Suma Yanasinee, Kang Christina S, Kim Han S
Department of Advanced Technology Fusion, Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul, 143-701, Korea.
Department of Environmental Engineering, Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul, 143-701, Korea.
Environ Sci Pollut Res Int. 2016 Jan;23(2):1015-24. doi: 10.1007/s11356-015-4168-5. Epub 2015 Feb 7.
The decomposition of various aromatic hydrocarbon intermediates was examined using a recombinant oxidative enzyme immobilized on single-walled carbon nanotubes (SWCNTs). Hydroxyquinol 1,2-dioxygenase (CphA-I), which catalyzes ring cleavage of catechol and its analogues, was obtained from Arthrobacter chlorophenolicus A6 via cloning, overexpression, and subsequent purification. This recombinant enzyme was immobilized on SWCNTs by physical adsorption and covalent coupling in the absence and presence of N-hydroxysuccinimide. The immobilization yield was as high as 52.1%, and a high level of enzyme activity of up to 64.7% was preserved after immobilization. Kinetic analysis showed that the substrate utilization rates (vmax) and catalytic efficiencies (kcat/KM) of the immobilized enzyme for all substrates evaluated were similar to those of the free enzyme, indicating minimal loss of enzyme activity during immobilization. The immobilized enzyme was more stable toward extreme pH, temperature, and ionic strength conditions than the free enzyme. Thus, the oxidative enzyme immobilized on SWCNTs can be used as an effective and stable biocatalyst for the biochemical remediation process if further investigations would be carried out under field conditions.
使用固定在单壁碳纳米管（SWCNTs）上的重组氧化酶研究了各种芳烃中间体的分解情况。通过克隆、过表达及后续纯化，从嗜氯节杆菌A6中获得了催化儿茶酚及其类似物环裂解的羟基喹啉1,2-双加氧酶（CphA-I）。在不存在和存在N-羟基琥珀酰亚胺的情况下，通过物理吸附和共价偶联将该重组酶固定在SWCNTs上。固定化产率高达52.1%，固定化后保留了高达64.7%的高水平酶活性。动力学分析表明，固定化酶对所有评估底物的底物利用率（vmax）和催化效率（kcat/KM）与游离酶相似，表明固定化过程中酶活性损失最小。固定化酶在极端pH、温度和离子强度条件下比游离酶更稳定。因此，如果在现场条件下进行进一步研究，固定在SWCNTs上的氧化酶可作为生化修复过程中一种有效且稳定的生物催化剂。
