The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis
Abstract Argonaute proteins (Agos) play essential roles in nucleic acid targeting across life domains. While eukaryotic Agos (eAgos) utilize small-interfering RNAs (siRNAs) or microRNAs (miRNAs) for RNA interference, the mechanisms driving prokaryotic Agos (pAgos) in bacterial defense remain underex...
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| Format: | Article |
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Nature Portfolio
2025-07-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-025-61926-4 |
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| author | Yuchan Liu Jiasu Zhang Ji Liu Shengchun Zhang Linfeng An Wenbing Xie Kaiming Zhang Shanshan Li |
| author_facet | Yuchan Liu Jiasu Zhang Ji Liu Shengchun Zhang Linfeng An Wenbing Xie Kaiming Zhang Shanshan Li |
| author_sort | Yuchan Liu |
| collection | DOAJ |
| description | Abstract Argonaute proteins (Agos) play essential roles in nucleic acid targeting across life domains. While eukaryotic Agos (eAgos) utilize small-interfering RNAs (siRNAs) or microRNAs (miRNAs) for RNA interference, the mechanisms driving prokaryotic Agos (pAgos) in bacterial defense remain underexplored. Here, we characterize the mesophilic pAgo from Clostridium perfringens (CpAgo), which exhibits robust guide-independent and DNA-guided activity at 37 °C. CpAgo efficiently degrades plasmids and structured RNAs into small fragments, generating DNA fragments that serve as guides for subsequent cleavage. Cryo-electron microscopy reveals a positively-charged PAZ nucleotide-binding pocket, critical for both guide-dependent and guide-independent substrate recognition and cleavage. Structural analysis identifies CpAgo’s dimerization as a prerequisite for catalytic activity, supporting both nucleic acid degradation and targeted action. Functional assays in Escherichia coli demonstrate CpAgo’s role in bacterial defense by mediating plasmid degradation and DNA-guided cleavage. These findings position CpAgo as a critical component of prokaryotic immunity and a promising tool for biotechnology. |
| format | Article |
| id | doaj-art-7858d8a0605e4334bb01a6378e30c1f7 |
| institution | DOAJ |
| issn | 2041-1723 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj-art-7858d8a0605e4334bb01a6378e30c1f72025-08-20T03:05:05ZengNature PortfolioNature Communications2041-17232025-07-0116111310.1038/s41467-025-61926-4The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysisYuchan Liu0Jiasu Zhang1Ji Liu2Shengchun Zhang3Linfeng An4Wenbing Xie5Kaiming Zhang6Shanshan Li7Department of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaDepartment of Urology, The First Affiliated Hospital of USTC, MOE Key Laboratory for Cellular Dynamics, Hefei National Research Center for Physical Sciences at the Microscale, Center for Advanced Interdisciplinary Science and Biomedicine of IHM, The RNA Institute, Division of Life Sciences and Medicine, University of Science and Technology of ChinaAbstract Argonaute proteins (Agos) play essential roles in nucleic acid targeting across life domains. While eukaryotic Agos (eAgos) utilize small-interfering RNAs (siRNAs) or microRNAs (miRNAs) for RNA interference, the mechanisms driving prokaryotic Agos (pAgos) in bacterial defense remain underexplored. Here, we characterize the mesophilic pAgo from Clostridium perfringens (CpAgo), which exhibits robust guide-independent and DNA-guided activity at 37 °C. CpAgo efficiently degrades plasmids and structured RNAs into small fragments, generating DNA fragments that serve as guides for subsequent cleavage. Cryo-electron microscopy reveals a positively-charged PAZ nucleotide-binding pocket, critical for both guide-dependent and guide-independent substrate recognition and cleavage. Structural analysis identifies CpAgo’s dimerization as a prerequisite for catalytic activity, supporting both nucleic acid degradation and targeted action. Functional assays in Escherichia coli demonstrate CpAgo’s role in bacterial defense by mediating plasmid degradation and DNA-guided cleavage. These findings position CpAgo as a critical component of prokaryotic immunity and a promising tool for biotechnology.https://doi.org/10.1038/s41467-025-61926-4 |
| spellingShingle | Yuchan Liu Jiasu Zhang Ji Liu Shengchun Zhang Linfeng An Wenbing Xie Kaiming Zhang Shanshan Li The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis Nature Communications |
| title | The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis |
| title_full | The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis |
| title_fullStr | The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis |
| title_full_unstemmed | The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis |
| title_short | The PAZ pocket and dimerization drive CpAgo’s guide-independent and DNA-guided dual catalysis |
| title_sort | paz pocket and dimerization drive cpago s guide independent and dna guided dual catalysis |
| url | https://doi.org/10.1038/s41467-025-61926-4 |
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