Characterization of the R572T point mutant of a putative cleavage site in human foamy virus Env
Characterization of the R572T point mutant of a putative cleavage site in human foamy virus Env
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DOI：
10.1128/jvi.74.6.2949-2954.2000
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发表时间：
2000-03-01
影响因子：
5.4
通讯作者：
Mulligan, MJ
中科院分区：
文献类型：
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作者：
Bansal, A;Shaw, KL;Mulligan, MJ
人泡沫病毒（HFV）包膜糖蛋白（Env）的一个假定切割位点发生了改变。瞬时env表达表明，R572 T突变体Env正常表达和修饰的天冬酰胺连接的寡糖链。然而，这种单氨基酸取代足以消除所有可检测到的裂解的gp 130前体多蛋白。细胞表面的生物素化表明，未切割的突变体gp 130被运输到质膜。未切割的突变蛋白不能形成合胞体。糖蛋白驱动的病毒体出芽，HFV装配的一个独特方面，发生尽管缺乏Env切割。我们然后将R572 T突变体env替换成具有复制能力的HFV分子克隆。将突变病毒DNA转染到BHK-21细胞中，然后用FAB进行病毒滴定（泡沫病毒激活的β-半乳糖苷酶表达）试验表明，HFV Env的蛋白酶解是病毒感染性所必需的，野生型HFV Env部分补充了缺陷型病毒的表型。切割对Env运输、加工和功能的影响;以及Env蛋白水解对病毒成熟和感染性的重要性。
A putative cleavage site of the human foamy virus (HFV) envelope glycoprotein (Env) was altered. Transient env expression revealed that the R572T mutant Env was normally expressed and modified by asparagine-linked oligosaccharide chains. However, this single-amino-acid substitution was sufficient to abolish all detectable cleavage of the gp130 precursor polyprotein. Cell surface biotinylation demonstrated that the uncleaved mutant gp130 was transported to the plasma membrane. The uncleaved mutant protein was incapable of syncytium formation. Glycoprotein-driven virion budding, a unique aspect of HFV assembly, occurred despite the absence of Env cleavage, We then substituted the R572T mutant env into a replication-competent HFV molecular clone. Transfection of the mutant viral DNA into BHK-21 cells followed by viral titration with the FAB (foamy virus-activated beta-galactosidase expression) assay revealed that proteolysis of the HFV Env was essential for viral infectivity, Wild-type HFV Env partially complemented the defective virus phenotype, Taken together, these experimental results established the Location of the HFV Env proteolytic site; the effects of cleavage on Env transport, processing, and function; and the importance of Env proteolysis for virus maturation and infectivity.