Enzyme specific activity in functionalized nanoporous supports
Enzyme specific activity in functionalized nanoporous supports
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DOI：
10.1088/0957-4484/19/12/125102
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发表时间：
2008-03-26
期刊：
影响因子：
3.5
通讯作者：
Ackerman, Eric J.
中科院分区：
文献类型：
--
作者：
Lei, Chenghong;Soares, Thereza A.;Ackerman, Eric J.
本研究发现，通过改变功能化纳米孔支架中的蛋白质负载密度（P(LD)），可以大幅提高酶的比活性，从而使酶的固定化效率（I(e)，即固定化酶的比活性与溶液中游离酶的比活性之比）大大高于100%。净带负电荷的葡萄糖氧化酶（GOX）和净带正电荷的有机磷水解酶（OPH）分别自发地包裹在NH(2)-和hooc功能化的介孔二氧化硅（300埃，FMS）中。FMS包埋的GOX比活性随着PLD的降低而升高。随着PLD的降低，FMS中GOX的Ie从< 35%增加到bb10150%。与GOX不同，HOOC-FMS中OPH的比活性随着PLD的增加而增加。随着PLD的增加，FMS中OPH的对应Ie从100%增加到bb0 - 200%。基于蛋白质结构的蛋白质表面电荷分析指导了FMS中蛋白质分子基于静电相互作用的取向，表明FMS中底物对GOX分子的访问在高PLD时受到限制，从而降低了GOX的比活性。相比之下，FMS中OPH分子的底物通路在高PLD下保持开放，并可能促进更有利的约束环境，从而增强OPH活性。
Here we reveal that enzyme specific activity can be increased substantially by changing the protein loading density (P(LD)) in functionalized nanoporous supports so that the enzyme immobilization efficiency (I(e), defined as the ratio of the specific activity of the immobilized enzyme to the specific activity of the free enzyme in solution) can be much higher than 100%. A net negatively charged glucose oxidase ( GOX) and a net positively charged organophosphorus hydrolase ( OPH) were entrapped spontaneously in NH(2)- and HOOC-functionalized mesoporous silica ( 300 angstrom, FMS) respectively. The specific activity of GOX entrapped in FMS increased with decreasing PLD. With decreasing PLD, Ie of GOX in FMS increased from < 35% to > 150%. Unlike GOX, OPH in HOOC-FMS showed increased specific activity with increasing PLD. With increasing PLD, the corresponding Ie of OPH in FMS increased from 100% to > 200%. A protein structure-based analysis of the protein surface charges directing the electrostatic interaction-based orientation of the protein molecules in FMS demonstrates that substrate access to GOX molecules in FMS is limited at high PLD, consequently lowering the GOX specific activity. In contrast, substrate access to OPH molecules in FMS remains open at high PLD and may promote a more favorable confinement environment that enhances the OPH activity.