Quantifying the local resolution of cryo-EM density maps

Kucukelbir, Alp; Sigworth, Fred J.; Tagare, Hemant D.

doi:10.1038/nmeth.2727

Cited by 1,834 publications

(1,475 citation statements)

References 18 publications

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“…This density can be further split along the long sheath axis into two layers. The first layer was resolved to 14 Å resolution (shown in red in Figs 2E and EV5C), and the second layer was resolved to 10 Å resolution at the Hcp tube part and to 12 Å resolution at the VipA/VipB sheath periphery (orange in Figs 2E and EV5C), while rings below are resolved to 7.5 Å resolution as predicted by ResMap (Fig 1D; Kucukelbir et al , 2014). Six sheath subunits can be fitted into the layer that is next to the end of the regular sheath–tube polymer with a correlation coefficient 0.86, suggesting that this layer is composed of sheath–tube ring (topmost ring in Fig EV5C).…”

Section: Resultsmentioning

confidence: 54%

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Cryo‐ EM reconstruction of Type VI secretion system baseplate and sheath distal end

et al. 2017

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The bacterial Type VI secretion system (T6SS) assembles from three major parts: a membrane complex that spans inner and outer membranes, a baseplate, and a sheath–tube polymer. The baseplate assembles around a tip complex with associated effectors and connects to the membrane complex by TssK. The baseplate assembly initiates sheath–tube polymerization, which in some organisms requires TssA. Here, we analyzed both ends of isolated non‐contractile Vibrio cholerae sheaths by cryo‐electron microscopy. Our analysis suggests that the baseplate, solved to an average 8.0 Å resolution, is composed of six subunits of TssE/F2/G and the baseplate periphery is decorated by six TssK trimers. The VgrG/PAAR tip complex in the center of the baseplate is surrounded by a cavity, which may accommodate up to ~450 kDa of effector proteins. The distal end of the sheath, resolved to an average 7.5 Å resolution, shows sixfold symmetry; however, its protein composition is unclear. Our structures provide an important step toward an atomic model of the complete T6SS assembly.

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Section: Resultsmentioning

confidence: 54%

“…The resulting 11 Å resolution model showed the same features as a sixfold symmetrized model and trimeric features of the VgrG spike. Local resolution variations of the baseplate and distal‐end maps were estimated with Resmap (Kucukelbir et al , 2014). …”

Section: Methodsmentioning

confidence: 99%

Cryo‐ EM reconstruction of Type VI secretion system baseplate and sheath distal end

et al. 2017

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“…The Gaussians overlap to provide better sampling of the volume. The resulting resolution map is similar to what is produced by ResMap 85 , but measures FSC (which is filter-independent) unlike ResMap 85 , which requires unfiltered volumes and wouldn’t work well on subtomogram averages. The global resolution of our Cryo-ET map at the standard FSC 0.143 cutoff is ~28 Å and the local resolution distribution ranges from 20 to 38 Å, with the inner ring being in the 20–25 Å range (Extended Data Fig.…”

Section: Methodsmentioning

confidence: 92%

Integrative structure and functional anatomy of a nuclear pore complex

Kim

Fernández-Martı́nez

Nudelman

et al. 2018

Nature

490

894

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Summary Despite the central role of Nuclear Pore Complexes (NPCs) as gatekeepers of RNA and protein transport between the cytoplasm and nucleoplasm, their large size and dynamic nature have impeded a full structural and functional elucidation. Here, we have determined a subnanometer precision structure for the entire 552-protein yeast NPC by satisfying diverse data including stoichiometry, a cryo-electron tomography map, and chemical cross-links. The structure reveals the NPC’s functional elements in unprecedented detail. The NPC is built of sturdy diagonal columns to which are attached connector cables, imbuing both strength and flexibility, while tying together all other elements of the NPC, including membrane-interacting regions and RNA processing platforms. Inwardly-directed anchors create a high density of transport factor-docking Phe-Gly repeats in the central channel, organized in distinct functional units. Taken together, this integrative structure allows us to rationalize the architecture, transport mechanism, and evolutionary origins of the NPC.

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“…All density maps were corrected for the modulation transfer function (MTF) of the K2 summit direct detector and then sharpened by applying temperature-factor that was estimated using post-processing in RELION. Local resolution was determined using ResMap 50 with half-reconstructions as input maps.…”

Section: Methodsmentioning

confidence: 99%

Cryo-EM structure of the activated GLP-1 receptor in complex with a G protein

Zhang

Sun

Feng

et al. 2017

Nature

503

568

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Summary Glucagon-like peptide-1 (GLP-1) is a hormone with essential roles in regulating insulin secretion, carbohydrate metabolism and appetite. GLP-1 effects are mediated through binding to GLP-1R, a family B G protein-coupled receptor (GPCR) signaling primarily through the stimulatory G protein Gs. Family B GPCRs are important therapeutic targets, however our understanding of their mechanism of action is limited by the lack of structural information on activated and full-length receptors. Here we show the electron cryo-microscopy structure of the peptide-activated GLP-1R:Gs complex at near atomic resolution. The peptide is clasped between the N-terminal domain and transmembrane core of the receptor, further stabilized by extracellular loops. Conformational changes in the transmembrane domain result in a sharp kink in the middle of transmembrane helix 6, which pivots its intracellular half outward to accommodate the α5 helix of GαsRas. These results provide a structural framework for understanding family B receptor activation through hormone binding.

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Quantifying the local resolution of cryo-EM density maps

Cited by 1,834 publications

References 18 publications

Cryo‐ EM reconstruction of Type VI secretion system baseplate and sheath distal end

Cryo‐ EM reconstruction of Type VI secretion system baseplate and sheath distal end

Integrative structure and functional anatomy of a nuclear pore complex

Cryo-EM structure of the activated GLP-1 receptor in complex with a G protein

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