|Entry||Database: EMDB / ID: 0459|
|Title||Structure of Dot1L-H2BK120ub nucleosome complex|
|Sample||Dot1L-H2BK120ub nucleosome complex (Classes 3 and 4):|
|Source||Homo sapiens (human)|
|Method||single particle reconstruction / cryo EM / 3.5 Å resolution|
|Authors||Anderson CJ / Baird MR / Hsu A / Barbour EH / Koyama Y / Borgnia MJ / McGinty RK|
|Citation||Journal: Cell Rep / Year: 2019|
Title: Structural Basis for Recognition of Ubiquitylated Nucleosome by Dot1L Methyltransferase.
Authors: Cathy J Anderson / Matthew R Baird / Allen Hsu / Emily H Barbour / Yuka Koyama / Mario J Borgnia / Robert K McGinty
Abstract: Histone H3 lysine 79 (H3K79) methylation is enriched on actively transcribed genes, and its misregulation is a hallmark of leukemia. Methylation of H3K79, which resides on the structured disk face of ...Histone H3 lysine 79 (H3K79) methylation is enriched on actively transcribed genes, and its misregulation is a hallmark of leukemia. Methylation of H3K79, which resides on the structured disk face of the nucleosome, is mediated by the Dot1L methyltransferase. Dot1L activity is part of a trans-histone crosstalk pathway, requiring prior histone H2B ubiquitylation of lysine 120 (H2BK120ub) for optimal activity. However, the molecular details describing both how Dot1L binds to the nucleosome and why Dot1L is activated by H2BK120 ubiquitylation are unknown. Here, we present the cryoelectron microscopy (cryo-EM) structure of Dot1L bound to a nucleosome reconstituted with site-specifically ubiquitylated H2BK120. The structure reveals that Dot1L engages the nucleosome acidic patch using a variant arginine anchor and occupies a conformation poised for methylation. In this conformation, Dot1L and ubiquitin interact directly through complementary hydrophobic surfaces. This study establishes a path to better understand Dot1L function in normal and leukemia cells.
|Date||Deposition: Jan 14, 2019 / Header (metadata) release: Feb 6, 2019 / Map release: Feb 13, 2019 / Last update: Feb 27, 2019|
|Structure viewer||EM map: |
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|File||emd_0459.map.gz (map file in CCP4 format, 143749 KB)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.08 Å|
CCP4 map header:
-Entire Dot1L-H2BK120ub nucleosome complex (Classes 3 and 4)
|Entire||Name: Dot1L-H2BK120ub nucleosome complex (Classes 3 and 4) / Number of components: 1|
-Component #1: protein, Dot1L-H2BK120ub nucleosome complex (Classes 3 and 4)
|Protein||Name: Dot1L-H2BK120ub nucleosome complex (Classes 3 and 4) / Recombinant expression: No|
|Mass||Theoretical: 270 kDa|
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Escherichia coli (E. coli)|
|Specimen||Specimen state: particle / Method: cryo EM|
|Sample solution||Specimen conc.: 0.93 mg/ml / pH: 7.5|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277.15 K / Humidity: 100 %|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Imaging||Microscope: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 48 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Imaging mode: BRIGHT FIELD|
|Specimen Holder||Model: OTHER|
|Camera||Detector: FEI FALCON III (4k x 4k)|
|Image acquisition||Number of digital images: 1000|
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