Mutations in the poliovirus P1 capsid precursor at arginine residues VP4-ARG34, VP3-ARG223, and VP1-ARG129 affect virus assembly and encapsidation of genomic RNA.
Mutations in the poliovirus P1 capsid precursor at arginine residues VP4-ARG34, VP3-ARG223, and VP1-ARG129 affect virus assembly and encapsidation of genomic RNA.
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脊髓灰质炎病毒 P1 衣壳前体精氨酸残基 VP4-ARG34、VP3-ARG223 和 VP1-ARG129 处的突变会影响病毒组装和基因组 RNA 衣壳化。
DOI：
10.1006/viro.1994.1094
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发表时间：
1994
期刊：
影响因子：
3.7
通讯作者：
Morrow,CD
中科院分区：
文献类型：
--
作者：
Ansardi,DC;Luo,M;Morrow,CD
为了开始鉴定组装和 RNA 衣壳化所需的脊髓灰质炎病毒衣壳蛋白决定簇，我们研究了脊髓灰质炎病毒衣壳的三个精氨酸残基在病毒组装和基因组 RNA 衣壳化中的功能意义。这些研究是通过使用最近描述的系统进行的，其中重组牛痘病毒用于将脊髓灰质炎病毒衣壳蛋白转变成脊髓灰质炎病毒亚基因组复制子[D. C. Ansardi、D. C. Porter 和 C. D. Morrow (1993)J。病毒.67, 3684-3690]。两个精氨酸残基位于 VP4 的第 34 位 (VP4-R034) 和 VP1 的第 129 位 (VP1-R129)，位于脊髓灰质炎病毒衣壳内部的空腔内，而第三个精氨酸，即 VP3 的第 223 位残基 (VP3-R223)，位于原体-原体界面。通过脊髓灰质炎病毒 P1 衣壳前体 cDNA 的定点诱变构建了 5 个突变体，分别编码 VP4-R034 处的赖氨酸或谷氨酰胺取代（VP4-R034K、VP4-R034Q）、VP1 残基 129 处的赖氨酸或谷氨酰胺取代（VP1-R129K、VP1-R129Q）或赖氨酸取代在VP3 (VP3-R223K)的残基223处。源自 VP3-R223K、VP1-R129K 和 VP1-R129Q 突变体前体的加工衣壳蛋白不稳定，无法在 37° 下组装亚病毒颗粒或病毒体。 VP3-R223K前体裂解产物的组装缺陷在33°时被部分克服，因为在较低温度下由突变衣壳亚基组装成空衣壳，但不是成熟病毒粒子。对于分析的第三个精氨酸残基，VP4-R034，源自 VP4-R034K 和 VP4-R034Q 突变体前体的加工衣壳蛋白在 37° 组装 155S 病毒体，然而，源自 VP4-R034Q 前体的衣壳蛋白在 39.5° 下对病毒体形成温度敏感。 39.5°处病毒体形成的减少显然反映了形成组装能力亚基的缺陷，这也阻止了剩余VP4-R034Q亚基作为空衣壳的积累。通过使用图形显示脊髓灰质炎病毒的三维结构，这些残基在脊髓灰质炎病毒衣壳内部的位置及其与相邻氨基酸的相互作用被可视化，从而为观察到的组装缺陷提供结构解释，强调这些残基在衣壳组装和RNA衣壳化中发挥的重要作用。
To begin to identify poliovirus capsid protein determinants required for assembly and RNA encapsidation, we have addressed the functional significance of three arginine residues of the poliovirus capsid in virus assembly and encapsidation of genomic RNA. These studies were conducted by using a recently described system in which recombinant vaccinia viruses are used to supply poliovirus capsid proteinsin transto a poliovirus subgenomic replicon [D. C. Ansardi, D. C. Porter, and C. D. Morrow (1993)J. Virol.67, 3684-3690]. Two of the arginine residues, located at position 34 of VP4 (VP4-R034) and position 129 of VP1 (VP1-R129), are located within a cavity on the poliovirus capsid interior, whereas the third arginine, residue 223 of VP3 (VP3-R223), is located at a protomer-protomer interface. Five mutants were constructed by site-directed mutagenesis of poliovirus P1 capsid precursor cDNA to separately encode lysine or glutamine substitutions at VP4-R034 (VP4-R034K, VP4-R034Q), lysine or glutamine substitutions at residue 129 of VP1 (VP1-R129K, VP1-R129Q), or a lysine substitution at residue 223 of VP3 (VP3-R223K). Processed capsid proteins derived from the VP3-R223K, VP1-R129K, and VP1-R129Q mutant precursors were unstable and failed to assemble subviral particles or virions at 37°. The assembly defect for cleavage products of the VP3-R223K precursor was partially overcome at 33°, as empty capsids, but not mature virions, assembled from the mutant capsid subunits at the lower temperature. With regard to the third arginine residue analyzed, VP4-R034, processed capsid proteins derived from both the VP4-R034K and the VP4-R034Q mutant precursors assembled 155S virions at 37° however, capsid proteins derived from the VP4-R034Q precursor were temperature-sensitive for virion formation at 39.5°. The reduced virion formation at 39.5° was apparently a reflection of a defect in forming assembly competent subunits which also prevented accumulation of surplus VP4-R034Q subunits as empty capsids. By using graphics to display the poliovirus three-dimensional structure, the locations of these residues on the poliovirus capsid interior and their interactions with adjacent amino acids were visualized to provide structural explanations for the observed assembly defects which highlight the important role these residues play in capsid assembly and RNA encapsidation.