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TitleFunctionalized graphene-oxide grids enable high-resolution cryo-EM structures of the SNF2h-nucleosome complex without crosslinking.
Journal, issue, pagesNat Commun, Vol. 15, Issue 1, Page 2225, Year 2024
Publish dateMar 12, 2024
AuthorsUn Seng Chio / Eugene Palovcak / Anton A A Smith / Henriette Autzen / Elise N Muñoz / Zanlin Yu / Feng Wang / David A Agard / Jean-Paul Armache / Geeta J Narlikar / Yifan Cheng /
PubMed AbstractSingle-particle cryo-EM is widely used to determine enzyme-nucleosome complex structures. However, cryo-EM sample preparation remains challenging and inconsistent due to complex denaturation at the ...Single-particle cryo-EM is widely used to determine enzyme-nucleosome complex structures. However, cryo-EM sample preparation remains challenging and inconsistent due to complex denaturation at the air-water interface (AWI). Here, to address this issue, we develop graphene-oxide-coated EM grids functionalized with either single-stranded DNA (ssDNA) or thiol-poly(acrylic acid-co-styrene) (TAASTY) co-polymer. These grids protect complexes between the chromatin remodeler SNF2h and nucleosomes from the AWI and facilitate collection of high-quality micrographs of intact SNF2h-nucleosome complexes in the absence of crosslinking. The data yields maps ranging from 2.3 to 3 Å in resolution. 3D variability analysis reveals nucleotide-state linked conformational changes in SNF2h bound to a nucleosome. In addition, the analysis provides structural evidence for asymmetric coordination between two SNF2h protomers acting on the same nucleosome. We envision these grids will enable similar detailed structural analyses for other enzyme-nucleosome complexes and possibly other protein-nucleic acid complexes in general.
External linksNat Commun / PubMed:38472177 / PubMed Central
MethodsEM (single particle)
Resolution2.3 - 3.2 Å
Structure data

42977

8v4y

EMDB-42977, PDB-8v4y:
Cryo-EM structure of singly-bound SNF2h-nucleosome complex with SNF2h at inactive SHL2 (conformation 1)
Method: EM (single particle) / Resolution: 2.8 Å

EMDB-43000: Cryo-EM structure of SNF2h-nucleosome complex (consensus structure)
Method: EM (single particle) / Resolution: 2.3 Å

EMDB-43001: Cryo-EM structure of SNF2h-nucleosome complex (single-bound structure)
Method: EM (single particle) / Resolution: 2.5 Å

43002

8v6v

EMDB-43002, PDB-8v6v:
Cryo-EM structure of doubly-bound SNF2h-nucleosome complex
Method: EM (single particle) / Resolution: 2.8 Å

43003

8v7l

EMDB-43003, PDB-8v7l:
Cryo-EM structure of singly-bound SNF2h-nucleosome complex with SNF2h at inactive SHL2 (conformation 2)
Method: EM (single particle) / Resolution: 2.9 Å

EMDB-43004: Cryo-EM structure of doubly-bound SNF2h-nucleosome complex (conformation 1)
Method: EM (single particle) / Resolution: 3.1 Å

EMDB-43005: Cryo-EM structure of doubly-bound SNF2h-nucleosome complex (conformation 2)
Method: EM (single particle) / Resolution: 3.2 Å

Chemicals

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM

ChemComp-MG:
Unknown entry

ChemComp-BEF:
BERYLLIUM TRIFLUORIDE ION

Source
  • homo sapiens (human)
  • xenopus laevis (African clawed frog)
  • synthetic construct (others)
KeywordsDNA BINDING PROTEIN/DNA / nucleosome / chromatin remodeler / ISWI / SNF2h / DNA BINDING PROTEIN-DNA complex

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