磁响应交联糖化酶聚集体的制备及催化特性

发布时间：2018-08-07 21:34

【摘要】：提出了一种采用羧基磁性纳米粒子制备杂化磁响应交联酶聚集体(M-CLEAs)的方法。表面羧基修饰的约10 nm的磁性纳米粒子与酶分子表面的氨基位点通过静电相互作用,形成复合物,在磁场作用下可将磁性纳米粒子-酶复合物从溶液中分离,经戊二醛交联即形成M-CLEAs。传统的表面氨基修饰的磁性纳米粒子与酶需在沉淀剂作用下,从溶液中分离,而后采用戊二醛共交联,而本方法无须沉淀剂,过程更为简化。以糖化酶为对象,对该过程的影响因素(交联时间、pH、酶浓度、戊二醛浓度等条件)进行了探索,并对制得的M-CLEAs的酶学性质进行了较为详细考察。结果表明,最优制备条件为:酶浓度1 mg·ml-1,磁流体浓度10 mg·ml-1,戊二醛浓度0.25%(质量体积比),在pH 6.0下交联反应6 h,最终载酶量可达80 mg·g~(-1)、比活为50 U·mg~(-1)。制得的固定化酶pH稳定性、热稳定性和储存稳定性均显著改善,可实现糖化酶重复使用10次,仍保留接近60%的酶活。
[Abstract]:A method for preparing hybrid magnetic response crosslinking enzyme aggregates (M-CLEAs) using carboxyl magnetic nanoparticles was proposed. The magnetic nanoparticles modified by carboxyl group on the surface interact with the amino sites on the surface of enzyme molecules by electrostatic interaction to form a complex, which can separate the magnetic nanoparticles and enzyme complexes from the solution under the action of magnetic field. M-CLEAs were formed by crosslinking with glutaraldehyde. The traditional surface modified magnetic nanoparticles and enzymes need to be separated from solution under the action of precipitator and then cross-linked by glutaraldehyde. The influencing factors of Glucoamylase (such as crosslinking time pH, enzyme concentration, glutaraldehyde concentration, etc.) were explored and the enzymatic properties of the prepared M-CLEAs were investigated in detail. The results showed that the optimal preparation conditions were as follows: enzyme concentration 1 mg ml -1, magnetic fluid concentration 10 mg ml -1, glutaraldehyde concentration 0.25% (mass / volume ratio), crosslinking reaction at pH 6.0 for 6 h, the final enzyme loading capacity was 80 mg g ~ (-1) and the specific activity was 50 U mg ~ (-1). The pH stability, thermal stability and storage stability of the immobilized enzyme were improved significantly. The enzyme activity could be reused for 10 times, and the enzyme activity was about 60%.
【作者单位】：华侨大学化工学院生物工程与技术系;福建省生物化工技术重点实验室(华侨大学);华侨大学分析测试中心;
【基金】：国家自然科学基金面上基金项目(21576108) 福建省新世纪优秀人才项目华侨大学研究生科研创新能力培育计划资助项目~~
【分类号】：Q814.2;TB383.1

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