STRUCTURE OF AS160, AKT SUBSTRATE REGULATING GLUCOSE TRANSPORTER TRANSLOCATION

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 一旦感觉到血液中的高血糖水平，胰岛β细胞就会分泌胰岛素来降低动态平衡所需的葡萄糖水平。胰岛素与胰岛素受体结合，触发多个激酶级联反应，最终导致葡萄糖转运体从细胞内的储存囊转移到细胞膜。膜结合的葡萄糖转运体允许外部葡萄糖进入细胞，最终降低血糖。这种胰岛素作用降低导致的高血糖水平是糖尿病疾病的特征。与葡萄糖转运蛋白转运有关的最远下游靶点是最近发现的蛋白AS160(Akt底物160 kDa)被胰岛素激活的激酶Akt磷酸化。最近的研究表明，AS160在胰岛素作用下的磷酸化会使RabGTP酶失活，这是葡萄糖转运蛋白内化过程中的一个关键事件。我们的结构目标是这种AS160蛋白充当开关来开启/关闭胰岛素信号。我们特别感兴趣的是AS160 C-末端有一个300个残基的RabGAP结构域，它负责激活RabGTP酶。在自制的旋转阳极X射线源上，我们获得了人AS160RabGAP结构域的晶体，其衍射率可达6°。我们相信，我们项目的成功在很大程度上取决于能否使用同步加速器设施。由于我们成功地结合了重金属并稳定了晶体，我们打算在可调谐的光束线上进行MAD实验，这对相位确定将是重要的。我们相信RabGAP结构域本身将是信息丰富的，因为结构可以引导我们设计一种使AS160失活的药物来模拟胰岛素的作用。这一新的方法瞄准了降低血糖的最大下游途径，可能是一种很有前途的治疗方法，用于测试对胰岛素无效的糖尿病患者。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Upon sensing high glucose levels in blood, pancreatic beta-cells secret insulin to lower the glucose level necessary for homeostasis. Insulin binds to insulin receptor and triggers multiple kinase cascades, which in the end lead to the translocation of glucose transporters from the intracellular storage vesicles to the cell membrane. Membrane bound glucose transporters allow the influx of the outside glucose into the cell to ultimately lower the blood glucose. High blood glucose levels caused by a decrease in this insulin action characterize the condition of the disease diabetes. The farthermost downstream target involving this glucose transporter translocation is the phosphorylation of a recently discovered protein AS160 (Akt substrate of 160kDa) by an insulin-activated kinase, Akt. Recent studies suggest that AS160 phosphorylation upon insulin action deactivates RabGTPases, which is a crucial event in the internalization of glucose transporters. Our structural target is this AS160 protein acting as a switch to turn on/off the insulin signal. We are especially interested in a 300-residue RabGAP domain at the C-terminus of AS160, which is responsible for activating the RabGTPase. We have obtained crystals of human AS160 RabGAP domain, which diffracts to 6¿¿¿ resolution at our home rotating anode X-ray source. We believe that a success in our project depends heavily on the access to a synchrotron facility. Since we were successful at incorporating heavy metals and stabilizing the crystals we intend to perform MAD experiments at a tunable beamline, which would be important for the phase determination. We believe that RabGAP domain by itself will be informative since structure can lead us in designing a drug that inactivates AS160 to simulate the action of insulin. This novel approach targeting the utmost downstream pathway in lowering blood glucose may be a promising treatment to test in diabetics to whom their response to insulin is failing.