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缓冲区的选择和pH强烈影响SARS-CoV-2核体与RNA相位分离

Nina C Kathe¹, Mihajlo Novakovic¹, Frédéric H-T Allain¹

¹Department of Biology, Institute of Biochemistry, ETH Zurich, Zurich, Switzerland.

Molecular biology of the cell

|April 3, 2024

概括

pH 和缓冲组合显著影响SARS-CoV-2核体 (N) 蛋白质液体-液体相分离 (LLPS). 伊斯蒂丁质子和静电相互作用是N蛋白LLPS的关键驱动因素,对于理解病毒复制至关重要.

科学领域:

生物化学生物化学
病毒学病毒学
分子生物学分子生物学

背景情况:

SARS-CoV-2 核体 (N) 蛋白对于病毒复制和基因组包装至关重要.
N蛋白经历液-液相分离 (LLPS) 形成生物分子凝聚物,这一过程对其功能至关重要.
控制N蛋白LLPS的精确因素仍然不完全理解.

研究的目的:

调查pH值和缓冲成分对SARS-CoV-2 N蛋白质的LLPS的影响.
阐明在N蛋白LLPS中的histidine质子和静电相互作用的作用.
突出实验条件在研究生物分子相位分离方面的重要性.

主要方法:

在不同pH条件下对SARS-CoV-2 N蛋白相分离的体外研究.
对希斯蒂丁残留质子状态的分析及其与LLPS的相关性.
评估与N蛋白和RNA的缓冲器特异性静电相互作用.

主要成果:

N蛋白质的LLPS对溶液pH高度敏感,与histidine质突相关.
在酸盐缓冲区中,对于LLPS而言,在位置356 (H356) 的histidine的质子化是必不可少的.
缓冲分子与N蛋白残留的静电相互作用改变了其净电荷,并影响了依赖RNA的相分离.

结论:

氨基酸质子和表面电荷的微妙变化,由pH和缓冲剂决定,深刻影响N蛋白LLPS.
电静电相互作用被确定为N蛋白相分离背后的主要驱动力.
像pH和缓冲值这样的实验参数是研究生物分子相位分离的关键考虑因素,特别是在改变细胞内环境的病毒感染中.

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