The Mimic Construction Of Nanocylinder-shaped Hemoprotein And Its Catalytic Oxidation Performance And Applications

In biology systems,catalytic oxidations are catalyzed by metal containing proteins.This includes reactions taking place in water at room temperature under atmospheric pressure that perfectly fit the current concept of green chemistry.Since the beginning of this century,many teams have addressed the question of clean oxidation by elaborating artificial metalloproteins.In many of the synthesis methods,using the hostgust interaction is one of the most simple and straightforward method,which directly introduce guest hemin into host protein.However,due to the differences in the structures and properties of proteins and the different binding force,they also have some problems at present,such as low stability of protein structure,insufficient functionality,incomplete theoretical research,not prominent of specificity and so on.We still need to explore more studies of artificial enzyme synthesis and functionalization,and get more excellent artificial enzymes,and expand their in biological analysis and environmental monitoring.In this paper,we studied the synthesis and properties of artificial hemozyme,and explored the practical application values.The chaperonin protein Gro EL was used as the host protein.It is tetrameric protein complex composed of two chemically identical heptameric rings stacked back-toback.The Gro EL takes on a cylindrical structure with a diameter of 13.7nm and a length of 14.6nm.It assists the misfolded proteins into the correct folding.On the one hand,it can seve as a carrier for hemin binding,and on the other hand,it can serve as a nanoscale reaction vessel.The main contents are as follows:1)The artificial enzyme was synthesized throng detergent dialysis mothed,which has peroxidase-like activity using Gro EL and hemin.The effects of temperature,p H,and concentration on enzyme activity and the stability of artificial enzymes were investigated.Gro EL effectively dispersed and located the hemin moleculas,and the molecular structure of Gro EL did not change significantly.The constructed hemin-Gro EL artificial enzyme has high peroxidase-like activity,and in catalytic experiments,its kinetic performance follows the ping-pong mechanism.2)The colorimetric detection assay based on hemin-GroEL artificial enzyme has been successfully established by using colorimetric substrates such as 3,3',5,5'-Tetramethylbenzidine(TMB)and o-Phenylenediamine(OPD).In the kinetic analysis of the colorimetric reaction,the Hemin-Gro EL enzyme also has peroxidase-like activity which follows the typical Michealis-Menten kinetics and ping-pong mechanism.The colorimetric assay based on hemin-Gro EL can get the detection of H2O2 and glucose in solution.In addition,the enzyme can also be used for the oxidation removal of catechol,which provides an important reference for practical applications,such as clinical analysis and environmental monitoring.3)The fluorescence detection assay based on Hemin-Gro EL artificial enzyme has been successfully established by using fluorescent substrates homovanillic acid(HVA).The kinetic analysis of fluorescent reaction and the competitive inhibition assay of the substrate shows that Hemin-Gro EL artificial enzyme have low substrate specificity.And with increasing the substrate concentration,the substrate inhibition could be reduced or even eliminated.Fluorescence detection assay based on Hemin-Gro EL can also determine H2O2 and glucose in solution.In addition,the assay can also be used for the detection of enzymes related substrate,such as glucose oxidase,which is a potential new type artificial enzymes.