KINETICS, REGULATION AND MECHANISMS OF BIOCHEMICAL REACTIONS

蛋白质泛素化与许多细胞内 流程. 所涉及的酶是：泛素活化酶，E1， 泛素载体酶E2，其将活化的泛素递送至 其连接酶E3或直接泛素化其靶蛋白。 E1是 发现存在两种异构体，命名为E1- 117 kDa和E1- 110 kDa。 E1 在HeLa细胞中发现磷酸化，表明蛋白质 泛素化可以通过磷酸化/去磷酸化 机制 这与体外研究结果雅阁，表明E1是 被蛋白激酶C磷酸化，和两种E2亚型，E2- 32 kDa 和E2- 20 kDa被酪氨酸激酶磷酸化，并被一种新的激酶磷酸化， 在HeLa细胞的胞质组分中，分别。 转录 发现Fos和Jun因子经历多重泛素化。 当完整细胞膜受到某一临界电场作用时， 强度，它们变得瞬时透化。 该瞬态时间为 与悬浮介质的离子强度成反比。 在 此外，离子强度也影响了 相对于电极的透化，以及体对体的透化。 膜电位 电子顺磁共振谱和自旋捕获方法， 用于识别和监测自由的形成和利用 根的 以前，我们发现Cu，Zn-SOD（Cu，Zn-SOD） 催化过氧化氢转化为羟基自由基， 从酶结合的阴离子配体产生二级自由基，或 食腐动物 因此，Cu，Zn-SOD活性的增强会产生损伤作用 影响细胞。 当NCB-20细胞，小鼠神经母细胞瘤-中国 仓鼠脑杂交细胞系，用人pSVneo Cu，Zn- SOD基因表达载体，还原型谷胱甘肽水平为 大幅减少。 这表明Cu，Zn-SOD在细胞内过量表达 由于自由基的产生，细胞维持慢性促氧化状态 由Cu，Zn-SOD的高水平催化。 肝细胞和系膜细胞胞浆Ca（II）振荡已被证实， 研究了 结果发现，加压素诱导的肌醇产生 三磷酸盐以初始峰值进行，然后降低到升高的 相对于基础水平的稳态水平。 这些数据表明 Ca（II）振荡的频率，其取决于 激动剂，与肌醇的稳态水平相关 三磷酸盐

Protein ubiquitination has been implicated in numerous intracellular processes. The enzymes involved are: the ubiquitin activating enzyme, E1, the ubiquitin carrier enzyme, E2, which delivers the activated ubiquitin to its ligase, E3, or directly ubiquitinates its target proteins. E1 was found to exist as two isoforms, designated as E1-117kDa and E1-110kDa. E1 in HeLa cells was found to be phosphorylated, suggesting protein ubiquitination may be regulated by the phosphorylation/dephosphorylation mechanism. This is in accord with the in vitro studies showing E1 is phosphorylated by protein kinase C, and two of the E2 isoforms, E2-32kDa and E2-20kDa are phosphorylated by a tyrosine kinase, and by a novel kinase in the cytosolic fraction of HeLa cells, respectively. The transcription factors, fos and jun, were found to undergo multiple ubiquitinations. When membrane of intact cells is subject to some critical electric field strength, they become transiently permeabilized. This transient time is inversely proportional to the ionic strength of the suspending medium. In addition, the ionic strength also affects the location of the permeabilization with respect to the electrodes, and the bulk-to-bulk membrane potential. Electron paramagnetic resonance spectroscopy and spin-trapping methods were used to identify and monitor the formation and the utilization of free radicals. Previously, we found that Cu,Zn-superoxide dismutase (Cu,Zn-SOD) catalyzes the conversion of hydrogen peroxide to hydroxyl radical which can generate secondary radicals from enzyme-bound anionic ligands or scavengers. Therefore, enhanced Cu,Zn-SOD activity can produced damaging effects to the cells. When NCB-20 cells, a mouse neuroblastoma-Chinese hamster brain hybrid cell line, is transfected with a human pSVneo Cu,Zn- SOD gene expression vector, the level of reduced glutathione is significantly reduced. This indicates that the Cu,Zn-SOD overexpressed cells maintain chronic pro-oxidant states due to free radical production catalyzed by the elevated level of Cu,Zn-SOD. Cytosolic Ca(II) oscillation in hepatocytes and mesangial cells has been investigated. It was found that vasopressin-induced production of inositol triphosphate proceeds with an initial spike and then reduces to an elevated steady-state level relative to that of basal. The data suggest that the frequency of Ca(II) oscillation, which is dependent on the concentration of agonist, is correlated with the steady-state level of inositol triphosphate.