Ebola virus VP35 has novel NTPase and helicase-like activities

Shu, Ting; Gan, Tianyu; Bai, Peng; Wang, Xiaotong; Qian, Qi; Zhou, Hui; Cheng, Qi; Qiu, Yang; Yin, Lei; Zhong, Jin; Zhou, Xi

Because a number of viral RNA remodelling proteins also have NTPase activity (22, 34, 47) , we next examined if EBOV VP35 has NTPase activity that can hydrolyze NTPs (ATP, GTP, CTP and UTP) by using a sensitive colorimetric assay that measures the total amount of free orthophosphate released after ATP hydrolysis. Our data showed that MBP-VP35 hydrolyzed all four types of NTPs, with a preference on ATP, GTP and UTP over CTP ( Figure 1A) . Moreover, when increasing concentrations of MBP-VP35 were incubated with ATP, MBP-VP35 can efficiently hydrolyze ATP in a dose-dependent manner ( Figure 1B ; Supplementary Figure S3 ). Furthermore, because VP35 has NTPase activity, it is intriguing to examine if this protein has NTP-binding activity. Our data show that 5 -ATP agarose can readily pull-down MBP-VP35 but not MBP alone ( Figure 1C) , showing that VP35 does bind to ATP. Of note, MBP protein was expressed and purified exactly same as MBP-VP35, and showed negligible NTP hydrolysis and binding.