Study On Protein Nitration And Protein Nitration Sites Selectivity By FeTPPS

Fe(III)meso-Tetra(4-sulfonatophenyl)porphine chloride(FeTPPS) is a kind of synthetic iron porphyrin compound with pseudo-peroxidase activity, because of the hydrophily property, it provided us with good conditions for biomimetic catalysis research, which solved the problem that heme cannot effectively simulate biochemical reactions in vivo because of its hydrophobicity. Moreover, FeTPPS is an effective scavenger of peroxynitrite(ONOO-), which can inhibit nitration reaction mediated by ONOO-. However there has not been any paper about FeTPPS catalyzed protein nitration in the presence of nitrite and hydrogen peroxide.In this paper, firstly we studied the peroxidase activities of FeTPPS: The data showed that FeTPPS has good pseudo-peroxidase activity. Then western blotting and spectrophotometer methods were used to determine whether the FeTPPS-NaNO2-H2O2 system could nitrate and oxidize bovine serum albumin(BSA) as well as a series of corresponding conditions. At the same time this system caused mice brain protein nitration was also studied, all these results were compared with that caused by the heme-NaNO2-H2O2 system. Both of the two systems could well nitrate and oxidize BSA,and reactions respectively have the concentration, pH and time dependence. In addition,there were some difference of protein nitration sites in vivo casused by different iron porphyrins. Finally, to compare the possible nitration sites of glyceraldehyde-3-phosphate dehydrogenase(GAPDH) catalyzed by heme or FeTPPS that mass spectrometric(LC-MS/MS) and Expasy ProtScale software were used. The results indicated that different iron porphyrin had different nitration site selectivity: heme tended to catalyze the nitration sites located in relatively hydrophobic environments; while FeTPPS selectively catalyzed the nitration sites which were located in relatively hydrophilic environments.The above research results show that although FeTPPS could be used as ONOOdecomposition catalyst to reduce the content of nitrotyrosine(3-NT), due to its chemical properties(pseudo-peroxidase activity), FeTPPS can cause tyrosine nitration in the presence of nitrite and hydrogen peroxide. Therefore, when using FeTPPS as ONOOscavengers to inhibit nitration reaction, we must pay attention that FeTPPS may alsopromote nitration reaction under oxidative conditions. And our study further confirmed the conclusion that protein nitration sites are affected by the solubility of iron porphyrin.Although having the same active center, the sites of nitrated tyrosine residues are depended on iron porphyrin's hydrophily.