3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM
The Volta phase plate is a recently developed electron cryo-microscopy (cryo-EM) device that enables contrast enhancement of biological samples. Here we have evaluated the potential of combining phase-plate imaging and single particle analysis to determine the structure of a small protein–DNA comple...
| Main Authors: | , , , , , , , |
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| Format: | Journal Article |
| Language: | English |
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2016
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| Online Access: | https://hdl.handle.net/10356/83403 http://hdl.handle.net/10220/41490 |
| _version_ | 1811694242715664384 |
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| author | Chua, Eugene Yue Dao Vogirala, Vinod Kumar Inian, Oviya Wong, Andrew See Weng Nordenskiold, Lars Plitzko, Juergen M. Danev, Radostin Sandin, Sara |
| author2 | School of Biological Sciences |
| author_facet | School of Biological Sciences Chua, Eugene Yue Dao Vogirala, Vinod Kumar Inian, Oviya Wong, Andrew See Weng Nordenskiold, Lars Plitzko, Juergen M. Danev, Radostin Sandin, Sara |
| author_sort | Chua, Eugene Yue Dao |
| collection | NTU |
| description | The Volta phase plate is a recently developed electron cryo-microscopy (cryo-EM) device that enables contrast enhancement of biological samples. Here we have evaluated the potential of combining phase-plate imaging and single particle analysis to determine the structure of a small protein–DNA complex. To test the method, we made use of a 200 kDa Nucleosome Core Particle (NCP) reconstituted with 601 DNA for which a high-resolution X-ray crystal structure is known. We find that the phase plate provides a significant contrast enhancement that permits individual NCPs and DNA to be clearly identified in amorphous ice. The refined structure from 26,060 particles has an overall resolution of 3.9 Å and the density map exhibits structural features consistent with the estimated resolution, including clear density for amino acid side chains and DNA features such as the phosphate backbone. Our results demonstrate that phase-plate cryo-EM promises to become an important method to determine novel near-atomic resolution structures of small and challenging samples, such as nucleosomes in complex with nucleosome-binding factors. |
| first_indexed | 2024-10-01T07:04:28Z |
| format | Journal Article |
| id | ntu-10356/83403 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T07:04:28Z |
| publishDate | 2016 |
| record_format | dspace |
| spelling | ntu-10356/834032023-02-28T17:00:47Z 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM Chua, Eugene Yue Dao Vogirala, Vinod Kumar Inian, Oviya Wong, Andrew See Weng Nordenskiold, Lars Plitzko, Juergen M. Danev, Radostin Sandin, Sara School of Biological Sciences NTU Institute of Structural Biology Nucleosome Core Particle cryo-EM The Volta phase plate is a recently developed electron cryo-microscopy (cryo-EM) device that enables contrast enhancement of biological samples. Here we have evaluated the potential of combining phase-plate imaging and single particle analysis to determine the structure of a small protein–DNA complex. To test the method, we made use of a 200 kDa Nucleosome Core Particle (NCP) reconstituted with 601 DNA for which a high-resolution X-ray crystal structure is known. We find that the phase plate provides a significant contrast enhancement that permits individual NCPs and DNA to be clearly identified in amorphous ice. The refined structure from 26,060 particles has an overall resolution of 3.9 Å and the density map exhibits structural features consistent with the estimated resolution, including clear density for amino acid side chains and DNA features such as the phosphate backbone. Our results demonstrate that phase-plate cryo-EM promises to become an important method to determine novel near-atomic resolution structures of small and challenging samples, such as nucleosomes in complex with nucleosome-binding factors. MOE (Min. of Education, S’pore) Published version 2016-09-26T02:56:11Z 2019-12-06T15:21:44Z 2016-09-26T02:56:11Z 2019-12-06T15:21:44Z 2016 2016 Journal Article Chua, E. Y. D., Vogirala, V. K., Inian, O., Wong, A. S. W., Nordenskiold, L., Plitzko, J. M., et al. (2016). 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM. Nucleic Acids Research, 44(17), 8013-8019. 0305-1048 https://hdl.handle.net/10356/83403 http://hdl.handle.net/10220/41490 10.1093/nar/gkw708 27563056 195117 en Nucleic Acids Research © The Author(s) 2016. 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 License (http://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. 7 p. application/pdf |
| spellingShingle | Nucleosome Core Particle cryo-EM Chua, Eugene Yue Dao Vogirala, Vinod Kumar Inian, Oviya Wong, Andrew See Weng Nordenskiold, Lars Plitzko, Juergen M. Danev, Radostin Sandin, Sara 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM |
| title | 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM |
| title_full | 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM |
| title_fullStr | 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM |
| title_full_unstemmed | 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM |
| title_short | 3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM |
| title_sort | 3 9 a structure of the nucleosome core particle determined by phase plate cryo em |
| topic | Nucleosome Core Particle cryo-EM |
| url | https://hdl.handle.net/10356/83403 http://hdl.handle.net/10220/41490 |
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