Immobilization Of Two-enzyme In Metal-organic Framework

Metal-Organic Framework(MOFs)is a kind of special crystal material formed by the self-assembly of metal ions and organic ligands.In recent years,MOFs can also be used as an immobilization matrix,which can fix certain biological macromolecules.This feature has increasingly attracted people's research interest.This article uses one of the MOFs materials called zeolitic imidazolate frameworks(ZIFs)and uses ZIF-8 in the ZIFs material class to immobilize two dual enzymes,DAAO-CAT and TH-DAAO.This is also a difficult point to study all the time,that is,the two enzymes are fixed together inside the support,which is a method that can use two enzymes simultaneously for continuous catalysis.Because of the precise control of the enzyme molar ratio,it is very difficult to spatially arrange the enzymes in close proximity.Therefore,immobilizing the double enzymes in this way not only achieves the precise control of the molar ratio of the two enzymes,but also achieves the co-immobilization of two enzymes that are in close proximity to each other.DAAO-CAT@ZIF-8 and TH-DAAO@ZIF-8 were slightly less active than free enzymes at 30? and 40? after the immobilized DAAO-CAT@ZIF-8 was tested for enzyme activity.With increasing temperature until 70? and 80?,the free enzyme has completely lost activity at this time,while the two immobilized enzymes,DAAO-CAT@ZIF-8 and TH-DAAO@ZIF-8,still retain about 65%of the free enzyme at 30?.It can be seen that neither of the two immobilized double enzymes showed a significant decrease in enzyme activity at normal temperature,but also maintained a good temperature tolerance at a very mode temperature,exhibiting considerable activity.In addition,this chapter also performed a series of characterization tests on composite materials DAAO-CAT@ZIF-8 and TH-DAAO@ZIF-8.Using SEM observer shape changes,we can know that by adjusting the concentration of Zn2+ and 2-methyl.The concentration of imidazole,or the concentration of the regulated protein,can directly control the crystal morphology of the composite.Confocal microscopy images can show that the enzyme is encapsulated inside the carrier and not simply adsorbed on the surface of the carrier.The XRD pattern can show that the composite has a lower diffraction intensity than the standard ZIF-8,which also indicates that the crystallinity of the ZIF-8 crystal decreases due to the addition of the protein.Both BET and Fourier infrared spectroscopy data consistently show that the entrapment mechanism of the two dual enzymes is not through the adsorption of the ZIF-8 material.The protein immobilized by ZIF-8 is different from the traditional immobilization method.In this way,in-suit encapsulation of the protein into the carrier is not only more stable than direct adsorption,the enzyme is not easily released,and the enzyme is also better protected,so that it has a better temperature tolerance.This article also further proves that MOFs have great potential for the development of biological macromolecules.