Binding Site Geometry and Subdomain Valency Control Effects of Neutralizing Lectins on HIV-1 Viral Particles
Binding Site Geometry and Subdomain Valency Control Effects of Neutralizing Lectins on HIV-1 Viral Particles
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DOI：
10.1021/acsinfecdis.6b00139
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发表时间：
2016-11-01
影响因子：
5.3
通讯作者：
Bewley, Carole A.
中科院分区：
文献类型：
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作者：
Lusvarghi, Sabrina;Lohith, Katheryn;Bewley, Carole A.
碳水化合物结合蛋白如griffithsin、cyanovirin-N和BanLec是有效的HIV进入抑制剂和有前途的杀微生物剂。每一种都与表面包膜糖蛋白gp 120上的高甘露糖聚糖结合，但它们参与病毒尖峰并表现出纳摩尔至皮摩尔范围内的抑制常数的机制尚不清楚。为了确定识别和效力的结构和机制基础，我们选择了一组具有不同化合价的凝集素，每个亚基，寡聚状态，和碳水化合物结合位点的相对取向，以系统地探测它们对抑制病毒进入的贡献。冷冻电子显微照片和免疫金染色的凝集素处理的病毒颗粒揭示了两个不同的效果，即，病毒聚集或聚类的HIV-1包膜上的病毒膜，所决定的碳水化合物结合位点的几何形状和价。“三明治”表面等离子体共振实验显示，第二个结合事件只发生在那些凝集素，可以聚集病毒颗粒。此外，观察到第二结合事件的皮摩尔K-d值，提供了实现皮摩尔IC 50值的机制。我们认为，这些结合和聚集现象转化为中和效力。
Carbohydrate binding proteins such as griffithsin, cyanovirin-N, and BanLec are potent HIV entry inhibitors and promising microbicides. Each binds to high-mannose glycans on the surface envelope glycoprotein gp120, yet the mechanisms by which they engage viral spikes and exhibit inhibition constants ranging from nanomolar to picomolar are not understood. To determine the structural and mechanistic basis for recognition and potency, we selected a panel of lectins possessing different valencies per subunit, oligomeric states, and relative orientations of carbohydrate binding sites to systematically probe their contributions to inhibiting viral entry. Cryo-electron micrographs and immuno gold staining of lectin-treated viral particles revealed two distinct effects namely, viral aggregation or clustering of the HIV-1 envelope on the viral membrane that were dictated by carbohydrate binding site geometry and valency. "Sandwich" surface plasmon resonance experiments revealed that a second binding event occurs only for those lectins that could aggregate viral particles. Furthermore, picomolar K-d values were observed for the second binding event, providing a mechanism by which picomolar IC50 values are achieved. We suggest that these binding and aggregation phenomena translate to neutralization potency.