优化稳定剂提高丙酮酸氧化酶稳定性的研究  

Enhancement of pyruvate oxidase stability by chemical stabilizers

作　　者：刘冰 丛文杰 姚兵莉 周化岚[1] 张建国[1] LIU Bing;CONG Wenjie;YAO Bingli;ZHOU Hualan;ZHANG Jianguo(Institute of Food Science and Engineering,School of Health Science and Engineering,University of Shanghai for Science and Technology,Shanghai 200093,China)

机构地区：[1]上海理工大学健康科学与工程学院食品科学与工程研究所,上海200093

出　　处：《生物加工过程》2023年第6期616-622,共7页Chinese Journal of Bioprocess Engineering

基　　金：国家自然科学基金(31870045)。

摘　　要：来自绿色气球菌的丙酮酸氧化酶被广泛用于食品工业、医药工业及生物检测等方面,但其多聚体结构会导致酶稳定性低,因此提高丙酮酸氧化酶的稳定性有助于丙酮酸氧化酶的推广和应用。首先筛选了醇类、糖类和离子稳定剂,进而优化了稳定剂复配组成。结果表明:在4℃、pH 6.0的条件下,含有0.01 mmol/L CuSO4、30 g/L甘油和90 g/L PEG2000的复合稳定剂能够将丙酮酸氧化酶活性提升为对照组的1.5倍。丙酮酸氧化酶稳定性的提高归因于丙酮酸氧化酶亚基之间连接的强化,它们之间形成了更多有活性的四聚体。最后,添加复合稳定剂的丙酮酸氧化酶在-20℃下储存28 d后仍能保持46.7%的酶活。可见,复合稳定剂能显著提高丙酮酸氧化酶的稳定性,有利于丙酮酸氧化酶的更广泛应用。Aerococcu sviridans pyruvate oxidase was widely utilized in various analysis and diagnosis in food and medical industry,but it suffered from low stability because of its structure formed with multimeric subunits.To prolong the shelf life of pyruvate oxidase,chemical stabilizers were usually added into pyruvate oxidase solution to improve its stability.Multiple potential stabilizers,including alcohols,sugars,ions,and various combinations,were selected in our screening experiments.The optimized conditions,with a mixture of 0.01 mmol/L copper sulfate,30 g/L glycerol and 90 g/L PEG2000,could significantly improve the stability of pyruvate oxidase,and the final pyruvate oxidase activity was increased to 1.5 times at 4℃,pH 6.0.Such improvement of pyruvate oxidase stability was attributed to the enhanced interactions of subunits in the tetramer structure of pyruvate oxidase.Notably,pyruvate oxidase with stabilizers could maintain 46.7%of enzymatic activity after it was stored at-20℃for 28 days.Our research provided a general guidance how to improve the stability of pyruvate oxidase,which could benefit the application of pyruvate oxidase.

关 键 词：丙酮酸氧化酶 酶稳定性 稳定剂 多聚酶 优化 

分 类 号：TQ464[化学工程—制药化工] Q804[生物学]