| Protein plays a key role in cell life activities.Protein separation and purification is an important part of protein research.The traditional protein separation method generally has the limitations of large preparation workload,small adsorption capacity,expensive cost or cumbersome separation and purification process in the early stage.Therefore,it is of great significance to develop a simple and efficient method for protein separation.Magnetic separation technology has become one of the most widely used techniques in the field of protein separation due to its high biocompatibility,easy operation,regulation and modification.However,there are some drawbacks such as magnetic core aggregation,low adsorption capacity and poor repeatability.Aiming at the key scientific problems mentioned above,Fe3O4@PDA@NiAl-LDH and Fe3O4@PDA@NiFe-LDHwith large specific surface area were prepared by layer-by-layer self-assembly using polydopamine（Polydopamine,PDA）as binder.PDA could effectively inhibit the aggregation of magnetic core and grow the lamellar structure with large specific surface area on its surface.XRD,SEM,TEM and BET were used to characterize their physicochemical properties and histidine tag protein was used as the target for experimental exploration to evaluate the separation and purification efficiency of magnetic materials for the target protein.The specific contents are composed of the following two parts:（1）Fe3O4@PDA@NiAl-LDH and Fe3O4@PDA@NiFe-LDH were successfully prepar ed by solvothermal method.In order to prevent magnetic core aggregation,dopamine（Dopa mine,DA）can be self-polymerized into PDA films in alkaline environment.Finally,the LD H lamellar structure with large specific surface area on the surface of magnetic core was fur ther modified with PDA as adhesive,and finally Fe3O4@PDA@NiAl-LDH and Fe3O4@PD A@NiFe-LDH with uniform size were obtained,which were about 595 nm and 482 nm,res pectively.The prepared material has stable crystal form and is not easy to aggregate,has bet ter dispersibility,better thermal stability,recoverability and strong magnetism,and the speci fic surface area increases from 5.2 m2/g of pure Fe3O4 to 19.4 and 17.5 m2/g,and the additio n of lamellar structure increases the specific surface area to provide more active sites,which is conducive to the subsequent protein separation.（2）Isolation and purification of histidine-tagged protein:The isolation and purification of histidine-tagged protein by Fe3O4@PDA@NiAl-LDH and Fe3O4@PDA@NiFe-LDH were systematically evaluated using E.coli cell lysate containing histidine-tagged protein as a research template,and their adsorption effect,adsorption ability and reusability under different reaction conditions were investigated.According to the complex cell lysate,the prepared magnetic nanoparticles（Magnetic Nanoparticles,MNP）with core-shell structure can realize the rapid adsorption and efficient purification of histidine-tagged protein,and its adsorption capacity is 612 mg·g-1 and 492 mg·g-1,respectively;when the material is mixed with the complex cell lysate for 2 h,the eluted protein band is the thickest,demonstrating that the adsorption capacity of the two to histidine-tagged protein is the strongest;the content of Ni（NO3）2·6H2O is regulated,and it is known that the optimal addition amount of Fe3O4@PDA@NiAl-LDH and Fe3O4@PDA@NiFe-LDH is 1.48 g and 0.35 g;after five repeated uses,the adsorption capacity does not change significantly,reflecting good stability and repeated utilization rate.The prepared three-dimensional graded core-shell structure can reduce the aggregation of magnetic nuclei,and the lamellar structure of LDH increases the specific surface area of the material and provides more active sites,which is conducive to the separation and purification of histidine-tagged proteins.The results provide an economical,efficient and rapid material for the rapid isolation and purification of histidine tag proteins and have a wide range of application prospects. |
