Mechanism of GLUT4 Inhibition by HIV Protease Inhibitor

描述（由申请人提供）：本项目的长期目标 是了解促进葡萄糖代谢的分子机制 运输 最近发现HIV蛋白酶抑制剂能够 选择性 抑制GLUT 4， 的 主要 胰岛素应答 葡萄糖 运输，是第一份报告，它是可能的选择性和 可逆地抑制单个易化葡萄糖转运蛋白的活性 同种型 这种抑制发生的机制仍然未知。的 本研究的具体目标是调查的机制基础， HIV蛋白酶抑制剂对GLUT 4的抑制作用，并确定 对GLUT 4的体外作用可以在体内复制，导致急性和 可逆性胰岛素抵抗首先，仔细的动力学分析 将使用异源表达的GLUT 4进行抑制过程， 非洲爪蟾卵母细胞。 接下来，HIV蛋白酶抑制剂结合GLUT 4的位点 将通过爪蟾卵母细胞中转运蛋白的光标记来确定 和/或3 T3-L1脂肪细胞， 抑制剂衍生物。 修饰蛋白将通过表面增强分析仪进行分析。 激光解吸/电离（SELDI）质谱法。最后，能力 HIV蛋白酶抑制剂急性可逆地引起胰岛素抵抗 将通过测量正常血糖下的葡萄糖处置来研究体内 正常人和糖尿病易感者高胰岛素钳夹状态 啮齿动物更好地理解生物多样性活动的机制 促进性葡萄糖转运蛋白可以在同种型中被急性调节 为研究葡萄糖稳态提供了一种新的手段， 正常个体和具有葡萄糖调节障碍的个体，例如 糖尿病的这项研究的结果也可能有助于 开发新的HIV蛋白酶抑制剂， 艾滋病毒治疗，同时避免其不利的代谢后果。

DESCRIPTION (provided by applicant): The long-term objective of this project is to understand the molecular mechanisms involved in facilitative glucose transport. The recent discovery that HIV protease inhibitors are capable of selectively inhibiting GLUT4, the major insulin responsive glucose transporter, is the first report that it is possible to selectively and reversibly inhibit the activity of a single facilitative glucose transporter isoform. The mechanism by which this inhibition occurs remains unknown. The specific goals of this research are to investigate the mechanistic basis for the inhibition of GLUT4 by HIV protease inhibitors and determine whether the in vitro effects on GLUT4 can be replicated in vivo, leading to acute and reversible insulin resistance. First, a careful kinetic analysis of the inhibition process will be conducted using GLUT4 heterologously expressed in Xenopus oocytes. Next, the site of HIV protease inhibitor binding to GLUT4 will be determined through photolabeling of the transporter in Xenopus oocytes and/or 3T3-L1 adipocytes using a synthesized reactive retroviral protease inhibitor derivative. Modified protein will be analyzed by Surface Enhanced Laser Desorption/Ionization (SELDI) mass spectrometry. Finally, the ability of HIV protease inhibitors to acutely and reversibly cause insulin resistance in vivo will be investigated by measuring glucose disposal under euglycemic hyperinsulinemic clamp conditions in both normal and diabetes susceptible rodents. A better understanding of the mechanism by which the activity of facilitative glucose transporters can be acutely modulated in an isoform specific manner will provide a new means of studying glucose homeostasis in normal individuals and those with disorders of glucose regulation, such as diabetes mellitus. The results of this research may also facilitate the development of newer HIV protease inhibitors that maintain their efficacy in HIV treatment while avoiding their adverse metabolic consequences.