The lysine at position 151 of the duck hepatitis A virus 1 2C protein is critical for its NTPase activities.

Veterinary microbiology 2022 Jan

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The duck hepatitis A virus 1 (DHAV-1) 2C protein was predicted to be a superfamily III helicase member and includes nucleotide binding (NTB) and putative RNA helicase activity motifs. To study whether DHAV-1 2C protein has NTB activity, we expressed DHAV-1 2C protein with maltose binding protein (MBP) to solve its poor solubility in a prokaryotic expression system. We showed that the DHAV-1 2C protein has nucleoside triphosphatase (NTPase) activity by measuring the released phosphate. The NTPase of the DHAV-1 2C protein is Mg2+ indispensable and affected by other biochemical characteristics such as Mn2+, Ca2+, Zn2+, Na+ and pH. Guanidine hydrochloride (GdnHCl), a potent inhibitor of viral RNA replication, inhibited ATPase activity of the DHAV-1 2C protein in a dose-dependent manner. Finally, we constructed three mutants to identify the key site for the ATPase activity of the DHAV-1 2C protein. These results indicate that lysine at position 151 of the DHAV-1 2C protein is very important for NTPase activity. Here, we demonstrated and partially characterized that the DHAV-1 2C protein has NTPase activity and showed that mutation of the lysine in the conserved Walker A impairs that activity. The results serve to confirm what is readily predicted from previous work on picornavirus 2C proteins. It also provides a basis for further study of the 2C protein and the function of NTPase activity on the viral life cycle. Copyright © 2021 Elsevier B.V. All rights reserved.

Citation

Xinhong Li, Xiaosi Tang, Mingshu Wang, Anchun Cheng, Xumin Ou, Sai Mao, Di Sun, Qiao Yang, Ying Wu, Shaqiu Zhang, Dekang Zhu, Renyong Jia, Shun Chen, Mafeng Liu, Xinxin Zhao, Juan Huang, Qun Gao, Bin Tian, Yunya Liu, Yanling Yu, Ling Zhang, Leichang Pan. The lysine at position 151 of the duck hepatitis A virus 1 2C protein is critical for its NTPase activities. Veterinary microbiology. 2022 Jan;264:109300