Construction And Application Of Novel Magnetic Nanocarriers In Biological Enzymes And Artificial Metalloenzymes

As a green and efficient catalyst,bioenzymes play important roles in many industrial processes.The appearance of artificial metalloenzyme enriches the types of catalysts to some extent.However,both biological enzymes and artificial metalloenzymes are faced with problems such as instability,inability to recycle,and short storage period.Therefore,we need to prepare a suitable carrier to improve the stability of biological enzymes or artificial metalloenzymes.In this paper,we have functionalized the magnetic silica nanomaterials according to the different types of enzymes to obtain new magnetic nanocarriers,and we explored the application value of these carriers in the immobilization of biological enzymes and artificial metalloenzymes.The main work contents and results are as follows:1.We prepared Ni-NTA functionalized magnetic silica nanocarriers and constructed HpaB and HpaC with histidine tag.The immobilization of bi-enzyme from the crude enzyme solution could be achieved by the affinity between nickel ions and histidine tags.The enzyme activity of immobilized bi-enzyme was 2.9 times higher than that of equivalent free enzyme activity.The pH stability,temperature stability and storage of immobilized enzyme were improved,and 60.94%activity remained after 7 times of use.2.We constructed a ruthenium-loaded artificial metalloenzyme that catalyzes the production of NAD(P)H by NAD(P)+and the regeneration of coenzyme in the presence of a hydrogen donor..At the same time,a biotin-functionalized magnetic silica nanocarier was prepared and immobilized on the ruthenium-loaded artificial metalloenzyme.The immobilized artificial metalloenzyme achieved recyclability and improved stability.After 10 days of storage at 4?,the immobilized enzyme retained 54.61%activity.After the immobilized ruthenium-loaded artificial metalloenzyme is combined with salicylic acid monooxygenase,the amount of catechol formed by catalyzing salicylic acid is about 10 times higher than that in the case where it is not used.3.Using the interaction between streptavidin and V02+,we constructed a vanadium-loaded artificial metalloenzyme with a simpler preparation method.The artificial metalloenzyme can catalyze the formation of benzathionine from thioanisole.After immobilization with biotinylated magnetic silica nanocarriers,pH and temperature stability,storage,and reusability are improved.