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A programmable pAgo nuclease with RNA target preference from the psychrotolerant bacteria Mucilaginibacter paludis

View ORCID ProfileWenqiang Li, Yang Liu, Fei Wang, View ORCID ProfileLixin Ma
doi: https://doi.org/10.1101/2021.06.08.447469
Wenqiang Li
1State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan, Hubei 430062, China
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  • ORCID record for Wenqiang Li
Yang Liu
1State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan, Hubei 430062, China
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Fei Wang
1State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan, Hubei 430062, China
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  • For correspondence: malixing@hubu.edu.cn wangfei@hubu.edu.cn
Lixin Ma
1State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan, Hubei 430062, China
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  • For correspondence: malixing@hubu.edu.cn wangfei@hubu.edu.cn
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ABSTRACT

Argonaute (Ago) proteins are programmable nuclease found in both eukaryotes and prokaryotes. Prokaryotic Argonaute proteins (pAgos) share a high degree of structural homology with eukaryotic Argonaute proteins (eAgos) and eAgos are considered to evolve from pAgos. However, the majority of studied pAgos prefer to cleave DNA targets, and eAgos exclusively cleave RNA targets. Here, we characterize a novel pAgo, MbpAgo, from psychrotolerant bacteria Mucilaginibacter paludis that can be programmed with DNA guides and prefers to cleave RNA targets rather than DNA targets. MbpAgo can be active at a wide range of temperatures (4-65°C). In comparison with previously studied pAgos, MbpAgo is able to utilize 16-nt long 5’phosphorylated and 5’hydroxylated DNA guides for efficient and precise cleavage and displays no obvious preference for the 5’end nucleotide of a guide. Furthermore, the cleavage efficiency can be regulated by mismatches in the central and 3’supplementary regions of the guide. MbpAgo can efficiently cleave highly-structured RNA targets using both 5’phosphorylated and 5’hydroxylated DNA guides in the presence of Mg2+ or Mn2+. In conclusion, we have demonstrated that MbpAgo is a unique programmable nuclease that has a strong preference for RNA targets, with great potential applications in the field of nucleic acid biotechnology.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

  • ↵† The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted June 08, 2021.
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A programmable pAgo nuclease with RNA target preference from the psychrotolerant bacteria Mucilaginibacter paludis
Wenqiang Li, Yang Liu, Fei Wang, Lixin Ma
bioRxiv 2021.06.08.447469; doi: https://doi.org/10.1101/2021.06.08.447469
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A programmable pAgo nuclease with RNA target preference from the psychrotolerant bacteria Mucilaginibacter paludis
Wenqiang Li, Yang Liu, Fei Wang, Lixin Ma
bioRxiv 2021.06.08.447469; doi: https://doi.org/10.1101/2021.06.08.447469

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