4WHV: E3 ubiquitin-protein ligase RNF8 in complex with Ubiquitin-conjugating enzyme E2 N and Polyubiquitin-B

DNA double strand break (DSB) responses depend on the sequential actions of the E3 ubiquitin ligases RNF8 and RNF168 plus E2 ubiquitin-conjugating enzyme Ubc13 to specifically generate histone Lys-63-linked ubiquitin chains in DSB signaling. Here, we defined the activated RNF8-Ubc13 approximately ubiquitin complex by x-ray crystallography and its functional solution conformations by x-ray scattering, as tested by separation-of-function mutations imaged in cells by immunofluorescence. The collective results show that the RING E3 RNF8 targets E2 Ubc13 to DSB sites and plays a critical role in damage signaling by stimulating polyubiquitination through modulating conformations of ubiquitin covalently linked to the Ubc13 active site. Structure-guided separation-of-function mutations show that the RNF8 E2 stimulating activity is essential for DSB signaling in mammalian cells and is necessary for downstream recruitment of 53BP1 and BRCA1. Chromatin-targeted RNF168 rescues 53BP1 recruitment involved in non-homologous end joining but not BRCA1 recruitment for homologous recombination. These findings suggest an allosteric approach to targeting the ubiquitin-docking cleft at the E2-E3 interface for possible interventions in cancer and chronic inflammation, and moreover, they establish an independent RNF8 role in BRCA1 recruitment.
PDB ID: 4WHVDownload
MMDB ID: 133051
PDB Deposition Date: 2014/9/23
Updated in MMDB: 2017/12
Experimental Method:
x-ray diffraction
Resolution: 8.3  Å
Source Organism:
Similar Structures:
Biological Unit for 4WHV: hexameric; determined by author
Molecular Components in 4WHV
Label Count Molecule
Proteins (6 molecules)
Ubiquitin-conjugating Enzyme E2 N(Gene symbol: UBE2N)
Molecule annotation
E3 Ubiquitin-protein Ligase Rnf8(Gene symbol: RNF8)
Molecule annotation
Polyubiquitin-b(Gene symbol: UBB)
Molecule annotation
Chemicals (4 molecules)
* Click molecule labels to explore molecular sequence information.

Citing MMDB