Functionalized graphene-oxide grids enable high-resolution cryo-EM structures of the SNF2h-nucleosome complex without crosslinking
Year: 2024
Type: article
Abstract: 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.
Source: Nature Communications
Language: English
Subfield: Structural Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $6,790
Funders: Howard Hughes Medical Institute, U.S. Department of Health and Human Services, Lundbeckfonden, Novo Nordisk, Danmarks Frie Forskningsfond, Novo Nordisk Fonden, H. Lundbeck A/S, National Institutes of Health, University of California, San Francisco, Stanford Bio-X, National Institute of General Medical Sciences
Awards: G1414818376, Molecular Biophysics Training Grant, Molecular mechanisms of the ACF chromatin remodeling complex, G2219480109, Glacios™ Cryo Transmission Electron Microscope with 200 kV XFEG optics, Acquisition of an electron microscope for high-resolution single particle cryo-EM, G3865440040, Advancing cryo-EM technology to address difficult biological questions, G3998664971, G4455162931, G5134627149, G5146311565, G5542357563, G575095112, G6335145789, G670161810, G7292143881, Linux cluster for near atomic resolution single particle cryo-EM, G857049419, G8873047370, G9000203011, ATP-dependent and independent mechanisms of regulating chromatin states
OpenAlex ID: W4392683734