Laboratory of Immunoregulation and Bioinformatics and Computational Biosciences Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892.
Proc Natl Acad Sci U S A. 2014 Feb 25;111(8):3152-7. doi: 10.1073/pnas.1314718111. Epub 2014 Feb 3.
Elicitation of broadly neutralizing antibodies is essential for the development of a protective vaccine against HIV-1. However, the native HIV-1 envelope adopts a protected conformation that conceals highly conserved sites of vulnerability from antibody recognition. Although high-definition structures of the monomeric core of the envelope glycoprotein subunit gp120 and, more recently, of a stabilized soluble gp140 trimer have been solved, fundamental aspects related to the conformation and function of the native envelope remain unresolved. Here, we show that the conserved central region of the second variable loop (V2) of gp120 contains sulfated tyrosines (Tys173 and Tys177) that in the CD4-unbound prefusion state mediate intramolecular interaction between V2 and the conserved base of the third variable loop (V3), functionally mimicking sulfated tyrosines in CCR5 and anti-coreceptor-binding-site antibodies such as 412d. Recombinant gp120 expressed in continuous cell lines displays low constitutive levels of V2 tyrosine sulfation, which can be enhanced markedly by overexpression of the tyrosyl sulfotransferase TPST2. In contrast, virion-associated gp120 produced by primary CD4(+) T cells is inherently highly sulfated. Consistent with a functional role of the V2 sulfotyrosines, enhancement of tyrosine sulfation decreased binding and neutralization of HIV-1 BaL by monomeric soluble CD4, 412d, and anti-V3 antibodies and increased recognition by the trimer-preferring antibodies PG9, PG16, CH01, and PGT145. Conversely, inhibition of tyrosine sulfation increased sensitivity to soluble CD4, 412d, and anti-V3 antibodies and diminished recognition by trimer-preferring antibodies. These results identify the sulfotyrosine-mediated V2-V3 interaction as a critical constraint that stabilizes the native HIV-1 envelope trimer and modulates its sensitivity to neutralization.
诱导广谱中和抗体对于开发针对 HIV-1 的保护性疫苗至关重要。然而，天然的 HIV-1 包膜采用一种受保护的构象，使抗体无法识别高度保守的脆弱部位。尽管已经解决了单体核心包膜糖蛋白亚基 gp120 的高清晰度结构，以及最近稳定的可溶性 gp140 三聚体的高清晰度结构，但与天然包膜的构象和功能相关的基本方面仍未解决。在这里，我们表明 gp120 第二可变环（V2）的保守中心区域含有磺酸酪氨酸（Tys173 和 Tys177），在 CD4 未结合的预融合状态下，介导 V2 与第三可变环（V3）保守基础之间的分子内相互作用，在功能上模拟 CCR5 和抗核心受体结合位点抗体中的磺酸酪氨酸，如 412d。在连续细胞系中表达的重组 gp120 显示出低水平的 V2 酪氨酸磺酸化，通过过表达酪氨酸磺基转移酶 TPST2 可以显著增强。相比之下，由原代 CD4(+) T 细胞产生的病毒相关 gp120 固有地高度磺酸化。与 V2 磺酸酪氨酸的功能作用一致，增强酪氨酸磺酸化降低了单体可溶性 CD4、412d 和抗 V3 抗体对 HIV-1 BaL 的结合和中和作用，并增加了对三聚物偏好性抗体 PG9、PG16、CH01 和 PGT145 的识别。相反，抑制酪氨酸磺酸化增加了对可溶性 CD4、412d 和抗 V3 抗体的敏感性，并降低了对三聚物偏好性抗体的识别。这些结果确定了磺酸酪氨酸介导的 V2-V3 相互作用是稳定天然 HIV-1 包膜三聚体并调节其对中和的敏感性的关键约束。
