Centro de Biotecnología y Genómica de Plantas, Universidad Politécnica de Madrid-Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (CBGP, UPM-INIA), Campus Montegancedo, Autopista M-40, km 38, Pozuelo de Alarcón, 28223 Madrid, Spain.
Viruses. 2020 Jun 19;12(6):661. doi: 10.3390/v12060661.
The involvement of different structural domains of the coat protein (CP) of turnip mosaic virus, a potyvirus, in establishing and/or maintaining particle assembly was analyzed through deletion mutants of the protein. In order to identify exclusively those domains involved in protein-protein interactions within the particle, the analysis was performed by agroinfiltration "in planta", followed by the assessment of CP accumulation in leaves and the assembly of virus-like particles lacking nucleic acids, also known as empty virus-like particles (eVLP). Thus, the interactions involving viral RNA could be excluded. It was found that deletions precluding eVLP assembly did not allow for protein accumulation either, probably indicating that non-assembled CP protein was degraded in the plant leaves. Deletions involving the CP structural core were incompatible with particle assembly. On the N-terminal domain, only the deletion avoiding the subdomain involved in interactions with other CP subunits was incorporated into eVLPs. The C-terminal domain was shown to be more permissive to deletions. Assembled eVLPs were found for mutants, eliminating the whole domain. The C-terminal domain mutants were unusually long, suggesting some role of the domain in the regulation of particle length. The identification of the CP domains responsible for eVLP formation will allow for new approaches to protein stretch replacement with peptides or proteins of nanobiotechnological interest. Finally, specific cases of application are considered.
通过对芜菁花叶病毒（一种马铃薯 Y 病毒属病毒）外壳蛋白（CP）的不同结构域进行缺失突变，分析了 CP 参与建立和/或维持粒子组装的作用。为了专门鉴定那些参与粒子内蛋白-蛋白相互作用的结构域，通过农杆菌浸润“体内”进行了分析，随后评估了叶片中 CP 的积累以及缺乏核酸的病毒样颗粒（也称为空病毒样颗粒，eVLP）的组装。这样就可以排除涉及病毒 RNA 的相互作用。结果发现，阻止 eVLP 组装的缺失也不允许 CP 积累，这可能表明未组装的 CP 蛋白在植物叶片中被降解。涉及 CP 结构核心的缺失与粒子组装不兼容。在 N 端结构域中，只有避免与其他 CP 亚基相互作用的亚结构域缺失的突变才能被整合到 eVLP 中。C 端结构域对缺失更为宽容。对于消除整个结构域的突变体，组装的 eVLP 被发现。C 端结构域突变体异常长，这表明该结构域在调节粒子长度方面可能具有一定作用。鉴定负责 eVLP 形成的 CP 结构域将允许采用新方法用肽或具有纳米生物技术应用前景的蛋白替代蛋白质的伸展部分。最后，考虑了具体的应用案例。
