Mutations at KFRDI and VGK domains of enterovirus 71 3C protease affect its RNA binding and proteolytic activities |
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Authors: | Shin-Ru Shih Chiayn Chiang Tzu-Chun Chen Cheng-Nan Wu John Tsu-An Hsu Jin-Ching Lee Ming-Jing Hwang Mei-Ling Li Guang-Wu Chen Mei-Shan Ho |
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Affiliation: | (1) Clinical Virology Laboratory, Department of Clinical Pathology, Chang Gung Memorial Hospital, Tao-Yuan;(2) Division of Biomedical Sciences, Academia Sinica, Taiwan;(3) Division of Biotechnology and Pharmaceutical Research, National Health Research Institutes, Taipei;(4) Department of Chemical Engineering, National Tsing Hua University, Taiwan;(5) Division of Biostatistics and Bioinformatics, National Health Research Institutes, Taipei, Taiwan;(6) School of Medical Technology, Chang Gung University, 259 Wen-Hua 1st Rd., Kwei-Shan, Tao-Yuan, Taiwan |
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Abstract: | The 3C proteases (3Cpro) of enterovirus 71 (EV71) is a good molecular target for drug discovery. Notably, this protease was found to possess RNA-binding activity. The regions responsible for RNA binding were classified as KFRDI (positions 82–86) and VGK (positions 154–156) in 3Cpro by mutagenesis study. Although the RNA-binding regions are structurally distinct from the catalytic site of EV71 3Cpro, mutations in the RNA-binding regions influenced 3Cpro proteolytic activity. In contrast, mutations at the catalytic site had almost no influence on RNA binding ability. We identified certain mutations within 3Cpro which abrogated both the RNA-binding activity of the expressed, recombinant, protease and the ability to rescue virus from an infectious full-length clone of EV71 (pEV71). Interestingly, mutation at position 84 from Arg(R) to Lys(K) was found to retain good RNA binding and proteolytic activity for the recombinant 3Cpro; however, no virus could be rescued when pEV71 with the R84K mutation was introduced into the infectious copy. Together, these results may provide useful information for using 3Cpro as the molecular target to develop anti-EV71 agents.The second and the third authors contributed equally to this work. |
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Keywords: | Enterovirus 71 3C protease RNA binding |
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