摘要
以温敏性材料N-羟基琥珀酰亚胺活化的聚(N-异丙基丙烯酰胺)为载体制备固定化木瓜蛋白酶,对其酶学性质进行了研究。结果表明:固定化酶的最适pH值为7.5,最适温度为65℃,米氏常数为2.72 mg/mL;经过24次温敏循环操作,剩余相对活力86.6%;在4℃条件下放置60 d,剩余相对活力为71.5%。与游离酶相比,固定化酶pH值稳定性、热稳定性、操作稳定性和储存稳定性都有显著提高。将此固定化酶应用于HCG单克隆抗体消化,与纤维素固定化酶相比,水解能力更强,更适合于大分子的酶解,特别是抗体片段的制备。
Traditional solid phase immobilized enzyme shows low catalytic efficiency,especially when acting on macromolecular substrate. Here the thermally sensitive material poly （N-isopmpylacrylamide）, which was esterified by N-hydroxysuccinimide, was employed as a carrier for the immobilization of papain, and some properties of the immobilized papain were investigated. From the study it was found that the optimal pH value and temperature for immobilized papain activity were 7.5 and 65℃ respectively, and the Michaelis constant was 2.72 mg/mL. The immobilized papain left was 86.6% of the activity after 24 thermal cycles. After stored at 4 ℃ for 60 d, the immobilized papain activity retained 71.5 %. Compared with free enzyme, the pH value, thermal, operational and storage stabilities of immobilized papain were obviously improved. The immobilized papain was applied to the digestion of HCG mono- clonal antibody. Compared with the papain immobilized on cellulose, the hydrolysis capacity of the thermally sensitive immobilized papain was higher and the operation was more convenient. It was more suitable for the enzymatic hydrolysis of the macromolecules, especially the preparation of antibody fragments.
出处
《药物生物技术》
CAS
2013年第3期210-214,共5页
Pharmaceutical Biotechnology
基金
浙江省科技计划项目(No.2012C23076)
