糊精-尿酸酶结合物的制备、表征及酶学性质。
Dextrin-uricase conjugate: Preparation, characterization, and enzymatic properties.
机构信息
State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang 330047, China; School of Food Science and Technology, Nanchang University, Nanchang 330047, China.
School of Food Science and Technology, Nanchang University, Nanchang 330047, China.
出版信息
Int J Biol Macromol. 2018 May;111:28-32. doi: 10.1016/j.ijbiomac.2017.12.131. Epub 2017 Dec 29.
Uricase was conjugated with dextrin to improve its stability. Firstly, dextrin was succinylated into dextrin monosuccinate with the functional group, carboxylic acid, and dextrin monosuccinates with different degree of substitution (DS) were prepared. Secondly, dextrin-uricase conjugate was synthesized by uricase and dextrin monosuccinate, which was verified by size-exclusion chromatography and anion exchange chromatography. Thirdly, it was found that the conjugate degree of the dextrin-uricase conjugate was positively related to the DS of dextrin monosuccinate, but the activity was seriously lost and difficult to recover by α-amylase. When the molar ratio of dextrin monosuccinate (DS=0.283) to uricase was 30:1, the conjugate degree of dextrin-uricase conjugate reached 22.1% and the activity was decreased to 40.4%. After triggered by α-amylase, the activity was recovered to 83.4%. Finally, the enzymatic properties of this dextrin-uricase conjugate were investigated and compared with those of free uricase. The optimal pH and temperature of the dextrin-uricase conjugate was 9.0 and 45°C, respectively, whereas the optimal pH and temperature of free uricase was 8.5 and 45°C, respectively. Furthermore, dextrin-uricase conjugate was more resistant to simulated physiological conditions and trypsin. These results suggested that the stability of uricase could be ameliorated by conjugation with dextrin.
尿酸酶与糊精偶联以提高其稳定性。首先，糊精通过功能基团羧酸酐被琥珀酸化，制备具有不同取代度（DS）的糊精单琥珀酸酯。其次，通过尿酸酶和糊精单琥珀酸酯合成了糊精-尿酸酶偶联物，通过尺寸排阻色谱和阴离子交换色谱进行了验证。第三，发现糊精-尿酸酶偶联物的偶联度与糊精单琥珀酸酯的 DS 呈正相关，但活性严重丧失且难以通过α-淀粉酶恢复。当糊精单琥珀酸酯（DS=0.283）与尿酸酶的摩尔比为 30:1 时，糊精-尿酸酶偶联物的偶联度达到 22.1%，活性降低至 40.4%。经α-淀粉酶触发后，活性恢复至 83.4%。最后，研究了该糊精-尿酸酶偶联物的酶学性质，并与游离尿酸酶进行了比较。糊精-尿酸酶偶联物的最适 pH 和温度分别为 9.0 和 45°C，而游离尿酸酶的最适 pH 和温度分别为 8.5 和 45°C。此外，糊精-尿酸酶偶联物在模拟生理条件和胰蛋白酶下更稳定。这些结果表明，通过与糊精偶联可以改善尿酸酶的稳定性。
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