TY - JOUR
T1 - Structure, folding and flexibility of co-transcriptional RNA origami
AU - McRae, Ewan K.S.
AU - Rasmussen, Helena Østergaard
AU - Liu, Jianfang
AU - Bøggild, Andreas
AU - Nguyen, Michael T.A.
AU - Sampedro Vallina, Nestor
AU - Boesen, Thomas
AU - Pedersen, Jan Skov
AU - Ren, Gang
AU - Geary, Cody
AU - Andersen, Ebbe Sloth
N1 - Funding Information:
We dedicate our work to the late Ned Seeman, who pioneered the research field of structural nucleic acid nanotechnology and greatly inspired our work and directly or indirectly influenced our careers. We acknowledge the EMBION Cryo-EM Facility at iNANO, Aarhus University, for time under application ID 0042. We thank P. Nissen and A. Briegel for their early support of the project and valuable discussion, S. Sparvath for early design and experiments with the 6HB and A. Briegel and H. C. Høiberg for early NS-TEM characterization of 6HBs. We thank R. Rosendahl and C. Bus for technical assistance. The work at iNANO was supported by the Independent Research Fund Denmark under the Research Project 1 grant (9040-00425B) to E.S.A., the Canadian Natural Sciences and Engineering Research Council post-doctoral fellowship (532417) to E.K.S.M., the Carlsberg Foundation Research Infrastructure grant (CF20-0635) to E.S.A., the European Research Council (ERC) Consolidator grant (683305) to E.S.A. and Novo Nordisk Foundation Ascending Investigator grant (NNF20OC0060694) and Interdisciplinary Synergy grant (NNF21OC0070452) to E.S.A. The work at the Molecular Foundry, Lawrence Berkeley National Laboratory, was supported by the Office of Science, Office of Basic Energy Sciences, of the US Department of Energy (contract no. DE-AC02-05CH11231) and the US National Institutes of Health (nos. R01HL115153, R01 GM104427, R01MH077303 and R01DK042667) to J.L. and G.R.
Funding Information:
We dedicate our work to the late Ned Seeman, who pioneered the research field of structural nucleic acid nanotechnology and greatly inspired our work and directly or indirectly influenced our careers. We acknowledge the EMBION Cryo-EM Facility at iNANO, Aarhus University, for time under application ID 0042. We thank P. Nissen and A. Briegel for their early support of the project and valuable discussion, S. Sparvath for early design and experiments with the 6HB and A. Briegel and H. C. Høiberg for early NS-TEM characterization of 6HBs. We thank R. Rosendahl and C. Bus for technical assistance. The work at iNANO was supported by the Independent Research Fund Denmark under the Research Project 1 grant (9040-00425B) to E.S.A., the Canadian Natural Sciences and Engineering Research Council post-doctoral fellowship (532417) to E.K.S.M., the Carlsberg Foundation Research Infrastructure grant (CF20-0635) to E.S.A., the European Research Council (ERC) Consolidator grant (683305) to E.S.A. and Novo Nordisk Foundation Ascending Investigator grant (NNF20OC0060694) and Interdisciplinary Synergy grant (NNF21OC0070452) to E.S.A. The work at the Molecular Foundry, Lawrence Berkeley National Laboratory, was supported by the Office of Science, Office of Basic Energy Sciences, of the US Department of Energy (contract no. DE-AC02-05CH11231) and the US National Institutes of Health (nos. R01HL115153, R01 GM104427, R01MH077303 and R01DK042667) to J.L. and G.R.
Publisher Copyright:
© 2023, The Author(s), under exclusive licence to Springer Nature Limited.
PY - 2023/7
Y1 - 2023/7
N2 - RNA origami is a method for designing RNA nanostructures that can self-assemble through co-transcriptional folding with applications in nanomedicine and synthetic biology. However, to advance the method further, an improved understanding of RNA structural properties and folding principles is required. Here we use cryogenic electron microscopy to study RNA origami sheets and bundles at sub-nanometre resolution revealing structural parameters of kissing-loop and crossover motifs, which are used to improve designs. In RNA bundle designs, we discover a kinetic folding trap that forms during folding and is only released after 10 h. Exploration of the conformational landscape of several RNA designs reveal the flexibility of helices and structural motifs. Finally, sheets and bundles are combined to construct a multidomain satellite shape, which is characterized by individual-particle cryo-electron tomography to reveal the domain flexibility. Together, the study provides a structural basis for future improvements to the design cycle of genetically encoded RNA nanodevices.
AB - RNA origami is a method for designing RNA nanostructures that can self-assemble through co-transcriptional folding with applications in nanomedicine and synthetic biology. However, to advance the method further, an improved understanding of RNA structural properties and folding principles is required. Here we use cryogenic electron microscopy to study RNA origami sheets and bundles at sub-nanometre resolution revealing structural parameters of kissing-loop and crossover motifs, which are used to improve designs. In RNA bundle designs, we discover a kinetic folding trap that forms during folding and is only released after 10 h. Exploration of the conformational landscape of several RNA designs reveal the flexibility of helices and structural motifs. Finally, sheets and bundles are combined to construct a multidomain satellite shape, which is characterized by individual-particle cryo-electron tomography to reveal the domain flexibility. Together, the study provides a structural basis for future improvements to the design cycle of genetically encoded RNA nanodevices.
KW - RNA/chemistry
KW - Nanotechnology/methods
KW - Nanostructures/chemistry
KW - Molecular Conformation
KW - Nanomedicine
KW - Nucleic Acid Conformation
UR - http://www.scopus.com/inward/record.url?scp=85148959656&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85148959656&partnerID=8YFLogxK
U2 - 10.1038/s41565-023-01321-6
DO - 10.1038/s41565-023-01321-6
M3 - Article
C2 - 36849548
AN - SCOPUS:85148959656
SN - 1748-3387
VL - 18
SP - 808
EP - 817
JO - Nature Nanotechnology
JF - Nature Nanotechnology
IS - 7
ER -