Saturation Mutagenesis of the HIV-1 Envelope CD4 Binding Loop Reveals Residues Controlling Distinct Trimer Conformations.

The conformation of HIV-1 envelope (Env) glycoprotein trimers is key in ensuring protection against waves of neutralizing antibodies generated during infection, while maintaining sufficient exposure of the CD4 binding site (CD4bs) for viral entry. The CD4 binding loop on Env is an early contact site for CD4 while penetration of a proximal cavity by CD4 triggers Env conformational changes for entry. The role of residues in the CD4 binding loop in regulating the conformation of the trimer and trimer association domain (TAD) was investigated using a novel saturation mutagenesis approach. Single mutations identified, resulted in distinct trimer conformations affecting CD4bs exposure, the glycan shield and the TAD across diverse HIV-1 clades. Importantly, mutations that improve access to the CD4bs without exposing the immunodominant V3 loop were identified. The different trimer conformations identified will affect the specificity and breadth of nabs elicited in vivo and are important to consider in design of Env immunogens for vaccines. Spike proteins on the surface of HIV virus particles bind to CD4 receptors on the surface of immune cells and trigger infection. The immune system in an infected person attacks the virus spikes by producing antibodies that bind and neutralize them. To combat this immune attack, HIV continually alters the structure of the spike and thus escapes host antibodies. However, this process must still preserve sites on the spike that bind CD4 receptors for infection. Here, we investigated how the spike regulates its structure. We used a systematic approach to investigate every possible mutation covering a region of the spike critical for binding the CD4 receptor and controlling overall structure. We identified different sites and mechanisms that control the spike structure for diverse HIV-1 strains and impact the exposure of the binding site for CD4 along with targets for neutralizing antibodies. Our observations will help guide the design of spike structures for vaccines that induce neutralizing antibodies effective against different HIV-1 strains across the globe.

HIV - 1包膜（Env）糖蛋白三聚体的构象对于确保在感染期间抵御产生的中和抗体波至关重要，同时要保持CD4结合位点（CD4bs）有足够的暴露以利于病毒进入。Env上的CD4结合环是与CD4的早期接触位点，而CD4对近端腔的穿透会触发Env的构象变化以利于进入。利用一种新的饱和诱变方法研究了CD4结合环中的残基在调节三聚体和三聚体结合域（TAD）构象中的作用。所鉴定出的单个突变导致了不同的三聚体构象，影响了不同HIV - 1分支中CD4bs的暴露、聚糖屏蔽和TAD。重要的是，鉴定出了在不暴露免疫优势V3环的情况下改善对CD4bs可及性的突变。所鉴定出的不同三聚体构象将影响体内引发的中和抗体的特异性和广度，并且在设计用于疫苗的Env免疫原时需要加以考虑。 HIV病毒颗粒表面的刺突蛋白与免疫细胞表面的CD4受体结合并引发感染。感染者的免疫系统通过产生结合并中和病毒刺突的抗体来攻击病毒刺突。为了对抗这种免疫攻击，HIV不断改变刺突的结构从而逃避宿主抗体。然而，这个过程仍必须保留刺突上与CD4受体结合以引发感染的位点。在此，我们研究了刺突如何调节其结构。我们采用一种系统的方法研究了覆盖刺突上对结合CD4受体和控制整体结构至关重要的一个区域的每一种可能的突变。我们鉴定出了不同的位点和机制，它们控制着不同HIV - 1毒株的刺突结构，并影响CD4结合位点的暴露以及中和抗体的作用靶点。我们的观察结果将有助于指导疫苗刺突结构的设计，这些结构可诱导出能有效中和全球不同HIV - 1毒株的中和抗体。