Reconstruction and validation of entire virus model with complete genome from mixed resolution cryo-EM density.
Reconstruction and validation of entire virus model with complete genome from mixed resolution cryo-EM density.
复制标题
通过混合分辨率冷冻电镜密度重建和验证具有完整基因组的整个病毒模型。
DOI：
10.1039/d2fd00053a
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发表时间：
2022
影响因子：
3.4
通讯作者：
Farafonov VS
中科院分区：
文献类型：
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作者：
Farafonov VS
从实验数据中很难计算出一个大的生物分子复合体的整个生物结构。由此产生的原子论模型不应该包含结构上的间隙，并且它应该产生稳定的动力学。我们首次从公布的不完全冷冻-EM密度重建了一个原子分辨率的完整MS2病毒，也就是基因组的衣壳，并用显式水的全原子分子动力学验证了结果。可用的实验数据包括高分辨率的蛋白质衣壳和不均匀分辨率的基因组图。对于基因组RNA，除了16个发夹具有原子性分辨外，衣壳内表面附近的链被分辨到核骨架水平，而最里面的密度完全没有分辨。结果，只有242个核苷酸被定位(3569个核苷酸中)，而基因组的其余部分只勾勒出一个碎裂的骨架，因此有必要进行详细的模型重建。对于模型重建，除了可用的原子结构信息外，我们还广泛使用了预测的基因组二级结构(碱基配对)。这项技术是基于半自动构建相对较大的RNA链，然后在被追踪的主干上进行人工定位。整个病毒结构(衣壳+基因组)通过在标准条件下与离子一起在生理溶液中运行的分子动力学来验证，证实了模型的稳定性。
It is very difficult to reconstruct computationally a large biomolecular complex in its biological entirety from experimental data. The resulting atomistic model should not contain gaps structurally and it should yield stable dynamics. We, for the first time, reconstruct from the published incomplete cryo-EM density a complete MS2 virus at atomistic resolution, that is, the capsid with the genome, and validate the result by all-atom molecular dynamics with explicit water. The available experimental data includes a high resolution protein capsid and an inhomogeneously resolved genome map. For the genomic RNA, apart from 16 hairpins with atomistic resolution, the strands near the capsid’s inner surface were resolved up to the nucleic backbone level, and the innermost density was completely unresolved. As a result, only 242 nucleotides (out of 3569) were positioned, while only a fragmented backbone was outlined for the rest of the genome, making a detailed model reconstruction necessary. For model reconstruction, in addition to the available atomistic structure information, we extensively used the predicted secondary structure of the genome (base pairing). The technique was based on semi-automatic building of relatively large strands of RNA with subsequent manual positioning over the traced backbone. The entire virus structure (capsid + genome) was validated by a molecular dynamics run in physiological solution with ions at standard conditions confirming the stability of the model.