Reversible aggregation of HIV-1 Gag proteins mediated by nucleic acids.
Reversible aggregation of HIV-1 Gag proteins mediated by nucleic acids.
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DOI：
10.1016/j.bbrc.2016.12.054
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发表时间：
2017-01-22
影响因子：
3.1
通讯作者：
Cheng W
中科院分区：
文献类型：
--
作者：
Chen Z;Cheng W
HIV-1 Gag蛋白是病毒粒子组装的主要结构蛋白。虽然已经使用部分纯化的Gag蛋白来研究病毒颗粒组装的机制，但组装反应的结果仍然存在争议。在这里，我们已经开发了一种改进的程序，用于从大肠杆菌中纯化几种未标记的逆转录病毒Gag蛋白。大肠杆菌中纯化至超过95%的纯度，并表征了溶液中的Gag组装体。我们发现HIV-1 Gag蛋白可以发生核酸依赖性聚集，并具有几个意想不到的特征：（1）它们形成直径达微米的球形颗粒;（2）聚集体的大小随核酸与蛋白质的摩尔比而变化，这些颗粒的平均大小在核酸与蛋白质的摩尔比为1：2时达到最大;和（3）这些颗粒可以简单地在向溶液中加入过量核酸时有效地分解，表明存在有序组装。10个核苷酸或更短的单链DNA寡核苷酸不支持这些颗粒的形成。此外，Gag蛋白的基质结构域显著促进这些聚集体的形成。这些研究揭示了一个以前未知的途径，HIV Gag组装在体外，并在体内的HIV-1 Gag组装和发病机制的影响。
HIV-1 Gag protein is the major structural protein for the assembly of virion particles. Although studies have been carried out using partially purified Gag proteins to investigate the mechanisms of viral particle assembly, the outcomes of an assembly reaction remain controversial. Here we have developed an improved procedure for purification of several untagged retroviral Gag proteins from E. coli to more than 95% purity and characterized Gag assembly in solution. We found that HIV-1 Gag proteins can undergo nucleic acid-dependent aggregation with several unexpected features: (1) they form spherical particles that are as large as microns in diameter; (2) the size of the aggregates vary with the molar ratio between nucleic acids and proteins, with the average size of these particles reaching maximal at a molar ratio of 1:2 between nucleic acids and proteins; and (3) these particles can be efficiently disassembled simply upon addition of excess nucleic acids into the solution, suggesting the presence of an ordered assembly. Single-stranded DNA oligos that are 10 nucleotides or shorter do not support the formation of these particles. Furthermore, the matrix domain of the Gag protein dramatically facilitates the formation of these aggregates. These studies uncover a previously uncharacterized pathway of HIV Gag assembly in vitro, and have implications for HIV-1 Gag assembly and pathogenesis in vivo.