Accelerated cryo-EM-guided determination of three-dimensional RNA-only structures
Abstract
The discovery and design of biologically important RNA molecules is outpacing three-dimensional structural characterization. In this work, we demonstrate that cryo-electron microscopy can routinely resolve maps of RNA-only systems and that these maps enable subnanometer-resolution coordinate estimation when complemented with multidimensional chemical mapping and Rosetta DRRAFTER computational modeling. Furthermore ,this hybrid ‘Ribosolve’ pipeline detects and falsifies homologies and conformational rearrangements in 11 previously unknown 119- to 338-nucleotide protein-free RNA structures: full-length Tetrahymena ribozyme, hc16 ligase with and without substrate, full-length Vibrio cholerae and Fusobacterium nucleatum glycine riboswitch aptamers with and without glycine, Mycobacterium SAM-IV riboswitch with and without S-adenosylmethionine, and the computer-designed ATP-TTR-3 aptamer with and without AMP. Simulation benchmarks, blind challenges, compensatory mutagenesis, cross-RNA homologies and internal controls demonstrate that Ribosolve can accurately resolve the global architectures of RNA molecules but does not resolve atomic details. These tests offer guidelines for making inferences in future RNA structural studies with similarly accelerated throughput.
- Authors:
-
[1];
[2];
[2];
[2];
[1];
[1];
[5];
[1]
- Stanford Univ., CA (United States)
- Stanford Univ., CA (United States). James H. Clark Center
- Stanford Univ., CA (United States). James H. Clark Center; Sichuan Univ. (China). West China Hospital, State Key Lab. of Biotherapy
- Stanford Univ., CA (United States); Univ. of Nebraska, Lincoln, NE (United States)
- Stanford Univ., CA (United States); Stanford Univ., CA (United States). James H. Clark Center; SLAC National Accelerator Lab., Menlo Park, CA (United States). Stanford Synchrotron Radiation Lightsource (SSRL)
- Publication Date:
- Research Org.:
- SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC); National Institutes of Health (NIH); National Science Foundation (NSF)
- OSTI Identifier:
- 1646882
- Grant/Contract Number:
- AC02-76SF00515; R01GM079429; R35 GM112579; P01AI120943; S10 OD021600; R21 AI145647; U54GM103297; P41GM103832; GRFP
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Nature Methods
- Additional Journal Information:
- Journal Volume: 17; Journal Issue: 7; Journal ID: ISSN 1548-7091
- Publisher:
- Nature Publishing Group
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; Computational biophysics; cryoelectron microscopy; RNA
Citation Formats
Kappel, Kalli, Zhang, Kaiming, Su, Zhaoming, Watkins, Andrew M., Kladwang, Wipapat, Li, Shanshan, Pintilie, Grigore, Topkar, Ved V., Rangan, Ramya, Zheludev, Ivan N., Yesselman, Joseph D., Chiu, Wah, and Das, Rhiju. Accelerated cryo-EM-guided determination of three-dimensional RNA-only structures. United States: N. p., 2020.
Web. doi:10.1038/s41592-020-0878-9.
Kappel, Kalli, Zhang, Kaiming, Su, Zhaoming, Watkins, Andrew M., Kladwang, Wipapat, Li, Shanshan, Pintilie, Grigore, Topkar, Ved V., Rangan, Ramya, Zheludev, Ivan N., Yesselman, Joseph D., Chiu, Wah, & Das, Rhiju. Accelerated cryo-EM-guided determination of three-dimensional RNA-only structures. United States. https://doi.org/10.1038/s41592-020-0878-9
Kappel, Kalli, Zhang, Kaiming, Su, Zhaoming, Watkins, Andrew M., Kladwang, Wipapat, Li, Shanshan, Pintilie, Grigore, Topkar, Ved V., Rangan, Ramya, Zheludev, Ivan N., Yesselman, Joseph D., Chiu, Wah, and Das, Rhiju. Thu .
"Accelerated cryo-EM-guided determination of three-dimensional RNA-only structures". United States. https://doi.org/10.1038/s41592-020-0878-9. https://www.osti.gov/servlets/purl/1646882.
@article{osti_1646882,
title = {Accelerated cryo-EM-guided determination of three-dimensional RNA-only structures},
author = {Kappel, Kalli and Zhang, Kaiming and Su, Zhaoming and Watkins, Andrew M. and Kladwang, Wipapat and Li, Shanshan and Pintilie, Grigore and Topkar, Ved V. and Rangan, Ramya and Zheludev, Ivan N. and Yesselman, Joseph D. and Chiu, Wah and Das, Rhiju},
abstractNote = {The discovery and design of biologically important RNA molecules is outpacing three-dimensional structural characterization. In this work, we demonstrate that cryo-electron microscopy can routinely resolve maps of RNA-only systems and that these maps enable subnanometer-resolution coordinate estimation when complemented with multidimensional chemical mapping and Rosetta DRRAFTER computational modeling. Furthermore ,this hybrid ‘Ribosolve’ pipeline detects and falsifies homologies and conformational rearrangements in 11 previously unknown 119- to 338-nucleotide protein-free RNA structures: full-length Tetrahymena ribozyme, hc16 ligase with and without substrate, full-length Vibrio cholerae and Fusobacterium nucleatum glycine riboswitch aptamers with and without glycine, Mycobacterium SAM-IV riboswitch with and without S-adenosylmethionine, and the computer-designed ATP-TTR-3 aptamer with and without AMP. Simulation benchmarks, blind challenges, compensatory mutagenesis, cross-RNA homologies and internal controls demonstrate that Ribosolve can accurately resolve the global architectures of RNA molecules but does not resolve atomic details. These tests offer guidelines for making inferences in future RNA structural studies with similarly accelerated throughput.},
doi = {10.1038/s41592-020-0878-9},
journal = {Nature Methods},
number = 7,
volume = 17,
place = {United States},
year = {Thu Jul 02 00:00:00 EDT 2020},
month = {Thu Jul 02 00:00:00 EDT 2020}
}
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