MAPPING OF FUNCTIONALLY IMPORTANT RESIDUES OF A CYSTEINE-HISTIDINE BOX IN THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NUCLEOCAPSID PROTEIN
MAPPING OF FUNCTIONALLY IMPORTANT RESIDUES OF A CYSTEINE-HISTIDINE BOX IN THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NUCLEOCAPSID PROTEIN
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DOI：
10.1128/jvi.67.10.6159-6169.1993
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发表时间：
1993-10-01
影响因子：
5.4
通讯作者：
GOTTLINGER, HG
中科院分区：
文献类型：
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作者：
DORFMAN, T;LUBAN, J;GOTTLINGER, HG
人类免疫缺陷病毒 1 型 (HIV-1) 核衣壳蛋白含有两个拷贝的序列基序，即半胱氨酸-组氨酸盒，在逆转录病毒中是保守的。为了鉴定半胱氨酸-组氨酸盒的功能相关位置，通过定点诱变单独替换基序近端拷贝中的每个氨基酸。 14 个位置中的 5 个位置的突变消除了病毒复制，并将突变颗粒的病毒 RNA 含量降低至亲代水平的 10% 至 20%。其他位置的突变对病毒复制和病毒粒子 RNA 含量没有影响或影响很小。 RNA 与细菌表达的突变体 Pr55gag 多蛋白的体外结合与突变对颗粒相关病毒 RNA 水平的影响密切相关。 HIV-1核衣壳蛋白中基序的两个不同拷贝在功能上并不等同，因为近端基序转换为远端基序的精确拷贝会导致病毒复制缺陷和突变体颗粒的病毒RNA含量减少。两个基序中功能相关位置的同时替换导致gag蛋白输出显着下降，表明gag前体的核衣壳结构域也是病毒粒子的有效组装或释放所必需的。
The human immunodeficiency virus type 1 (HIV-1) nucleocapsid protein contains two copies of a sequence motif, the cysteine-histidine box, that is conserved among retroviruses. To identify the functionally relevant positions of a cysteine-histidine box, each amino acid in the proximal copy of the motif was individually substituted by site-directed mutagenesis. Mutations at 5 of 14 positions abolished virus replication and reduced the viral RNA content of mutant particles to between 10 and 20% of parental levels. Mutations at other positions had either no or only a minor effect on virus replication and virion RNA content. In vitro binding of RNA to bacterially expressed mutant Pr55gag polyprotein correlated well with the effects of the mutations on particle-associated viral RNA levels. The two different copies of the motif in the HIV-1 nucleocapsid protein are not functionally equivalent, since the conversion of the proximal motif to an exact copy of the distal motif results in a defect in virus replication and a reduction in the viral RNA content of mutant particles. The simultaneous substitution of functionally relevant positions in both motifs led to a significant decline in gag protein export, indicating that the nucleocapsid domain of the gag precursor is also required for efficient assembly or release of the virion.