Comparative Glycomics of the HIV Protein gp 120

描述(申请人提供)：糖蛋白gp120覆盖HIV病毒表面。它在HIV感染中起着至关重要的作用，因为gp120结合到体内的受体位置，作为病毒入侵宿主细胞的第一步。由于病毒在不进入宿主细胞的情况下无法增殖，因此针对治愈艾滋病的研究的主要重点是阻止最初的细胞入侵。在细胞入侵之前摧毁病毒的策略尚未实现，因为gp120上的碳水化合物提供了抗体攻击的屏障。虽然HIV病毒上存在的多糖已被确定为病毒避免被免疫系统破坏的主要原因，但几乎没有办法克服病毒的防御机制，即它的糖蛋白屏障，因为人们对蛋白质上24个糖基化位点上存在的碳水化合物知之甚少。我们已经确定了更好地理解病毒防御机制所需的关键信息：1)每个糖基化位点的碳水化合物组成是什么？2)这些位点的拓扑结构(分支程度)是什么？3)由于碳水化合物带负电荷，蛋白质表面哪里有可能发生静电相互作用？为了回答这些问题，我们将使用先进的质谱学仪器来探索糖肽结构，包括MALDI TOF-TOF质谱仪和线性陷阱-傅里叶变换离子回旋共振质谱仪(LTQ-FTMS)。我们正在开发新的质谱学方法来探索gp120中带负电荷的糖形式的位置，以及识别糖肽拓扑的方法。此外，通过比较这些因素在不同的gp120分离株中的变化，我们可以确定糖蛋白的哪些部分在不同的gp120分离株中表现出最小的碳水化合物含量变化趋势。这些信息将直接影响gp120的结构-功能模型，并为未来的疫苗靶点提供方向。该项目的合作者包括威奇托州立大学的糖生物学家George Bousfield博士，以及堪萨斯大学蛋白质组学设施的联合主任Mike Alterman博士和Todd Williams博士。

DESCRIPTION (provided by applicant): The glycoprotein, gp120, covers the surface of the HIV virus. It plays an essential role in HIV infection, because gp120 binds to receptor sites in the body, as the first step of viral invasion into host cells. Since the virus cannot proliferate without entering host cells, a major thrust of research directed at curing AIDS has focused on stopping the initial cellular invasion. A strategy to destroy the virus prior to cellular invasion has not yet been achieved, because the carbohydrates on gp120 present a barrier to antibody attack. While the glycans present on the HIV virus have been identified as the primary reason that the virus avoids destruction by the immune system, little can be done to overcome the virus's defense mechanism, its glycan shield, because not enough is known about the carbohydrates present at each of the 24 glycosylation sites on the protein. We have identified critical information needed to better understand the virus's defense mechanism: 1) What is the carbohydrate composition at each glycosylation site? 2) What is the topology (the degree of branching) at these sites? 3) Where is there a potential for electrostatic interaction, due to negatively charged carbohydrates, on the surface of the protein? To answer these questions, we will probe glycopeptide structure using advanced mass spectrometric instrumentation, including a MALDI TOF-TOF mass spectrometer and Linear Trap-Fourier Transform Ion Cyclotron Resonance Mass Spectrometer (LTQ-FTMS). We are developing new mass spectrometric methods to probe the location of negatively charged glycoforms within gp120, as well as methods to identify glycopeptide topology. Furthermore, by comparing how each of these factors change from different gp120 isolates, we can determine which portions of the glycoprotein show the least propensity for variation in the carbohydrate content, among the different gp120 isolates. This information will directly impact both structure-function models of gp120, as well as provide direction for future vaccine targets. Collaborators on the project include Dr. George Bousfield, a glycobiologist from Wichita State University, and Drs. Mike Alterman and Todd Williams, Co-Directors of the University of Kansas Proteomics Facility.