the causative agent of Chagas' disease. This approach is based on a subcellular fractionation protocol to produce fractions enriched in either organelle or plasma membrane/cytoplasmic proteins. Through lectin affinity capture of the glycopeptides from each subcellular fraction and stable isotope labeling of the glycan attachment sites with (H2O)-O-18, we unambiguously identified 36 glycosylation sites on 35 glycopeptides which mapped to 29 glycoproteins. We also present the first expression evidence for 11 T. cruzi specific glycoproteins and provide experimental data indicating that the mucin associated surface protein family (MASP) and dispersed gene family (DGF-1) are post-translationally modified by N-linked glycans.