Association of HIV-1 gp120 with its coreceptors

超出提供的空间。在这个继续的提案中，我们试图扩大和完善我们对HIV-1包膜糖蛋白及其辅助受体CCR5之间相互作用的理解。在上一次资助期间，我们证实了我们初步观察到CCR5的N-末端存在酪氨酸硫化及其在HIV-1进入过程中的关键作用。我们还证明了基于CCR5 N末端序列的硫酸化多肽可以阻止HIV-1进入人PBMC和巨噬细胞，并且已经开始详细地定位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CXCR4上类似硫酸盐部分的存在和作用。在这次更新中，将继续研究HIV-1包膜糖蛋白和HIV-1辅助受体之间的联系的分子细节。为实现这一目标提供了以下具体目标。在具体目标1中，我们将利用我们开发的Ig融合构建体和生物硫化CCR5多肽，进一步分析gp120区域与CCR5各个结构域的分子相互作用。在具体目标2中，我们将利用_NR5(N-末端缺失的CCR5)和硫酸化的CCR5多肽来研究HIV-1的融合过程。在上一个资助期间，我们已经证明了This_NR5本身不支持HIV-1进入，但当与N-硫化CCR5肽互补时，它支持HIV-1进入，为剖析复杂的HIV-1融合过程提供了一个极好的系统。在具体目标3中，我们试图证明CCR5‘S酪氨酸硫化在原代HIV-1靶细胞中的水平和调节。我们还将研究酪氨酸硫化在CCR5内化和循环中的作用，以响应其趋化因子配体，这是趋化因子介导的抑制HIV-1进入的关键过程。表演网站========================================Section End===========================================

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===========================================