HIV VIRION RELEASE AND VIRAL PROTEIN U (VPU)

本提案中描述的实验旨在 了解“病毒蛋白U”(VPU)介导 细胞中存在HIV-1颗粒。潘加尼班博士的实验 这里描述的是一种新的细胞蛋白的作用 在酵母、体外和人类细胞中直接与VPU相互作用。 这种新的细胞蛋白，U-结合蛋白(UBP)是一个 包括免疫亲和素和一类 含四肽重复序列的丝氨酸/苏氨酸磷酸酶 (TPR)。Panganiban博士和他的同事们有几个补充 目标。他们将进行实验，以检验 更详细的VPU和UBP。尤其是。尤其是他们会 确定蛋白质-蛋白质所需的两种蛋白质的区域 相互作用，分离表现出结合稳定性改变的突变体，以及 验证VPU作用是通过UBP介导的假设。潘加尼班医生的 初步数据还表明，UBP与HIV-1 Gag直接相互作用 人HIV-1 Gag蛋白衣壳结构域体外表达 细胞。因此，他将确定UBP和GAG上需要的站点 为了这次互动。为了考察UBP对颗粒释放的影响， UBP和Gag野生型和突变型表达的影响 是此交互所必需的。要考察UBP对 颗粒释放、野生型和突变型表达的影响 将确定UBP在颗粒释放上的浓度。同样，他们也可能 确定内源性UBP表达的消融是否影响 颗粒释放效率。最后，由于UBP以前没有 Panganiban博士和他的同事们将研究 UBP的内在属性，并试图洞察其正常作用 在牢房里的UBP。这些研究的总体目标是制定一项 全面的图像，解释了分子机制 VPU介导了颗粒的释放。

The experiments described in this proposal are directed toward understanding the mechanism by which "viral protein U" (Vpu) mediates the exist of HIV-1 particles from cells. Dr. Panganiban's experiments described here are centered around the role of a novel cellular protein that interacts directly with Vpu in yeast, in vitro and in human cells. This novel cellular protein, "U-binding protein (Ubp) is a member of a protein superfamily that includes the immunophilins and a class of serine/threonine phosphatases containing tetratrico-peptide repeats (TPRs). Dr. Panganiban and his colleagues have several complementary goals. They will carry out experiments to examine the interaction between Vpu and Ubp in more detail. In particular. In particular they will identify regions of the two proteins that are required for protein-protein interaction, isolate mutants that exhibit altered binding stability, and test the hypothesis that Vpu action is mediated via Ubp. Dr. Panganiban's preliminary data also indicated that Ubp interacts directly with Hiv-1 Gag protein in vitro, and with the capsid domain of HIV-1 Gag protein in human cells. Thus, he will identify the sites on Ubp and Gag that are required for this interaction. To examine the effect on Ubp on particle release, the effect of expression of wild type and mutant forms of Ubp and Gag that are required for this interaction. To examine the effect of Ubp on particle release, the effect of expression of wild type and mutant forms of Ubp on particle release will be determined. Similarly, they may determine whether ablation of endogenous Ubp expression affects the efficiency of particle release. Finally, since Ubp has not been previously characterized, Dr. Panganiban and his colleagues will examine the intrinsic properties of Ubp and try to gain insight into the normal role of Ubp in the cell. The overall goal of these studies to develop a comprehensive picture that accounts for the molecular mechanism by which Vpu mediates particle release.