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Fabrication of Ni2+-nitrilotriacetic acid functionalized magnetic mesoporous silica nanoflowers for one pot purification and immobilization of His-tagged ω-transaminase

Cao, Guangxiu, Gao, Jing, Zhou, Liya, Huang, Zhihong, He, Ying, Zhu, Meng, Jiang, Yanjun
Biochemical engineering journal 2017 v.128 pp. 116-125
X-ray diffraction, amines, catalytic activity, magnetism, nanoflowers, pH, porous media, scanning electron microscopy, silica, storage quality, temperature, thermal stability, transmission electron microscopy
ω-transaminase (ω-TA) has gained much attention due to its application in the preparation of chiral amines. It is of increasing interest to obtain stable and reusable immobilized ω-TA for economic catalytic process. A major approach for simple and efficient immobilization is introduction of an affinity tag to the target enzyme. Thus, in this study, the recombinant ω-TA with His-tag was successfully expressed in E.coli Rosetta (DE3). To realize the one pot purification and immobilization of ω-TA, Ni²⁺-nitrilotriacetic acid functionalized magnetic mesoporous silica nanoflowers (Ni-NTA/MMS-NF) were synthesized for the first time. The morphology, structure, and composition of the Ni-NTA/MMS-NF were characterized by scanning electron microscopy, transmission electron microscopy, X-ray diffraction, etc. The immobilized ω-TA had the same optimum temperature with free ω-TA, and the optimum pH of the immobilized ω-TA was shifted from 7.5 to 7.0. Compared to free ω-TA, the immobilized ω-TA showed increased thermal stability, better pH stability, and excellent storage stability. Additionally, the immobilized ω-TA preserved 67.38% of initial activity after reusing twelve times. These results revealed that Ni-NTA/MMS-NF might have great potential in the purification and immobilization of His-tagged enzyme.