Association of HIV-1 gp120 with its coreceptors

超出所提供的空间。在这一继续提案中，我们寻求扩大和完善我们对HIV-1包膜糖蛋白及其辅助受体CCR5之间相互作用的理解。在上一个资助期内，我们证实了我们最初观察到的CCR5 n端酪氨酸硫酸化的存在及其在HIV-1进入中的关键作用。我们还证明了基于CCR5 N-末端序列的硫酸化肽可以阻断HIV-1进入人PBMC和巨噬细胞，并开始详细绘制HIV-1包膜糖蛋白中与CCR5 N-末端相互作用的区域。最后，我们记录了类似硫酸盐基团在CXCR4上的存在和作用。在这次更新中，将继续研究HIV-1包膜糖蛋白和HIV-1共受体之间关联的分子细节。为实现这一目标，提出了以下具体目标。在Specific Aim 1中，我们将进一步分析gp120区域与CCR5单个结构域的分子相互作用，使用我们开发的ig融合构建物和生物硫酸化CCR5肽。在Specific Aim 2中，我们将利用_NR5 （n端缺失的CCR5）和硫酸化的CCR5肽来研究HIV-1融合过程。在上一个资助期内，我们已经证明该_NR5本身不支持HIV-1的进入，但当与N硫酸盐化的CCR5肽互补时，它可以提供一个很好的系统来解剖复杂的HIV-1融合过程。在Specific Aim 3中，我们试图记录CCR5酪氨酸磺酸在HIV-1原发靶细胞中的水平和调控。我们还将研究酪氨酸硫酸化对CCR5内化和循环的贡献，以响应其趋化因子配体，这是趋化因子介导的HIV-1进入抑制的关键过程。网站性能 ======================================== 节结束 ===========================================

EXCEED THE SPACE PROVIDED. In this continuation proposal, we seek to expand and refine our understanding of the interaction between HIV-1 envelope glycoprotein and its coreceptor CCR5. During the last funding period, we confirmed our initial observation of the presence of tyrosine sulfation in the N-terminus of CCR5 and its critical role in HIV-1 entry. We also demonstrated that sulfated peptides based on the sequence of CCR5 N- terminus can block HIV-1 entry in human PBMC and macrophages and have begun to map in detail the regions in HIV-1 envelope glycoprotein that interact with CCR5 N-terminus. Finally, we have documented the presence and role of analogous sulfate moieties on CXCR4. In this renewal, will continue to investigate the molecular details of the association between HIV-1 envelope glycoprotein and the HIV-1 coreceptors. The following specific aims are offered to accomplish this goal. In Specific Aim 1, we will further analyze the molecular interactions of regions of gp120 with individual domains of CCR5, using the Ig-fusion constructs and biosulfated CCR5 peptides we have developed. In Specific Aim 2, we will examine the HIV-1 fusion process by utilizing _NR5 (N-terminal deleted CCR5) and sulfated CCR5 peptides. We have demonstrated during last funding period that this _NR5 does not support HIV-1 entry by itself, but does when complemented with N sulfated CCR5 peptides, offering an excellent system to dissect the complex HIV-1 fusion process. In Specific Aim 3, we seek to document the levels and regulation of CCR5's tyrosine sulfation in primary HIV-1 target cells. We will also study the contribution of tyrosine sulfation to the internalization and recycling of CCR5 in response to its chemokine ligands, a process criticalto chemokine mediated inhibition of HIV-1 entry. PERFORMANCE SITE ========================================Section End===========================================