Structure, interactions and dynamics of PRD1 virus I. Coupling of subunit folding and capsid assembly.
Structure, interactions and dynamics of PRD1 virus I. Coupling of subunit folding and capsid assembly.
复制标题
PRD1 病毒的结构、相互作用和动力学 I。亚基折叠和衣壳组装的耦合。
DOI：
10.1006/jmbi.1996.0149
复制
发表时间：
1996
影响因子：
5.6
通讯作者：
ThomasJr,GJ
中科院分区：
文献类型：
--
作者：
Tuma,R;Bamford,JH;Bamford,DH;Russell,MP;ThomasJr,GJ
噬菌体 PRD1 感染大肠杆菌和鼠伤寒沙门氏菌，由二十面体衣壳组成，其中包裹着膜包老化的双链 DNA 基因组。使用时间和温度分辨拉曼光谱和紫外共振拉曼光谱对病毒外壳进行了研究，揭示了衣壳结构的新特征及其从 P3 亚基组装的途径。拉曼光谱显示，壳的热稳定性可达 50°C，并在 50 至 70°C 之间分解，P3 构象仅发生很小的变化。然而，热分解产物敏感地取决于总蛋白浓度。分析超速离心表征表明，低于 8 mg/ml，纯化的壳主要分解成 P3 三聚体；浓度较高时，会形成较大的 P3 多聚体。 P3 壳的盐酸胍 (GuHCl) 解离产生类似的结果。通过加热或 GuHCl 处理分离的纯化 P3 三聚体在 30 至 50°C 之间表现出结构敏感性。因此，壳的分解会降低 P3 的热稳定性。三聚体的低温转变（30°C 至 50°C）和壳的高温转变（50°C 至 70°C）都涉及约 5% 的 P3 肽主链从 α 螺旋转变为 β 链。 P3 肽主链的氘交换显示壳中的交换比三聚体中的交换更快，这与观察到的高浓度三聚体的非特异性聚合一致。相反，吲哚 1NH 基团的交换表明，约 65% 的色氨酸残基在组装的壳中受到保护，免于交换。结果表明了一种壳组装机制，其中三聚体与正确的壳结构的特定关联涉及亚基 α-螺旋结构域的稳定以及所选侧链与溶剂接触的隔离。我们提出了一个衣壳组装模型，它将 P3 壳的形成与 P3 亚基折叠的最后步骤结合起来。
BacteriophagePRD1, which infectsEscherichia coliandSalmonella typhimurium, consists of an icosahedral capsid enclosing a membrane-pack aged double-stranded DNA genome. The viral shell has been investigated using time and temperature resolved Raman and ultraviolet-resonance Raman spectroscopy to reveal novel features of the capsid structure and its pathway of assembly from P3 subunits. Raman spectra show that the shell is thermostable to 50°C, and disassembles between 50 and 70°C with only a small change in P3 conformation. However, the products of thermal dis- assembly depend sensitively upon total protein concentration. Charac- terization by analytical ultracentrifugation indicates that below 8 mg/ml, the purified shell dissassembles primarily into P3 trimers; at higher concen- trations, larger multimers of P3 are formed. Guanidine hydrochloride (GuHCl) dissociation of the P3 shell yields similar results. Purified P3 trimers, isolated either by heat or GuHCl treatment, exhibit structure sensitivity between 30 and 50°C. Thus, shell disassembly diminishes P3 thermostability. Both the lower temperature transition (30°C to 50°C) of the trimer and the higher temperature transition (50°C to 70°C) of the shell involve a conversion of ≈5% of the P3 peptide backbone from α-helix to β-strand. Deuterium exchange of the P3 peptide backbone reveals more rapid exchange in the shell than in the trimer, consistent with the observed non-specific polymerization of trimers at high concentration. Conversely, the exchange of indole 1NH groups shows that ≈65% of tryptophan residues are protected against exchange in the assembled shell. The results suggest a mechanism for shell assembly in which the specific association of trimers into the correct shell architecture involves stabilization of a subunit α-helical domain and sequestering of selected side-chains from solvent access. We propose a capsid assembly model which couples P3 shell formation with the final step in folding of the P3 subunit.