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A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase

During RNA elongation, the influenza A viral (IAV) RNA-dependent RNA polymerase (RdRp) residues in the active site interact with the triphosphate moiety of nucleoside triphosphate (NTP) for catalysis. The molecular mechanisms by which they control the rate and fidelity of NTP incorporation remain el...

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Autores principales: Xu, Xinzhou, Zhang, Lu, Chu, Julie Tung Sem, Wang, Yuqing, Chin, Alex Wing Hong, Chong, Tin Hang, Dai, Zixi, Poon, Leo Lit Man, Cheung, Peter Pak-Hang, Huang, Xuhui
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8421151/
https://www.ncbi.nlm.nih.gov/pubmed/34379778
http://dx.doi.org/10.1093/nar/gkab660
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author Xu, Xinzhou
Zhang, Lu
Chu, Julie Tung Sem
Wang, Yuqing
Chin, Alex Wing Hong
Chong, Tin Hang
Dai, Zixi
Poon, Leo Lit Man
Cheung, Peter Pak-Hang
Huang, Xuhui
author_facet Xu, Xinzhou
Zhang, Lu
Chu, Julie Tung Sem
Wang, Yuqing
Chin, Alex Wing Hong
Chong, Tin Hang
Dai, Zixi
Poon, Leo Lit Man
Cheung, Peter Pak-Hang
Huang, Xuhui
author_sort Xu, Xinzhou
collection PubMed
description During RNA elongation, the influenza A viral (IAV) RNA-dependent RNA polymerase (RdRp) residues in the active site interact with the triphosphate moiety of nucleoside triphosphate (NTP) for catalysis. The molecular mechanisms by which they control the rate and fidelity of NTP incorporation remain elusive. Here, we demonstrated through enzymology, virology and computational approaches that the R239 and K235 in the PB1 subunit of RdRp are critical to controlling the activity and fidelity of transcription. Contrary to common beliefs that high-fidelity RdRp variants exert a slower incorporation rate, we discovered a first-of-its-kind, single lysine-to-arginine mutation on K235 exhibited enhanced fidelity and activity compared with wild-type. In particular, we employed a single-turnover NTP incorporation assay for the first time on IAV RdRp to show that K235R mutant RdRp possessed a 1.9-fold increase in the transcription activity of the cognate NTP and a 4.6-fold increase in fidelity compared to wild-type. Our all-atom molecular dynamics simulations further elucidated that the higher activity is attributed to the shorter distance between K235R and the triphosphate moiety of NTP compared with wild-type. These results provide novel insights into NTP incorporation and fidelity control mechanisms, which lay the foundation for the rational design of IAV vaccine and antiviral targets.
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spelling pubmed-84211512021-09-09 A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase Xu, Xinzhou Zhang, Lu Chu, Julie Tung Sem Wang, Yuqing Chin, Alex Wing Hong Chong, Tin Hang Dai, Zixi Poon, Leo Lit Man Cheung, Peter Pak-Hang Huang, Xuhui Nucleic Acids Res Nucleic Acid Enzymes During RNA elongation, the influenza A viral (IAV) RNA-dependent RNA polymerase (RdRp) residues in the active site interact with the triphosphate moiety of nucleoside triphosphate (NTP) for catalysis. The molecular mechanisms by which they control the rate and fidelity of NTP incorporation remain elusive. Here, we demonstrated through enzymology, virology and computational approaches that the R239 and K235 in the PB1 subunit of RdRp are critical to controlling the activity and fidelity of transcription. Contrary to common beliefs that high-fidelity RdRp variants exert a slower incorporation rate, we discovered a first-of-its-kind, single lysine-to-arginine mutation on K235 exhibited enhanced fidelity and activity compared with wild-type. In particular, we employed a single-turnover NTP incorporation assay for the first time on IAV RdRp to show that K235R mutant RdRp possessed a 1.9-fold increase in the transcription activity of the cognate NTP and a 4.6-fold increase in fidelity compared to wild-type. Our all-atom molecular dynamics simulations further elucidated that the higher activity is attributed to the shorter distance between K235R and the triphosphate moiety of NTP compared with wild-type. These results provide novel insights into NTP incorporation and fidelity control mechanisms, which lay the foundation for the rational design of IAV vaccine and antiviral targets. Oxford University Press 2021-08-11 /pmc/articles/PMC8421151/ /pubmed/34379778 http://dx.doi.org/10.1093/nar/gkab660 Text en © The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research. https://creativecommons.org/licenses/by-nc/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (http://creativecommons.org/licenses/by-nc/4.0/ (https://creativecommons.org/licenses/by-nc/4.0/) ), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Nucleic Acid Enzymes
Xu, Xinzhou
Zhang, Lu
Chu, Julie Tung Sem
Wang, Yuqing
Chin, Alex Wing Hong
Chong, Tin Hang
Dai, Zixi
Poon, Leo Lit Man
Cheung, Peter Pak-Hang
Huang, Xuhui
A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase
title A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase
title_full A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase
title_fullStr A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase
title_full_unstemmed A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase
title_short A novel mechanism of enhanced transcription activity and fidelity for influenza A viral RNA-dependent RNA polymerase
title_sort novel mechanism of enhanced transcription activity and fidelity for influenza a viral rna-dependent rna polymerase
topic Nucleic Acid Enzymes
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8421151/
https://www.ncbi.nlm.nih.gov/pubmed/34379778
http://dx.doi.org/10.1093/nar/gkab660
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