Thermally responsive reversed micelles for immobilization of enzymes
Thermally responsive reversed micelles for immobilization of enzymes
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DOI：
10.1002/adfm.200600452
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发表时间：
2008-01-11
影响因子：
19
通讯作者：
Yang, Yi-Yan
中科院分区：
文献类型：
--
作者：
Chen, Hong;Liu, Li-Hong;Yang, Yi-Yan
本研究合成了具有热响应性的烷基封端的聚（N-异丙基丙烯酰胺-co-丙烯酸）两亲性共聚物，并成功地用于制备反胶束和酶催化剂。透射电子显微镜（TEM）研究表明，反胶团是球形的性质。并以裸酶为对照，研究了反胶束固定化脂肪酶催化月桂酸和正丙醇酯化反应的活性。固定化脂肪酶具有更高的催化活性。此外，还考察了pH、含水量、聚合物浓度和酶浓度对催化活性的影响。优化的负载脂肪酶的反胶团的制备条件为：聚合物浓度12 mg mL（-1），酶浓度25 mg mL（-1），磷酸盐缓冲液（PBS），pH 7.4，W-0（[水]/[表面活性剂]）83.3。固定化脂肪酶在胶束中的稳定性明显优于传统的琥珀酸双（2-乙基己基）酯磺酸钠胶束。更重要的是，脂肪酶固定化胶束的尺寸减小，并且当温度升高到略高于聚合物的低临界溶解温度（LCST）的值时，酶溶液从反应混合物中沉淀。酶的回收率在75%左右。回收的脂肪酶的α-螺旋结构保持完整。此外，在将环境温度升高至略高于LCST后终止酶促反应。这些热响应胶束可能成为一种有前途的酶固定化系统。
In this study, thermally responsive alkyl end-capped poly(N-isopropylacrylamide-co-acrylic acid) amphiphilic copolymers were synthesized, and successfully utilized to fabricate reversed micelles and immobilize enzymes. Transmission electron microscopy (TEM) study showed that the reversed micelles were spherical in nature. The activity of Candida rugosa lipase immobilized in the reversed micelles towards catalyzing the esterification of lauric acid and 1-propanol was analyzed in comparison with naked enzyme. Immobilized lipase provided much greater catalytic activity. In addition, the effects of pH, water content, polymer and enzyme concentration on the catalytic activity were investigated. The optimized fabrication conditions of lipase-loaded reversed micelles, under which lipase gave the highest activity, were as follows: polymer concentration, 12 mg mL(-1); enzyme concentration, 25 mg mL(-1) phosphate buffered saline (PBS); pH, 7.4; W-0 ([water]/[surfactant]), 83.3. Lipase immobilized in these micelles was much more stable than that in conventional sodium bis(2-ethylhexyl) sulfosuccinate micelles. More importantly, the size of lipase-immobilized micelles decreased, and the enzyme solution precipitated from the reaction mixture when the temperature increased to a value slightly higher than the lower critical solution temperature (LCST) of the polymer. The recovery rate of enzyme was about 75%. The alpha-helix structure of the recovered lipase remained intact. In addition, the enzymatic reaction was terminated after raising the environmental temperature slightly above the LCST. These thermally responsive micelles may make a promising system for enzyme immobilization.