Wu Chengcheng, Tao Lihan, Zhou Quanyong, Zhang Fanfan, Zeng Yanbing
Institute of Animal Husbandry and Veterinary Medicine, Jiangxi Academy of Agricultural Sciences, Nanchang 330200, China.
College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330045, China.
Animals (Basel). 2025 Mar 18;15(6):865. doi: 10.3390/ani15060865.
The nonstructural protein 13 (NSP13) of PDCoV is a highly conservative helicase and plays key roles in viral replication. NSP13 contains a zinc-binding domain (ZBD), a helical Stalk domain, a beta-barrel 1B domain, and a core helicase domain. However, the specific functions of these domains of PDCoV NSP13 remain largely unknown. Here, we expressed and purified the wild-type NSP13 and various mutants with domain deletions, and the activities of these proteins were analyzed using multiple methods. We found that NSP13 possessed the abilities to hydrolyze ATP and unwind double-stranded nucleic acids, but the unwinding efficiency was lower than that of NSP13. In contrast, NSP13, NSP13, and NSP13 all lost their unwinding activity, but not their ATPase activity. These results revealed that the deletion of the ZBD impaired the unwinding activity of PDCoV helicase NSP13, and the Stalk and 1B domains were critical for NSP13 to separate the duplexes. The identification of the roles of each domain in this study was helpful to gain an in-depth understanding of the overall functions of helicase NSP13, providing a theoretical basis for the development of antiviral drugs targeting helicase.
猪德尔塔冠状病毒（PDCoV）的非结构蛋白13（NSP13）是一种高度保守的解旋酶，在病毒复制中起关键作用。NSP13包含一个锌结合结构域（ZBD）、一个螺旋茎结构域、一个β桶1B结构域和一个核心解旋酶结构域。然而，PDCoV NSP13这些结构域的具体功能仍 largely未知。在此，我们表达并纯化了野生型NSP13和各种结构域缺失的突变体，并使用多种方法分析了这些蛋白的活性。我们发现NSP13具有水解ATP和解开双链核酸的能力，但解旋效率低于NSP13。相比之下，NSP13、NSP13和NSP13均失去了解旋活性，但ATP酶活性未丧失。这些结果表明，ZBD的缺失损害了PDCoV解旋酶NSP13的解旋活性，茎结构域和1B结构域对NSP13解开双链至关重要。本研究中各结构域作用的鉴定有助于深入了解解旋酶NSP13的整体功能，为开发靶向解旋酶的抗病毒药物提供理论依据。
