毛细管电泳法表征多肽及糖蛋白的稳定性

建立了毛细管电泳表征多肽和糖蛋白稳定性的方法。分别以血管紧张素II(Ang II)和植物血球凝集素(PHA)、牛凝血酶(B-Thr)、人凝血酶(H-Thr)、辣根过氧化物酶(HRP)4种糖蛋白为多肽和糖蛋白的模式分子。从样品浓度、电泳缓冲液、样品溶液pH和离子强度等方面优化了血管紧张素II的分离分析条件;从毛细管的选择、样品的电荷状态、电泳缓冲液的选择和分离电压的影响等方面讨论了糖蛋白的分离条件。Ang II和4种糖蛋白的稳定性试验结果表明:Ang II可在pH 7.4的硼酸盐缓冲液(0.02 mol/L)中于4℃下稳定放置48 h; 4种糖蛋白可在pH 7.4硼酸盐缓冲液(0.2 mol/L)中于20,4,-20℃下稳定放置48 h;放置时间大于一周且小于四周时,在-20℃下各蛋白质均保持稳定;放置时间大于两周且小于四周时,只有HRP在上述3个温度下均保持稳定。该方法具有高效、快速、简单、低成本的特点,可广泛应用于多肽和蛋白质的稳定性表征。

Characterization of stability of polypeptides and glycoproteins by capillary electrophoresis

A capillary electrophoresis (CE) method was developed for the stability characterization of polypeptides and glycoproteins. Angiotensin II (Ang II), phytagglutinin (PHA), bovine thrombin (B-Thr), human thrombin (H-Thr) and horseradish peroxidase (HRP) were used as polypeptide and glycoprotein mode molecules. The parameters affecting the analysis efficiency for Ang II, such as sample concentration, running buffer, pH and ionic strength of sample solution were optimized, as for the glycoprotein, capillary conditions, charge state of sample, running buffer and applied voltage were optimized. It showed that the Ang II was stable when kept in borate buffer (0.02 mol/L pH 7.4) at 4℃ for 48 h. The four glycoproteins were quite stable in borate buffer (0.2 mol/L pH 7.4) at 20, 4, -20℃ for 48 h, and also kept stable at -20℃ when deposited over one week and less than four weeks. HRP was the only one that kept stable when deposited over two weeks and less than four weeks. This method is effective, rapid, simple and low-cost and can be widely used for the stability characterization of polypeptides and glycoproteins.