疫苗病毒的DNA聚合酶全酶的结构和灵活性
Wim P Burmeister1, Laetitia Boutin2, Aurelia C Balestra1
1Institut de Biologie Structurale (IBS), Université Grenoble Alpes (UGA), Commissariat à l'Energie Atomique et aux Energies Alternatives (CEA), Centre National de la Recherche Scientifique (CNRS), Grenoble, France.
PLoS pathogens
|May 20, 2024
概括
研究人员特征了疫苗病毒 (VACV) DNA聚合酶全酶,对病毒DNA复制至关重要. 这项研究揭示了全酶.
科学领域:
- 结构生物学是结构生物学.
- 病毒学 病毒学
- 生物化学 生化学
背景情况:
- 2022年波克斯疫情突显了了解麻疹病毒 (MPXV) 和相关的骨麻疹病毒的重要性.
- 疫苗病毒 (VACV) 与MPXV具有显著的序列相同性,特别是在DNA复制蛋白中.
- 包含E9聚合酶和A20-D4异构体的VACV DNA聚合酶全酶对于病毒DNA复制至关重要.
研究的目的:
- 制造和确定VACV DNA聚合酶全酶的冷电子显微镜 (cryo-EM) 结构.
- 研究全酶复合体内的结构动态和相互作用.
- 了解E9-D4相互作用在病毒生存能力中的作用.
主要方法:
- 使用baculovirus-insect细胞系统生产VACV DNA聚合酶全酶.
- 低温电子显微镜 (cryo-EM) 用于确定无DNA全酶的3.8 Å结构.
- AlphaFold 2用于A20结构预测,与现有组件结构集成.
- 微角X射线散射 (SAXS) 用于研究溶液形状.
- 生物层干涉测量 (BLI) 来评估E9-D4相互作用强度.
主要成果:
- 冷-EM结构显示了E9-A20-D4全酶的紧布局,D4折叠在E9上,类似于MPXV全酶结构.
- 在全酶的背景下,E9指域变得混乱.
- 萨克斯数据表明,由于A20中的链区域,溶液中存在更开放,更灵活的形状.
- 在没有DNA的情况下,BLI证实了E9和D4之间的弱且短暂的相互作用.
- 在E9-D4接口的关键残留物中发生的突变使病毒无法生存,这凸显了它的重要性.
结论:
- VACV DNA聚合酶全酶采用一个紧的结构,由E9和D4.4之间保留的疏水界面稳定.
- 全酶表现出形状灵活性,允许在溶液中进行不同的安排.
- 微弱但至关重要的E9-D4相互作用对病毒复制和生存至关重要.
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