| Glutathione peroxidase (GPx) was the first mammalian enzyme characterized as selenoprotein. As the primary enzyme with antioxidant activity, it catalyses the reduction of harmful peroxides by glutathione and protects the cell membrane from oxidative damage. Consequently, it has the potential to cure various ROS-related diseases such as reperfusion injury, inflammatory process, age-related diseases, neuronal apoptosis, cancer and cataract. Due to the limitations associated with native GPX (solution instability, limited cellular accessibility, immunogenicity, short half-lives, costs of production, and proteolysis digestion), scientists have made a great deal of efforts to study the structure -activity correlation of native GPX, prepared many GPX mimics and studied the biological effects of GPX mimics. There are two principal phenomena that underlie the activity of enzyme: substrate binding and the subsequent intracomplex catalysis. When an enzyme model is to build, it is necessary to consider that enzyme model can recognize and bind substrate, and that the catalytic group is at a correct position to interact with the reactive group of substrate in order to promote catalysis. The well-known GPX mimic, Ebselen, has been undergoing Ⅲ phase clinical trial as antioxidant drug in Japan, but its GPX activity is only about 1 U/μmol. Based on the structure-function correlation of native GPX, we produce a new kind of cyclodextrin-derived GPX mimics, 6A, 6A'-Imidazole-6B, 6B'–selenium –bridged β-cyclodextrin to imitate the functional action of native GPX. Elemental analysis, IR, 13C NMR and X-ray photoelectron spectroscopy were used to characterize the mimic. The GPX activity of the mimic with H2O2 is 6.4 U/μmol. The activity of the mimic can maintain a high level at pH's ranging from 7 to 9. The optimal pH of the native enzyme is 8.8, so the mimic is more tolerant in pH change than its native counterpart. Detailed kinetic studies of 6A, 6A'-Imidazole -6B, 6B'– selenium–bridged β-cyclodextrin was undertaken. Saturation kinetics was observed for both H2O2 and GSH. Double reciprocal plots of the initial velocity versus the concentration of substrates were a family of parallel lines, which is consistent with a Ping-Pong mechanism involving at least one covalent enzyme intermediate. The kinetic equation was similar to native GPX, the high GPX activities of 6A, 6A'-Imidazole-6B, 6B'–selenium–bridged β-cyclodextrin can be explained by increase of ability of cyclodextrin to bind GSH. In the previous study, we synthesized a series of GPX mimic with GPX activity. The simulation of the catalytic cycle of GPX by computer-assisted molecular modeling suggested that there is interaction between selenium and amino group in the active site. The introduction of amino group proximal to the selenium can stabilize and activate it; therefore, it promotes the reaction.Since GPX plays a pivotal role in protection cells against oxidative damage, selenium compounds with GPX activity attract great attentions of scientists. We constructed the ferrous sulfate/ascorbate-induced mitochondria damage model system to evaluate the biological effects of the mimic. The model system was used to demonstrate that the damaged mitochondria have great changes in morphology, structure and function. The extent of swelling, lipid peroxide of mitochondria was chosen as measures to determine the extent of injury and protection of mitochondria. The mimic can reduce the swelling of mitochondria during damage and decrease the level of MDA accumulation. These data show that the mimics have great antioxidant activity.This mimic as well as the others derived from cyclodextrin, which generated by our group, has the advantage of low molecular weight, good solubility and stability. The feature of this mimic is that its activity is higher than 6-SeCD because we introduce an amino group which can stabilize intermediates. Due to special hindrance of the imidazoles that were introduced at the primary face of cyclodextrin, the activity of the mimic is not a...
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