Hydrophilization of immobilized model enzymes suggests a widely applicable method for enhancing protein stability in polar organic co-solvents
Hydrophilization of immobilized model enzymes suggests a widely applicable method for enhancing protein stability in polar organic co-solvents
复制标题
DOI：
10.1016/j.molcatb.2007.02.005
复制
发表时间：
2007-05-02
影响因子：
--
通讯作者：
Brena, Beatriz M.
中科院分区：
文献类型：
--
作者：
Irazoqui, Gabriela;Giacomini, Cecilia;Brena, Beatriz M.
使用来自大肠杆菌、乳酸克鲁维酵母和曲霉的β-半乳糖苷酶来表征两步策略的酶稳定化的潜力：（i）固定在戊二醛-琼脂糖（Glut 90）上，（ii）随后通过与聚葡聚糖-葡聚糖聚合物（Glut 90-Pal）反应产生亲水性纳米环境，然后与聚胺-葡聚糖聚合物（Glut 90-Pal-Pam）反应。通过动力学参数、共溶剂（乙醇和丙酮）和温度稳定性对衍生物进行了表征。在所有情况下，亲水化实现了重要的共溶剂稳定化。获得的最显著的结果之一是A的半衰期增加了25倍。将Glut 90-Pal-Pam衍生物溶于50%（v/v）丙酮中。在非常剧烈的共溶剂浓度下实现的稳定化与纳米环境的稳定化直接相关。K-M值表明，亲水性壳似乎表现为开放结构，并可能产生保护酶不变性的“分配效应”。这些结果表明，固定化酶的稳定性，这将使工业应用的发展成为可能。(c)2007 Elsevier B. V.保留所有权利。
beta-Galactosidases from Escherichia coli, Kluyveromyces lactis and Aspergillus oryzae were used to characterize the potential for enzyme stabilization of a two-step strategy: (i) immobilization on glutaraldehyde-agarose (Glut90), (ii) subsequent generation of a hydrophilic nano-environment by reaction with polyaldehyde-dextran polymer (Glut90-Pal), followed by polyamine-dextran polymer (Glut90-Pal-Pam). The derivatives were characterized by kinetics parameters, co-solvent (ethanol and acetone) and temperature stability. Hydrophilization achieved important co-solvent stabilization in all cases. One of the most remarkable results obtained was a 25-fold increase in the half-life of the A. oryzae Glut90-Pal-Pam derivative in 50% (v/v) acetone. Stabilization achieved in very drastic co-solvent concentrations is directly related to the hydrophilization of the nano-environment. The K-M values show that the hydrophilic shell appears to behave as an open structure and may create a "partition effect" that protects the enzymes from denaturation. These results show the potential of hydrophilization for building up additional stabilization of immobilized enzymes which would make possible the development of industrial applications. (c) 2007 Elsevier B.V. All rights reserved.