Encapsulation of enzymes in liposomes: High encapsulation efficiency and control of substrate permeability
Encapsulation of enzymes in liposomes: High encapsulation efficiency and control of substrate permeability
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DOI：
10.1081/bio-120028669
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发表时间：
2004-01-01
期刊：
影响因子：
--
通讯作者：
Fournier, D
中科院分区：
文献类型：
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作者：
Chaize, B;Colletier, JP;Fournier, D
将酶封装到脂质体中是一种稳定并防止其变性和蛋白水解的有前途的技术。我们使用乙酰胆碱酯酶证明了这一点，乙酰胆碱酯酶是农药的主要目标。为了达到合理的封装良率，我们分析了每个步骤或各种封装程序所涉及的参数。唯一不会使蛋白质变性的封装方法是脂膜水合技术，但封装效率通常较低。通过诱导酶和脂质表面之间的特定相互作用，效率可提高至 40% 以上。一旦被封装，酶就遇到了另一个问题：脂膜的渗透性屏障通过降低底物分子的进入速率，从而降低脂质体内的底物浓度，从而极大地降低了脂质体中包埋的酶的活性。为了解决这个问题，我们通过重构大肠杆菌的孔蛋白来控制脂质体壁的渗透性。我们成功地恢复了酶的全部功能，同时保留了对变性酶和蛋白水解酶的保护。
Enzyme encapsulation into liposomes is a promising technique to stabilize and prevent them from denaturation and proteolysis. We demonstrate this using acetylcholinesterase which is the main target for pesticides. In order to achieve a reasonable encapsulation yield, we analyzed the parameters involved in each step or various encapsulation procedures. The only encapsulation method which did not denature the protein was the lipid film hydration technique, however the encapsulation efficiency was usually low. The efficiency could be increased up to more than 40% by induction of a specific interaction between the enzyme and the lipid surface. Once encapsulated, the enzyme encountered another problem: the permeability barrier of the lipid membrane drastically diminished the activity of the enzyme entrapped in the liposome by reducing the entrance rate of the substrate molecules and then reducing the substrate concentration inside the liposome. To solve this problem, we controlled the permeability of the liposome wall by reconstituting a porin from Eycherichia coli. We succeeded to recover the full functionality of the enzyme, while retaining the protection against denaturation and proteolytic enzymes.