Improvement Of The Stability Of Immobilized Enzyme By Polyethylenimine-induced Biomimetic Silicification

In this work,polyethyleneimine(PEI)modified magnetic nanoparticles(MNPs)were prepared for the immobilization of phosphoenolpyruvate carboxylase(PEPcase)by electrostatic interaction,denoted as MNPs-ENZ.Then,the surface of MNPs-ENZ was coated by a silica layer through PEI-induced biomimetic silicification to improve the stability of the immobilized enzyme,denoted as MNPs-ENZ@Si O₂.The PEI modified nanoparticles(MNPs-PEI)and two immobilized enzymes(MNPs-ENZ and MNPs-ENZ@Si O₂)were characterized by transmission electron microscopy,X-ray photoelectron spectroscopy,vibrating sample magnetometer and nanometer particle size analyzer,etc.It was proved the modification of PEI,immobilization of PEPcase and the coating of silica layer were successfully achieved.Moreover,the biomimetic silicification did not affect the superparamagnetism of nanoparticles,but improved their dispersion in aqueous solution.Enzymatic kinetics results showed that the activities of the two immobilized enzymes(MNPs-ENZ and MNPs-ENZ@Si O₂)were significantly improved(150%),as comparing to the free enzyme.Moreover,coating of silica layer did not significantly change the activity of immobilized enzyme.Furthermore,the biomimetic silicification did greatly improve the thermal stability,p H stability and recyclability of MNPs-ENZ@Si O₂.For instance,93%of activity was maintained after incubation 1 h even at 50?or p H 10.0,and 77%of activity was reserved after 8 cycles.Above results indicated that MNPs-PEI could improve the stability of the PEPcase,and it is a suitable carrier for enzyme immobilization.In addition,the biomimetic silicification could further enhance the stability and dispersity of the immobilized enzyme,proving its potential applications in industry.Besides,immobilization of carbonic anhydrase was attempted,by PEI adsorption,covalent linking with polydopamine,and cross-linking,which is beneficial in the co-immobilization of double enzymes in further.The experimental results could help the selection of suitable enzyme immobilization method,as well as promote the application of magnetic nanoparticles,polyethyleneimine and biomimetic silicification in enzyme immobilization.