KINETICS, REGULATION, AND MECHANISMS OF BIOCHEMICAL REACTIONS

点击翻译按钮获取中文摘要

Using a model system and EPR techniques, we have shown that the yellow fluorescent products, observed in glycated proteins, were generated via the interaction between amino acids and methylglyoxal. We showed that three free radical species, the cross-linked radical cation, the methylglyoxal radical anion, and the superoxide anion radical, were formed. The latter is formed only in the presence of oxygen. When the amino acid was replaced with bovine serum albumin, the cross-linked Schiff base and the cross-linked Schiff base radical cation of the protein was found to function like an enzyme for catalyzing the generation of free radicals. These results suggest that glycated proteins accumulated in vivo could accelerate oxidative damage. Protein tyrosine phosphatase (PTP) has been implicated to play a major role in cell signaling. We have studied the redox regulation of PTP-1B and found that the inactivation of PTP-1B and its recovery are more efficient with superoxide anion radicals as a signal relative to hydrogen peroxide. The inability to fully recover the PTP-1B activity with DTT, after its reaction with hydrogen peroxide, could be attributed to the fact that more methionines and cysteines were oxidized. Using our home-build electroporator system and phospholipid vesicles, we have studied the induction, distribution, and lifetime of the electric field-induced membrane pores. We also explored the possibility of developing a rapid kinetic method using loaded vesicles as reaction vessels since they can be ruptured in the microsecond time range by electric pulses. However, the heterogeneity problems due to vesicle preparation and its rupture need to be resolved.