Mutation of the SP1 sequence impairs both multimerization and membrane-binding activities of human immunodeficiency virus type 1 Gag
Mutation of the SP1 sequence impairs both multimerization and membrane-binding activities of human immunodeficiency virus type 1 Gag
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DOI：
10.1128/jvi.79.3.1803-1812.2005
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发表时间：
2005-02-01
影响因子：
5.4
通讯作者：
Liang, C
中科院分区：
文献类型：
--
作者：
Guo, XF;Roldan, A;Liang, C
人类免疫缺陷病毒1型的Gag蛋白在衣壳和下游核衣壳序列之间包含一个14个氨基酸的区域，称为SP1。虽然已知SP1对于病毒产生是不可或缺的，但所涉及的机制大多不清楚。在这项研究中，我们证明了SP1内的M368 A突变严重降低了Gag与细胞膜结合的能力。虽然通过基质内的第二位点L20 K突变恢复了M368 A Gag的野生型膜结合水平，但所得Gag突变体L20 K-M368 A在病毒生产中仍然存在缺陷。后一种缺陷部分是由于L20 K-M368 A Gag与非筏膜的结合，而不是野生型Gag的筏结合。进一步的分析显示，大多数膜结合的M368 A Gag蛋白是小的寡聚体，表明多聚化缺陷。为了支持这一观察结果，含有M368 A突变的纯化重组Gag衍生物形成的高分子量复合物的量比野生型Gag低得多，这些复合物在21，000 x g下可形成颗粒。基于肉豆蔻基开关模型，我们提出M368 A突变抑制Gag多聚化，从而限制Gag与细胞膜的结合。
The Gag protein of human immunodeficiency virus type 1 contains a 14-amino-acid region, termed SP1, between the capsid and downstream nucleocapsid sequences. Although SP1 is known to be indispensable for virus production, the mechanisms involved are mostly unclear. In this study, we demonstrate that an M368A mutation within SP1 severely diminished the ability of Gag to associate with cellular membranes. Although wild-type levels of membrane binding were restored to the M368A Gag by a second-site L20K mutation within matrix, the resultant Gag mutant L20K-M368A remained defective in virus production. This latter deficit was partially consequent to the binding of L20K-M368A Gag to nonraft membranes as opposed to raft association seen for wild-type Gag. Further analysis revealed that the majority of membrane-bound M368A Gag proteins were small oligomers, indicating a multimerization defect. In support of this observation, purified recombinant Gag derivatives containing the M368A mutation formed much lower amounts of high-molecular-weight complexes that were pelletable at 21,000 x g than did wild-type Gag. Based on the myristyl switch model, we propose that the M368A mutation inhibits Gag multimerization and, as a result, restricts the binding of Gag to cellular membranes.