固定化粪产碱杆菌青霉素G酰化酶的制备

Preparation on immobilization of penicillin G acylase from Alcaligenes faecalis

粪产碱杆菌来源的青霉素G酰化酶通过共价结合在环氧型载体Eupergit C上,通过对酶质量浓度、固定化反应时间、pH值以及反应温度等条件的考察,确定了最优固定化条件:375mg比活力5 000U/g的重组粪产碱杆菌青霉素G酰化酶蛋白对应1g载体,最适pH 8.0,反应温度30℃。反应120h后制得固定化青霉素G酰化酶活力达到220U/g,固定化酶效率为10.7%。该固定化酶可在含饱和乙酸丁酯磷酸缓冲液中彻底水解青霉素G钾盐。经过15批连续水解反应,固定化酶仍保持95%的活力,展现出良好的稳定性。

The penicillin G acylase from Alcaligenes faecalis was covalently immobilized to epoxy-type carrier Eurpergit C.The immobilization conditions including the ratio of the enzyme to the support,reaction time,pH and temperature were studied.The optimized immobilization conditions are 375 mg recombinantAlcaligenes faecalis penicillin G acylase of 5 000 U/g specific activi-ty to per gram Eupergit C,pH 8.0 and 30 ℃.After stirring for 120 h,the activity of immobilized penicillin G acylase is up to 220 U/g and the efficiency is 10.7%.The immobilized enzyme can completely hydrolysis penicillin G potassium salt in saturated butyl-acetate phosphate buffer.After hydrolyzing penicillin G potassium salt 15 times in a reactor,the immobilized enzyme still remains 95% activity and it shows a good operational stability.