FUNCTIONS OF THE HEPATIC GAMMA-GLUTAMYL CYCLE

描述：(改编自研究人员摘要)还原型谷胱甘肽(GSH) 在许多生化过程和干扰中起着关键作用 在它的动态平衡中与许多疾病的病因和进展有关 疾病的威胁。这三个三肽的周转的第一步 哺乳动物细胞是其进入细胞外空间的运输；然而， 调节谷胱甘肽外流的运输系统仍不清楚。在肝脏中， 作为谷胱甘肽合成的主要部位，谷胱甘肽以很高的速率释放到两种血液中。 血浆和胆汁。谷胱甘肽转运到胆汁中是胆汁的驱动力 分泌，在药物的肝脏解毒中起重要作用， 金属，以及内源和外源的其他活性化合物。 谷胱甘肽也会通过肝窦膜释放到血浆中，以 递送到其他组织。我们最近的研究提供了重要的见解 谷胱甘肽转运的分子机制。特别是，我们证明了 Oatp1是一种正弦有机溶质转运蛋白，具有谷胱甘肽/有机转运蛋白的功能 溶质交换器。这一发现不仅确定了能量耦合 Oatp1的机制，但也阐明了GSH释放到血液中的途径 血浆，以及对GSH的一种新功能。此外，我们还展示了 Ycf1p是哺乳动物MRP蛋白的酵母同源物，介导ATP依赖， 低亲和力GSH转运，提示MRP可能介导GSH外流。 哺乳动物细胞。 拟议研究的总体目标是确定和表征GSH 和谷胱甘肽-S的偶联转运机制，特别是要测试 OATP和MRP转运蛋白家族介导细胞GSH的假说 放手。我们的具体目标是：(1)测试最近克隆的OATP2 与燕麦蛋白1同源的转运蛋白，也参与谷胱甘肽/有机溶质的调节 交换。(2)建立了谷胱甘肽在燕麦上转运的动力学和特异性。 如果该转运蛋白也被发现作为GSH交换器，则在OATP2上也是如此。 (3)测试谷胱甘肽是mrp2的底物还是共底物，如果是；a) 定义小管膜蛋白运输的能量学，b) 将这些参数与酵母Ycf1p和c)上GSH转运的参数进行比较。 检查谷胱甘肽是否也是mrp3和mrp6的底物 肝细胞侧膜蛋白；以及(4)继续确定其作用 肝内谷氨酰循环在谷胱甘肽处置中的作用 S-偶联物及其转化为相应的硫代糖酸 研究硫代尿酸的转运机制。

DESCRIPTION: (Adapted from investigator's abstract) Reduced glutathione (GSH) plays a critical role in a multitude of biochemical processes, and disturbances in its homeostasis are implicated in the etiology and progression of a number of diseases. The initial step in the turnover of this tri-peptide in all mammalian cells is its transport into the extracellular space; however, the transport systems that mediate GSH efflux remain poorly defined. In the liver, a major site of GSH synthesis, GSH is released at high rates into both blood plasma and bile. GSH transport into bile functions as a driving force for bile secretion and plays an important role in hepatic detoxification of drugs, metals, and other reactive compounds of both endogenous and exogenous origin. GSH is also released across the sinusoidal membrane into blood plasma for delivery to other tissues. Our recent studies provide important insight into molecular mechanisms of GSH transport. In particular, we demonstrated that oatp1, the sinusoidal organic solute transporter, functions as a GSH/organic solute exchanger. This finding not only identifies the energy coupling mechanism for oatp1, but also elucidates a pathway for GSH release into blood plasma, as well as a novel function for GSH. Moreover, we demonstrated that Ycf1p, the yeast orthologue of mammalian mrp proteins, mediates ATP-dependent, low-affinity GSH transport, indicating that mrp may mediate GSH efflux in mammalian cells. The overall goals of the proposed studies are to identify and characterize GSH and glutathione S-conjugate transport mechanisms, and in particular to test the hypothesis that the oatp- and mrp-family of transporters mediate cellular GSH release. Our specific aims are: (1) Test whether oatp2, a recently cloned transporter that is homologous to oatp1, also mediates GSH/organic solute exchange. (2) Establish the kinetics and specificity of GSH transport on oatp1, and on oatp2 if this transporter is also found to function as a GSH exchanger. (3) Test whether GSH is a substrate or a co-substrate for mrp2, and if so; a) define the energetics of transport on this canalicular membrane protein, b) compare these parameters with those for GSH transport on yeast Ycf1p, and c) examine whether GSH is also a substrate for mrp3 and mrp6, putative hepatocellular lateral membrane proteins; and (4) Continue to define the role of the intrahepatic -glutamyl cycle in the disposition of glutathione S-conjugates and their conversion to the corresponding mercapturic acids by examining mechanisms of mercapturic acid transport.