DNA origami signposts for identifying proteins on cell membranes by electron cryotomography
Electron cryotomography (cryoET), an electron cryomicroscopy (cryoEM) modality, has changed our understanding of biological function by revealing the native molecular details of membranes, viruses, and cells. However, identification of individual molecules within tomograms from cryoET is challenging...
| Main Authors: | , , , , , , , , , , |
|---|---|
| Format: | Journal article |
| Language: | English |
| Published: |
Cell Press
2021
|
| _version_ | 1797077538428682240 |
|---|---|
| author | Silvester, E Vollmer, B Pražák, V Vasishtan, D Machala, EA Whittle, C Black, S Bath, J Turberfield, AJ Grünewald, K Baker, LA |
| author_facet | Silvester, E Vollmer, B Pražák, V Vasishtan, D Machala, EA Whittle, C Black, S Bath, J Turberfield, AJ Grünewald, K Baker, LA |
| author_sort | Silvester, E |
| collection | OXFORD |
| description | Electron cryotomography (cryoET), an electron cryomicroscopy (cryoEM) modality, has changed our understanding of biological function by revealing the native molecular details of membranes, viruses, and cells. However, identification of individual molecules within tomograms from cryoET is challenging because of sample crowding and low signal-to-noise ratios. Here, we present a tagging strategy for cryoET that precisely identifies individual protein complexes in tomograms without relying on metal clusters. Our method makes use of DNA origami to produce “molecular signposts” that target molecules of interest, here via fluorescent fusion proteins, providing a platform generally applicable to biological surfaces. We demonstrate the specificity of signpost origami tags (SPOTs) in vitro as well as their suitability for cryoET of membrane vesicles, enveloped viruses, and the exterior of intact mammalian cells. |
| first_indexed | 2024-03-07T00:19:31Z |
| format | Journal article |
| id | oxford-uuid:7c0ab4ff-8681-4ab0-9609-f33f4dd5c9ed |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T00:19:31Z |
| publishDate | 2021 |
| publisher | Cell Press |
| record_format | dspace |
| spelling | oxford-uuid:7c0ab4ff-8681-4ab0-9609-f33f4dd5c9ed2022-03-26T20:54:34ZDNA origami signposts for identifying proteins on cell membranes by electron cryotomographyJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:7c0ab4ff-8681-4ab0-9609-f33f4dd5c9edEnglishSymplectic ElementsCell Press2021Silvester, EVollmer, BPražák, VVasishtan, DMachala, EAWhittle, CBlack, SBath, JTurberfield, AJGrünewald, KBaker, LAElectron cryotomography (cryoET), an electron cryomicroscopy (cryoEM) modality, has changed our understanding of biological function by revealing the native molecular details of membranes, viruses, and cells. However, identification of individual molecules within tomograms from cryoET is challenging because of sample crowding and low signal-to-noise ratios. Here, we present a tagging strategy for cryoET that precisely identifies individual protein complexes in tomograms without relying on metal clusters. Our method makes use of DNA origami to produce “molecular signposts” that target molecules of interest, here via fluorescent fusion proteins, providing a platform generally applicable to biological surfaces. We demonstrate the specificity of signpost origami tags (SPOTs) in vitro as well as their suitability for cryoET of membrane vesicles, enveloped viruses, and the exterior of intact mammalian cells. |
| spellingShingle | Silvester, E Vollmer, B Pražák, V Vasishtan, D Machala, EA Whittle, C Black, S Bath, J Turberfield, AJ Grünewald, K Baker, LA DNA origami signposts for identifying proteins on cell membranes by electron cryotomography |
| title | DNA origami signposts for identifying proteins on cell membranes by electron cryotomography |
| title_full | DNA origami signposts for identifying proteins on cell membranes by electron cryotomography |
| title_fullStr | DNA origami signposts for identifying proteins on cell membranes by electron cryotomography |
| title_full_unstemmed | DNA origami signposts for identifying proteins on cell membranes by electron cryotomography |
| title_short | DNA origami signposts for identifying proteins on cell membranes by electron cryotomography |
| title_sort | dna origami signposts for identifying proteins on cell membranes by electron cryotomography |
| work_keys_str_mv | AT silvestere dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT vollmerb dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT prazakv dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT vasishtand dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT machalaea dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT whittlec dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT blacks dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT bathj dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT turberfieldaj dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT grunewaldk dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography AT bakerla dnaorigamisignpostsforidentifyingproteinsoncellmembranesbyelectroncryotomography |