<br>麻疹病毒磷蛋白 X 结构域与大聚合酶蛋白的二分界面调节病毒聚合酶动力学。,PLoS Pathogens

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麻疹病毒磷蛋白 X 结构域与大聚合酶蛋白的二分界面调节病毒聚合酶动力学。
PLoS Pathogens ( IF 5.5 ) Pub Date : 2019-08-05 , DOI: 10.1371/journal.ppat.1007995
Venice Du Pont ₁ , Yi Jiang ₂ , Richard K Plemper ₁

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麻疹病毒（MeV）是一种具有高度传染性、重新出现的主要人类病原体。复制需要病毒 RNA 依赖性 RNA 聚合酶（RdRP），该聚合酶由大（L）聚合酶蛋白与同型四聚体磷蛋白（P）复合组成。此外，P 介导与核蛋白（N）包覆的病毒 RNA 基因组的相互作用。P：L 界面的性质和核糖核蛋白模板的 RdRP 协商知之甚少。基于生化界面映射、将中央 P 四聚化结构域（OD）交换为酵母 GCN4 和功能测定，我们证明 MeV P-to-L 界面是二分的，包括靠近 OD 的卷曲螺旋微结构域和三角棱柱形 C 末端 P X 结构域（P-XD）的未占据面，这与已知的结合 N 尾的 P-XD 面不同。混合的零突变 P 四聚体以比率依赖性方式恢复了 L 结合能力，并在 minireplicon 测定和重组 MeV 中完全回收了生物活性，表明单个 L 结合界面元件在物理和机制上是不同的。P-XD 与 L 和 N 的结合能力同样是反式互补的，这与数学建模相结合，能够通过化学计量和化学计量控制的三元互补来表征 P 的机理表征。每个 P 四聚体的四种 XD 中只有一个必须具有 L 或 N 生物活性的结合活性，但同一 P-XD 与 L 和 N 的相互作用是互斥的，并且 L 结合取代了参与 N。混合的 P 四聚体与单个指定的 L 结合感受态 P-XD 会导致显著的 RdRP 过度活跃，概述了 P-XD 的迭代解析和重组模型：L 界面调节聚合酶迁移率。

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Bipartite interface of the measles virus phosphoprotein X domain with the large polymerase protein regulates viral polymerase dynamics.

Measles virus (MeV) is a highly contagious, re-emerging, major human pathogen. Replication requires a viral RNA-dependent RNA polymerase (RdRP) consisting of the large (L) polymerase protein complexed with the homo-tetrameric phosphoprotein (P). In addition, P mediates interaction with the nucleoprotein (N)-encapsidated viral RNA genome. The nature of the P:L interface and RdRP negotiation of the ribonucleoprotein template are poorly understood. Based on biochemical interface mapping, swapping of the central P tetramerization domain (OD) for yeast GCN4, and functional assays, we demonstrate that the MeV P-to-L interface is bipartite, comprising a coiled-coil microdomain proximal to the OD and an unoccupied face of the triangular prism-shaped C-terminal P X-domain (P-XD), which is distinct from the known P-XD face that binds N-tail. Mixed null-mutant P tetramers regained L-binding competence in a ratio-dependent manner and fully reclaimed bioactivity in minireplicon assays and recombinant MeV, demonstrating that the individual L-binding interface elements are physically and mechanistically distinct. P-XD binding competence to L and N was likewise trans-complementable, which, combined with mathematical modeling, enabled the mechanistic characterization of P through two- and stoichiometrically-controlled three-way complementations. Only one each of the four XDs per P tetramer must be L or N binding-competent for bioactivity, but interaction of the same P-XD with L and N was mutually exclusive, and L binding superseded engaging N. Mixed P tetramers with a single, designated L binding-competent P-XD caused significant RdRP hyperactivity, outlining a model of iterative resolution and reformation of the P-XD:L interface regulating polymerase mobility.

更新日期：2019-08-05

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