Biophysical Signatures in HIV-1 Env Correlating with Mother-to-Child Transmission

描述：艾滋病毒的特征之一是其极端的变异性，这导致感染后病毒准种的广泛多样化。在粘膜传播过程中，病毒也经历了相反的现象，经历了基因多样性的严格缩小，使得大多数新感染是由准种的单一传播的“创始人”(T/F)变体建立的。传播的物理决定因素仍然没有明确的定义。虽然许多研究将T/F特征追溯到Env，即介导感染的表面糖蛋白，但大多数研究都研究了潜在差异的生物学后果，但无法明确区分不同的Env，因为经典方法不适合分析糖蛋白的结构变化。对结晶学的要求使得有必要截断和去糖基化糖蛋白，消除使每个变体与众不同的可变环和糖基化等特征，这些特征可以调节核心保守区域的结构。我们建议应用新的生物物理和结构方法来比较来自HIV T/F变种和来自供体变种的Env糖蛋白的结构，使用在一项具有良好特征的母婴传播(MTCT)研究中确定的传递对。我们发展了利用氢/氢交换质谱仪(HDX-MS)来探测天然条件下完整的糖蛋白结构。这种方法提供了识别变异体特异性差异所需的灵敏度和分辨率，产生了整个糖蛋白局部结构顺序和稳定性的指纹。在目标1中，我们将应用HDX-MS从内罗毕母乳喂养母婴传播研究中确定的匹配传播对中分析环境病毒。传递对提供了一种强大的手段来精确定位序列差异，这些差异可能与T/F变异体的传递相关的结构和表型差异有关。我们假设T/F环境中可能存在结构属性，包括赋予T/F变体选择性优势的更大稳定性。我们预计，更好的环境稳定性可能会导致复合体在AIM 2中抵抗灭活，我们将使用感染性分析来测试T/F变体是否具有增强的抗热灭活和低pH和乳酸灭活的能力，这些因素可能存在于传播部位，如婴儿的胃肠道。Lee和Overbaugh实验室的结构生物学和病毒学专业知识的结合应用于检查匹配的传播对，可以为病毒传播性的物理决定因素提供重要的新见解，并提供与疫苗免疫原设计相关的结构细节。

DESCRIPTION: One of the hallmarks of HIV is its extreme mutability, which leads to broad diversification of the viral quasispecies following infection. During mucosal transmission, the virus experiences the opposite phenomenon as well, undergoing a stringent narrowing of genotypic diversity, such that the majority of new infections are established by a single transmitted, "founder" (T/F) variant from the quasispecies. The physical determinants of transmission remain poorly defined. While many studies have traced T/F signature traits to Env, the surface glycoprotein that mediates infection, most have examined the biological consequences of underlying differences without being able to address what specifically differentiates one Env from another because classical methods are poorly suited to analyze structural variation of glycoproteins. The requirements for crystallography make it necessary to truncate and deglycosylate the glycoprotein, removing the very features such as variable loops and glycosylation that make each variant distinct and that can modulate structure within core, conserved regions. We propose to apply novel biophysical and structural approaches to compare the structure of Env glycoproteins from T/F variants of HIV with Env from donor variants using transmission pairs identified in a well-characterized mother-to-child-transmission (MTCT) study. We have developed the use of hydrogen/deuterium-exchange mass spectrometry (HDX-MS) to probe intact glycoprotein structure under native conditions. This approach provides the sensitivity and resolution necessary to identify variant-specific differences, producing a fingerprint of local structural order and stability throughout the glycoprotein. In AIM 1, we will apply HDX-MS to analyze Env from matched transmission pairs identified in a Nairobi Breastfeeding MTCT study. The transmission pairs provide a powerful means to pinpoint sequence differences that may be linked with structural and phenotypic differences relevant to transmission of T/F variants. We hypothesize that structural properties in T/F Env may exist including greater stability that confer a selective advantage to T/F variants. We anticipate that greater Env stability may result in complexes that are resistant to inactivation In AIM 2, using infectivity assays, we will test whether T/F variants exhibit enhanced resistance to heat inactivation and inactivation by low pH and lactic acid, factors that may be present at sites of transmission such as the infant gastrointenstinal tract. The combination of structural biology and virological expertise in the Lee and Overbaugh labs applied to examine the matched transmission pairs can provide significant new insights into the physical determinants of virus transmissibility and provide structural details relevant to vaccine immunogen design.