College of Chemistry, Jilin University, Changchun 130012, China.
Analyst. 2013 May 21;138(10):3032-7. doi: 10.1039/c3an00107e.
In this paper, a novel method has been proposed to achieve selective enrichment and purification of glycoproteins/glycopeptides on a surface patterned sample support, which consists of a hydrophobic outer layer (F-SAM) and an internal boronic acid-modified gold microspot (900 μm). Upon deposition, the sample solution is firstly concentrated in a small area by repulsion of the hydrophobic outer layer, and then the glycoproteins/glycopeptides are selectively captured through boronic acid covalently binding in the inner layer. However, the non-glycosylated proteins/peptides or high concentration salts are removed after rinsing with alkaline solution. As a result, the detection sensitivity is improved by an order of magnitude greater than when using a stainless steel MALDI plate. With surface patterned sample support, the glycoproteins/glycopeptides can be detected even under interference from the excessive existing non-glycosylated proteins/peptides (10 times more than glycoproteins/glycopeptides). Simultaneously, high-quality mass spectra can be obtained even in the presence of urea (1 M), NaCl (1 M), or NH4HCO3 (200 mM). Therefore, this novel technique may be applied to high-throughput analysis of low-abundance glycoproteins/glycopeptides in complicated proteome research.
本文提出了一种新的方法，可在表面图案化的样品支撑物上实现糖蛋白/糖肽的选择性富集和纯化，该支撑物由疏水性外层（F-SAM）和内部硼酸修饰的金微点（900 μm）组成。沉积后，样品溶液首先通过疏水性外层的排斥作用在小区域内浓缩，然后通过内层的硼酸共价键合选择性捕获糖蛋白/糖肽。然而，用碱性溶液冲洗后，会去除非糖基化的蛋白质/肽或高浓度盐。结果，检测灵敏度比使用不锈钢 MALDI 板提高了一个数量级。使用表面图案化的样品支撑物，即使在存在过量的现有非糖基化蛋白质/肽（比糖蛋白/糖肽多 10 倍）的情况下，也可以检测到糖蛋白/糖肽。同时，即使存在尿素（1 M）、NaCl（1 M）或 NH4HCO3（200 mM），也可以获得高质量的质谱。因此，这项新技术可能适用于复杂蛋白质组研究中低丰度糖蛋白/糖肽的高通量分析。
