Highly purified human immunodeficiency virus type 1 reveals a virtual absence of vif in virions
Highly purified human immunodeficiency virus type 1 reveals a virtual absence of vif in virions
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DOI：
10.1128/jvi.73.2.1460-1467.1999
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发表时间：
1999-02-01
影响因子：
5.4
通讯作者：
Yu, XF
中科院分区：
文献类型：
--
作者：
Dettenhofer, M;Yu, XF
人类免疫缺陷病毒1型（HIV-1）的vif基因对于原代血液来源的淋巴细胞、巨噬细胞和某些人类T细胞系的生产性感染是必不可少的。已经表明，Vif与通过蔗糖密度平衡梯度分析纯化的HIV-1病毒体相关。然而，Vif掺入病毒体的特异性尚未确定。此外，最近的研究表明，用蔗糖密度平衡梯度制备的标准HIV-1颗粒制剂被细胞衍生的微泡污染。在这里，我们证明，如前所述，Vif cosediments与HIV-1颗粒在蔗糖密度平衡梯度分析。然而，我们还发现，当Vif在所有其他HIV-1编码的基因产物的情况下表达，然后通过蔗糖密度平衡梯度离心从细胞外上清液中分离时，其沉降模式在很大程度上没有改变，这表明Vif可以从细胞中分泌。使用新开发的OptiPrep速度梯度方法，我们能够将大部分细胞外Vif与HIV-1病毒粒子物理分离，而不会破坏病毒的感染性。通过针对OptiPrep梯度中的病毒峰级分滴定纯化的Vif和Gag的系列稀释液，我们证明
The vif gene of human immunodeficiency virus type 1 (HIV-1) is essential for the productive infection of primary blood-derived lymphocytes, macrophages, and certain human T-cell lines. It has been shown that Vif is associated with HIV-1 virions purified by sucrose density-equilibrium gradient analysis. However, the specificity of Vif incorporation into virions has not been determined. Moreover, recent studies have demonstrated that standard HIV-1 particle preparations created with sucrose density-equilibrium gradients are contaminated with cell-derived microvesicles. Here we demonstrate, as previously reported, that Vif cosediments with HIV-1 particles in sucrose density-equilibrium gradient analysis. However, we also found that, when Vif was expressed in the absence of all other HIV-1-encoded gene products and then isolated by sucrose density-equilibrium gradient centrifugation from extracellular supernatants, its sedimentation pattern was largely unaltered, suggesting that Vif can be secreted from cells. Using a newly developed OptiPrep velocity gradient method, we were able to physically separate most of the extracellular Vif from the HIV-1 virions without disrupting the infectivity of the virus. By titrating serial dilutions of purified Vif and Gag against the viral peak fraction in the OptiPrep gradient, we demonstrate that