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A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacterium Kurthia massiliensis

Kropocheva, Ekaterina and Kuzmenko, Anton and Aravin, Alexei A. and Esyunina, Daria and Kulbachiskiy, Andrey (2021) A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacterium Kurthia massiliensis. Nucleic Acids Research, 49 (7). pp. 4054-4065. ISSN 0305-1048. PMCID PMC8053121. doi:10.1093/nar/gkab182.

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Argonaute proteins are programmable nucleases that are found in both eukaryotes and prokaryotes and provide defense against invading genetic elements. Although some prokaryotic argonautes (pAgos) were shown to recognize RNA targets in vitro, the majority of studied pAgos have strict specificity toward DNA, which limits their practical use in RNA-centric applications. Here, we describe a unique pAgo nuclease, KmAgo, from the mesophilic bacterium Kurthia massiliensis that can be programmed with either DNA or RNA guides and can precisely cleave both DNA and RNA targets. KmAgo binds 16–20 nt long 5′-phosphorylated guide molecules with no strict specificity for their sequence and is active in a wide range of temperatures. In bacterial cells, KmAgo is loaded with small DNAs with no obvious sequence preferences suggesting that it can uniformly target genomic sequences. Mismatches between the guide and target sequences greatly affect the efficiency and precision of target cleavage, depending on the mismatch position and the nature of the reacting nucleic acids. Target RNA cleavage by KmAgo depends on the formation of secondary structure indicating that KmAgo can be used for structural probing of RNA. These properties of KmAgo open the way for its use for highly specific nucleic acid detection and cleavage.

Item Type:Article
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URLURL TypeDescription Paper ItemData CentralArticle
Kuzmenko, Anton0000-0001-7169-0561
Aravin, Alexei A.0000-0002-6956-8257
Esyunina, Daria0000-0002-3706-4425
Kulbachiskiy, Andrey0000-0002-2292-6424
Additional Information:© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (, which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. Received February 04, 2021; Revised March 02, 2021; Editorial Decision March 03, 2021; Accepted March 06, 2021; Published: 21 March 2021. We thank Sergei Ryazansky for help with bioinformatic analysis of pAgo proteins and Denis Yudin for initial analysis of smDNA libraries. Funding: Russian Foundation for Basic Research [18-29-07086 to A.K.; biochemical analysis of KmAgo]; Russian Science Foundation [19-14-00359 to D.E.; analysis of in vivo activities of KmAgo]. Funding for open access charge: Russian Foundation for Basic Research. Data Availability: The smDNA sequencing datasets generated in this study are available from the Gene Expression Omnibus (GEO) database under the accession number GSE168010. Conflict of interest statement. None declared.
Funding AgencyGrant Number
Russian Foundation for Basic Research18-29-07086
Russian Science Foundation19-14-00359
Issue or Number:7
PubMed Central ID:PMC8053121
Record Number:CaltechAUTHORS:20210203-143505325
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Official Citation:Ekaterina Kropocheva, Anton Kuzmenko, Alexei A Aravin, Daria Esyunina, Andrey Kulbachinskiy, A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacterium Kurthia massiliensis, Nucleic Acids Research, Volume 49, Issue 7, 19 April 2021, Pages 4054–4065,
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:107892
Deposited By: Tony Diaz
Deposited On:03 Feb 2021 23:39
Last Modified:05 Aug 2021 16:46

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