Assembly-defective point mutants of the human immunodeficiency virus type 1 Gag precursor phenotypically expressed in recombinant baculovirus-infected cells
Assembly-defective point mutants of the human immunodeficiency virus type 1 Gag precursor phenotypically expressed in recombinant baculovirus-infected cells
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在重组杆状病毒感染细胞中表型表达的 1 型人类免疫缺陷病毒 Gag 前体的装配缺陷点突变体
DOI：
10.1128/jvi.67.5.2787-2798.1993
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发表时间：
1993
影响因子：
5.4
通讯作者：
P. Boulanger
中科院分区：
文献类型：
--
作者：
S. S. Hong;P. Boulanger
用重组杆状病毒进行亚硝酸盐诱变后，分离出了人类免疫缺陷病毒1型gag基因产物的两个替代突变体，该重组杆状病毒表达了非n-豆芽化、p6缺失的gag前体（Pr49）。这两个突变体都不能像亲本重组体那样在细胞内组装Gag病毒样颗粒，因此在昆虫细胞中表达了一种自组装缺陷（Sad）表型。突变包括p24结构域高度保守的疏水残基、268位的Leu到Pro （L268P）和322位的Leu到Ser （L322S）的非保守性变化。实验数据表明，这两个突变残基属于Gag前体的功能不同区域。(i)在共感染实验中观察到两个突变体在Gag前体组装上存在部分互补效应。（ii）两种突变在n - myristyated环境下表现出不同的表型，其中只有L268P突变使胞外出芽和Gag颗粒在质膜上释放消失。L268P和L322S突变体对共感染重组表达的非n-肉芽化全长（Pr55） Gag前体的细胞内组装具有反显性负作用。然而，没有一个突变体显示出反式对共表达的n -肉芽化野生型Gag前体的膜靶向和出芽有任何可检测的影响。
Two substitution mutants of the human immunodeficiency virus type 1 gag gene product were isolated after nitrous acid mutagenesis of a recombinant baculovirus expressing a non-N-myristylated, p6-deleted Gag precursor (Pr49). Both mutants failed to assemble intracellular Gag virus-like particles, as does the parental recombinant, and therefore expressed a self-assembly defective (Sad) phenotype in insect cells. The mutations consisted of nonconservative changes involving highly conserved hydrophobic residues in the p24 domain, Leu to Pro at position 268 (L268P) and Leu to Ser at amino acid 322 (L322S). Experimental data suggested that the two mutated residues belonged to functionally different regions of the Gag precursor. (i) A partial complementation effect between the two mutants for Gag precursor assembly was observed in coinfection experiments. (ii) The two mutations showed different phenotypes when placed in the N-myristylated context, of which only the L268P mutation abolished extracellular budding and release of Gag particles at the plasma membrane. Both L268P and L322S mutants had a trans-dominant negative effect on the intracellular assembly of a non-N-myristylated, full-length (Pr55) Gag precursor expressed by a coinfecting recombinant. None of the mutants, however, showed any detectable effect in trans on membrane targeting and budding of the coexpressed N-myristylated wild-type Gag precursor.