Sugiura Yoshiro, Shimizu Kenta, Takahashi Tatsuki, Ueno Shiori, Tanigou Haruka, Amarbayasgalan Sodbayasgalan, Kamitani Wataru
Department of Infectious Disease and Host Defense, Graduate School of Medicine, Gunma University, Gunma, Japan.
PLoS One. 2024 Dec 6;19(12):e0312800. doi: 10.1371/journal.pone.0312800. eCollection 2024.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) non-structural protein 5 (nsp5) is a cysteine protease involved in viral replication and suppression of the host immune system. The substrate-binding domain of nsp5 is important for its protease activity. However, the relationship between nsp5 protease activity and viral replication remains unclear. We confirmed the importance of amino acid T25 in the nsp5 substrate-binding domain for viral replication using a split luciferase assay. By generating recombinant viruses using bacterial artificial chromosomes, we found that the proliferation of viruses with the T25I mutation in nsp5 was cell-dependent in culture. Furthermore, mice infected with the T25I mutant recombinant virus with a mouse acclimation backbone showed weight loss and increased lung viral load, similar to the wild-type (WT) infected group, up to 3 days after infection. However, after day 4, the lung viral load was significantly reduced in the T25I-infected group compared to that in the WT-infected group. This suggests that nsp5 T25 is involved in the pathogenesis of SARS-CoV-2.
严重急性呼吸综合征冠状病毒2（SARS-CoV-2）非结构蛋白5（nsp5）是一种半胱氨酸蛋白酶，参与病毒复制和宿主免疫系统的抑制。nsp5的底物结合结构域对其蛋白酶活性很重要。然而，nsp5蛋白酶活性与病毒复制之间的关系仍不清楚。我们使用分裂荧光素酶测定法证实了nsp5底物结合结构域中氨基酸T25对病毒复制的重要性。通过使用细菌人工染色体产生重组病毒，我们发现nsp5中具有T25I突变的病毒在培养中的增殖是细胞依赖性的。此外，感染了带有小鼠适应骨架的T25I突变重组病毒的小鼠在感染后3天内出现体重减轻和肺部病毒载量增加，与野生型（WT）感染组相似。然而，在第4天后，与WT感染组相比，T25I感染组的肺部病毒载量显著降低。这表明nsp5 T25参与了SARS-CoV-2的发病机制。
