KINETICS, REGULATION, AND MECHANISMS OF BIOCHEMICAL REACTIONS

活性氧簇(ROS)被认为是导致非吞噬细胞酪氨酸磷酸化蛋白升高的信号转导。这种升高可以通过灭活蛋白酪氨酸磷酸酶(PTPs)来实现。我们和其他人在体外已经证明，高度丰富的PTP，PTP-1B，通过其催化部位C215的氧化被ROS失活。我们发现，对于钝化PTP-1B，超氧阴离子在动力学上比过氧化氢更有效，在化学上也更特异。最初的氧化产物C215亚磺酸衍生物很容易被进一步氧化成其不可逆衍生物，这一反应可以通过谷胱甘肽基化来防止。S谷胱甘肽基化的PTP-1B可以被硫代转移酶重新激活。因此，提出了一种由超氧阴离子和谷胱甘肽参与调节PTP-1B的信号转导机制。这一机制得到了体内实验的支持，即当A431细胞被表皮生长因子处理时，在C215处形成谷胱甘肽基化的PTP-1B。此外，超氧化物歧化酶(MnSOD)对超氧化物歧化酶的水平也有调节作用。我们发现佛波酯TPA通过蛋白激酶C催化的CREB-1/ATF-1样因子的磷酸化而不是通过已知的氧化还原敏感的转录因子，如NF-kappa-B或AP-1来诱导人MnSOD基因。过氧化氢通过激活磷脂酶C-伽马-2(PLC-伽马-2)的酪氨酸磷酸化和激活磷脂酰肌醇3-激酶来诱导细胞内钙释放。-蛋白质磷酸酶、自由基、氧化应激、快速反应、小泡、电穿孔、EPR、糖基化

Reactive oxygen species (ROS) have been implicated as signal transducers that cause an elevation of tyrosine phosphorylated proteins in non-phagocytic cells. This elevation can be achieved by inactivating protein tyrosine phosphatases (PTPs). We and others have shown in vitro that the highly abundant PTP, PTP-1B, is inactivated by ROS via oxidation of its catalytic site C215. We found that superoxide radical anion is a kinetically more efficient and chemically more specific oxidant than hydrogen peroxide for inactivating PTP-1B. The initial oxidized product, the C215 sulfenic derivative, can easily be oxidized further to its irreversible derivative, a reaction that can be prevented by glutathionylation. The S-glutathionylated PTP-1B can then be reactivated by thiotransferase. Thus, a signal transduction mechanism mediated by superoxide radical anion and the participation of glutathione is proposed for the regulation of PTP-1B. This mechanism is supported by the in vivo demonstration that glutathionylated PTP-1B at C215 is formed in A431 cells when they were treated with epidermal growth factor. In addition, the superoxide level is regulated by Mn(II)-superoxide dismutase (MnSOD). We found that a phorbol ester, TPA, induces human MnSOD gene via a protein kinase C-catalyzed phosphorylation of a CREB-1/ATF-1-like factor and not via known redox- sensitive transcription factors, such as NF-kappa-B or AP-1. Hydrogen peroxide has been shown to induce intracellular calcium release through the activation of phospholipase C-gamma-2 (PLC-gamma-2) via both tyrosine phosphorylation of PLC-gamma-2 and activation of phosphatidylinositol 3-kinase. - protein phosphatase, free radical, oxidative stress, fast reaction, vesicles, electroporation, EPR, glycation