Separation of Glycoproteins Based on Sugar Chains Using Novel Stationary Phases Modified with Poly(ethylene glycol)-Conjugated Boronic-Acid Derivatives
Separation of Glycoproteins Based on Sugar Chains Using Novel Stationary Phases Modified with Poly(ethylene glycol)-Conjugated Boronic-Acid Derivatives
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DOI：
10.1021/acs.analchem.2c01002
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发表时间：
2022-05-10
影响因子：
7.4
通讯作者：
Otsuka，Koji
中科院分区：
文献类型：
--
作者：
Kobayashi，Hiroshi;Masuda，Yusuke;Otsuka，Koji
硼酸(BA)与二元醇结构形成可逆络合物。BA衍生物根据糖链的不同来分离糖蛋白。分离通常发生在碱性条件下，这不能保证糖蛋白的结构稳定性。本文以聚乙二醇(PEG)为连接剂，聚乙二醇亚胺(PEI)为骨架，制备了5-硼酸(BPA)硅胶柱，以抑制非选择性吸附，提高BPA的部分密度。仅使用不含有机溶剂的水缓冲液的高效液相色谱(HPLC)证明了BPA柱对糖蛋白的选择性保留能力。双酚A与糖链上的二元醇相互作用，但不与蛋白质相互作用。在对N-葡聚糖的评估中，BPA柱对高甘露糖型和低亲和力的乙酰神经氨酸(Neu5Ac)显示出更高的保留能力。最后，由于非还原末端有Neu5Ac的存在，一对糖蛋白，胎球蛋白和积雪草素被选择性地分离出来。
Boronic acid (BA) reversibly complexes with the diol structure. BA derivatives separate glycoproteins based on the differences in the sugar chains. Separation typically occurs under basic conditions, which does not guarantee the structural stability of glycoproteins. Here, 5-boronopicolinic acid (BPA) is used to prepare silica-gel based columns with poly(ethylene glycol) (PEG) as a linker to suppress nonselective adsorption and poly(ethylene imine) (PEI) as a scaffold to increase the BPA moiety density. High-performance liquid chromatography (HPLC) using only aqueous buffer solutions without organic solvents demonstrates the selective retention ability of the BPA columns for glycoproteins. BPA interacts with the diols in the sugar chains but not the proteins. In an evaluation for N-glycans, the BPA columns show a higher retention ability toward high mannose type and a lower affinity toN-acetylneuraminic acid (Neu5Ac). Finally, a pair of glycoproteins, fetuin and asialofetuin, are selectively separated due to the presence of Neu5Ac on the nonreducing end.