A general approach for characterizing glycosylation sites of glycoproteins.
A general approach for characterizing glycosylation sites of glycoproteins.
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表征糖蛋白糖基化位点的通用方法。
DOI：
10.1016/0003-2697(91)90419-t
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发表时间：
1991
影响因子：
2.9
通讯作者：
P. Yuan
中科院分区：
文献类型：
--
作者：
K. Hsi;L. Chen;D. Hawke;L. Zieske;P. Yuan
糖生物学的最新进展极大地刺激了碳水化合物的研究;然而，改进用于在蛋白质结构分析中鉴定和分离特定糖基化位点的技术仍然是一个挑战。在这里，我们报告了一个实用的方法，利用膜染色技术的Problott，PVDF型膜，筛选糖蛋白和糖肽来自糖蛋白的酶促反应。为了提高检测灵敏度，采用了使用生物素化凝集素、抗生物素蛋白和生物素化过氧化物酶的扩增染色技术。此外，我们描述了一种微批量亲和结合程序，以分离糖肽从复杂的糖蛋白酶促反应。这些协议使我们能够开始与subnanomole量的糖蛋白和定位的糖基化位点，分离糖肽在一个均匀的形式，并进行氨基酸组成，氨基酸序列和质量分析分离的糖肽。对一种结构尚不清楚的糖蛋白羧肽酶P的糖基化位点进行了研究。
Recent advances in glycobiology have greatly stimulated carbohydrate research; however, improving techniques for identification and isolation of specific glycosylation sites in protein structure analysis remains a challenge. We report here a practical approach utilizing a membrane staining technique on Problott, a PVDF-type membrane, to screen glycoproteins and glycopeptides derived from enzymatic digests of glycoproteins. To improve the detection sensitivity, an amplifled staining technique using biotinylated lectins, avidin, and biotinylated peroxidase was employed. In addition, we describe a micro-batch affinity binding procedure to isolate glycopeptides from complex glycoprotein enzymatic digests. These protocols allow us to start with a subnanomole quantity of glycoprotein and locate the glycosylation sites; isolate glycopeptides in a homogeneous form; and perform amino acid composition, amino acid sequence, and mass analyses on the isolated glycopeptides. The characterization of glycosylation site of a model glycoprotein, carboxypeptidase P, of which the structure is still largely unknown, has been investigated.
酿酒酵母 mnn9 甘露糖蛋白的碳水化合物结构。
DOI：
--
发表时间：
1984
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Tsai,PK;Frevert,J;Ballou,CE
通讯作者：
Ballou,CE