College of Animal Science, Yangtze University, Jingzhou 430023, China.
State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China.
Viruses. 2022 May 12;14(5):1032. doi: 10.3390/v14051032.
Aquareovirus, which is a member of the Reoviridae family, was isolated from aquatic animals. A close molecular evolutionary relationship between aquareoviruses and mammalian orthoreoviruses was revealed. However, the functions of the aquareovirus genome-encoded proteins are poorly understood. We investigated the molecular characteristics of the outer capsid proteins, namely, VP5 and VP7, of grass carp reovirus (GCRV). The peptides VP5 and VP7 were determined using in-gel tryptic digestion and mass spectrometry. Recovered peptides represented 76% and 66% of the full-length VP5 and VP7 sequences, respectively. Significantly, two-lysine acetylation, as well as two-serine and two-threonine phosphorylation modifications, were first revealed in VP5. We found that the initial amino acid in VP5 was Pro43, suggesting that a lower amount of VP5 remained uncleaved in virions at the autocleavage site (Asn42-Pro43). Further biochemical evidence showed that the cleaved VP5N/VP5C conformation was the major constituent of the particles. Moreover, early cleavage fragments of VP7 and enhanced infectivity were detected after limited tryptic digestion of GCRV, indicating that stepwise VP7 cleavage is essential for VP5 conformational rearrangement. Our results provide insights into the roles of posttranslational modifications in VP5 and its association with VP7 in the viral life cycle.
水生呼肠孤病毒，属于呼肠孤病毒科的一员，从水生动物中分离出来。水生呼肠孤病毒与哺乳动物正呼肠孤病毒之间存在密切的分子进化关系。然而，水生呼肠孤病毒基因组编码蛋白的功能还知之甚少。我们研究了草鱼呼肠孤病毒（GCRV）的外壳蛋白，即 VP5 和 VP7 的分子特征。使用胶内酶切和质谱法确定了 VP5 和 VP7 的肽段。回收的肽段分别代表全长 VP5 和 VP7 序列的 76%和 66%。值得注意的是，首次在 VP5 中发现了两个赖氨酸乙酰化以及两个丝氨酸和两个苏氨酸磷酸化修饰。我们发现 VP5 中的起始氨基酸是 Pro43，这表明在自切割位点（Asn42-Pro43）处，病毒粒子中 VP5 的未切割量较少。进一步的生化证据表明，切割的 VP5N/VP5C 构象是颗粒的主要成分。此外，在对 GCRV 进行有限胰蛋白酶消化后，检测到 VP7 的早期切割片段和增强的感染性，这表明 VP7 的逐步切割对于 VP5 构象重排是必不可少的。我们的研究结果为 VP5 中的翻译后修饰及其在病毒生命周期中与 VP7 的关联提供了新的认识。
