An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides
Prokaryotic Argonaute (pAgo) nucleases with precise DNA cleavage activity show great potential for gene <sc>manipulation</sc>. Extensive biochemical studies have revealed that recognition of guides with different 5′ groups by Ago is important for biocatalysis. Here, we identified an Ago...
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| Format: | Article |
| Language: | English |
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China Science Publishing & Media Ltd.
2022-05-01
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| Series: | Acta Biochimica et Biophysica Sinica |
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| Online Access: | https://www.sciengine.com/doi/10.3724/abbs.2022047 |
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| author | Sun Yingying Guo Xiang Lu Hui Chen Liuqing Huang Fei Liu Qian Feng Yan |
| author_facet | Sun Yingying Guo Xiang Lu Hui Chen Liuqing Huang Fei Liu Qian Feng Yan |
| author_sort | Sun Yingying |
| collection | DOAJ |
| description | Prokaryotic Argonaute (pAgo) nucleases with precise DNA cleavage activity show great potential for gene <sc>manipulation</sc>. Extensive biochemical studies have revealed that recognition of guides with different 5′ groups by Ago is important for biocatalysis. Here, we identified an Ago from the thermophilic Thermus parvatiensis (TpsAgo) and analyzed the regulatory effect of 5′-modified guides on TpsAgo cleavage activity. Recombinant TpsAgo cleaves single-stranded DNA and RNA targets at 65–90°C, which is mediated by a 5′ hydroxyl or phosphate DNA guide. Notably, TpsAgo can utilize various 5′-modified DNA guides for catalysis, including 5′-NH<sub>2</sub>C<sub>6</sub>, 5′-Biotin, 5′-FAM and 5′-SHC<sub>6</sub> guides. Moreover, TpsAgo performs programmable cleavage of double-stranded DNA at temperatures over 80°C and strongly tolerates NaCl concentrations up to 3.2 M. These results provide insight into the catalytic performance of Agos by guide regulation, which may facilitate their biotechnological applications. |
| format | Article |
| id | doaj-art-6897a281a5594f2a88623bdaff01c4b3 |
| institution | Kabale University |
| issn | 1672-9145 |
| language | English |
| publishDate | 2022-05-01 |
| publisher | China Science Publishing & Media Ltd. |
| record_format | Article |
| series | Acta Biochimica et Biophysica Sinica |
| spelling | doaj-art-6897a281a5594f2a88623bdaff01c4b32025-08-20T03:54:01ZengChina Science Publishing & Media Ltd.Acta Biochimica et Biophysica Sinica1672-91452022-05-015468669510.3724/abbs.202204720d259ccAn Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guidesSun Yingying0Guo Xiang1Lu Hui2Chen Liuqing3Huang Fei4Liu Qian5Feng Yan6["State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China"]["State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China"]["State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China"]["Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen 518055, China"]["State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China"]["State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China"]["State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China"]Prokaryotic Argonaute (pAgo) nucleases with precise DNA cleavage activity show great potential for gene <sc>manipulation</sc>. Extensive biochemical studies have revealed that recognition of guides with different 5′ groups by Ago is important for biocatalysis. Here, we identified an Ago from the thermophilic Thermus parvatiensis (TpsAgo) and analyzed the regulatory effect of 5′-modified guides on TpsAgo cleavage activity. Recombinant TpsAgo cleaves single-stranded DNA and RNA targets at 65–90°C, which is mediated by a 5′ hydroxyl or phosphate DNA guide. Notably, TpsAgo can utilize various 5′-modified DNA guides for catalysis, including 5′-NH<sub>2</sub>C<sub>6</sub>, 5′-Biotin, 5′-FAM and 5′-SHC<sub>6</sub> guides. Moreover, TpsAgo performs programmable cleavage of double-stranded DNA at temperatures over 80°C and strongly tolerates NaCl concentrations up to 3.2 M. These results provide insight into the catalytic performance of Agos by guide regulation, which may facilitate their biotechnological applications.https://www.sciengine.com/doi/10.3724/abbs.2022047Argonaute<italic>Thermus parvatiensis</italic>endonuclease5′-modified guide |
| spellingShingle | Sun Yingying Guo Xiang Lu Hui Chen Liuqing Huang Fei Liu Qian Feng Yan An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides Acta Biochimica et Biophysica Sinica Argonaute <italic>Thermus parvatiensis</italic> endonuclease 5′-modified guide |
| title | An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides |
| title_full | An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides |
| title_fullStr | An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides |
| title_full_unstemmed | An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides |
| title_short | An Argonaute from Thermus parvatiensis exhibits endonuclease activity mediated by 5′ chemically modified DNA guides |
| title_sort | argonaute from thermus parvatiensis exhibits endonuclease activity mediated by 5 chemically modified dna guides |
| topic | Argonaute <italic>Thermus parvatiensis</italic> endonuclease 5′-modified guide |
| url | https://www.sciengine.com/doi/10.3724/abbs.2022047 |
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