| Reactive oxygen species(ROS)are products of the normal metabolic activities of aerobic living organism and are produced in response to various stimuli. ROS are harmful to cells when oxidative metabolism products increase or the antioxidant defenses lack in the living organism. Lots of researches showed that ROS are related to some physiologic and pathological processes and play key roles in explaining the pathogenesis of many diseases. ROS are very active and very strong oxidant and they can attack biomolecules to damage the structure and function of cells and tissues.GPX are the well-known antioxidant selenoenzymes in organisms which destroy several harmful hydroperoxides and then maintain the metabolicbalance of ROS in vivo, thus protecting the biomembranes and other cellular components from oxidative damage. In certain pathogenic states, the production of ROS is enhanced and the excessROS damage various biomacromolecules including RNA,DNA, protein, sugars and lipids, and therefore result in ROS-mediated diseases. ROS-related diseases include reperfusion injury, inflammatory process, age-related diseases, neuronal apoptosis cancer and cataract. Therefore, GPX can really act as antioxidant drugs. Unfortunately, scientists can not fully understand the structures of GPX as well as its catalysis mechanism in vivo at the present time. Fabrication of GPX models offers an ideal alternative for elucidating the origin of substrate binding and catalysis of enzyme.Mimicking the natures of molecular recognition and catalysis of enzymes by artificial enzymes is very essential for exploring the evolved biological process of enzymes as well as their properties of structures and functions. On the basis of structural understanding for GPX, we designed Se-3P and Se-6P to imitate the properties of GPX.1.Construction of selenium-containing 3-mer peptides to imitate GPXAccording to the results of Epp's study, around active sites architecture of nature GPX, Gln79 and Trp153 are located within hydrogen bonding distance to the selenium atom and have been suggested to play functional roles in catalysis. A further comparison of amino acid sequences indicated that these two residues are in fact conserved in the whole glutathione peroxidase superfamily. Thus, we design six Se-3P QUW,QWU,WQU,WUQ,UWQ,UQW, and we synthesis all of them with solid-phase synthesis technology based on the active center of native GPX. Among the six Se-3P, UWQ has the highest GPX activity and shows similar kinetic behavior as natural GPX, that is abide the Ping-Pong mechanism.2.Construction of selenium-containing 6-mer peptides to imitate GPX Among the active-site residues which could be of functional importance for substrate binding and catalysis are Arg-167, Arg-40, Gln-130 and Trp-148. During binding interaction theγ-glutamyl carboxyl group of the GSH molecule is presumably fixed by a salt bridge to Arg-167, whereas the N-terminal amino group could form a hydrogen bond to Gln-130. The C-terminal glycyl carboxyl group, seems to form an additional salt bridge with Arg-40. We design the 6-mer peptide RQUWRQ and QQRUWR, based on the theory of Surface-simulation synthesis and combining with molecular modeling and molecular dynamics methods. The optimal pH for RQUWRQ catalyzed reduction of H2O2 by GSH is found to be 8.94 and the optimal temperature was found to be 58.2℃. The kinetic analysis show that the mechanism of RQUWRQ catalyzed reduction of H2O2 by GSH is Ping-Pong mechanism.
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