Insights into Ubiquitination from the Unique Clamp-like Binding of the RING E3 AO7 to the E2 UbcH5B

Abstract

Background:The RING E3 AO7/RNF25 binds its E2 with unusually high affinity. Results: AO7 has a secondary E2 binding site adjacent to the RING. Conclusion: This site prevents the stimulatory effect of non-covalent backside binding of ubiquitin, has unique agonist properties, and allows for structural analysis of RING mutants. Significance: Knowledge of how RING E3s mediate ubiquitination is critical to understanding cellular protein regulation.

Structural Studies of HHARI/UbcH7∼Ub Reveal Unique E2∼Ub Conformational Restriction by RBR RING1

“…The overall architecture of the RING1/UbcH7 interface is similar to those in other RING/E2 pairs (Brown et al., 2014, Brzovic et al., 2003, DaRosa et al., 2015, Dou et al., 2012, Koliopoulos et al., 2016, Lechtenberg et al., 2016, Li et al., 2015, Plechanovova et al., 2012, Pruneda et al., 2012, Scott et al., 2014, Xu et al., 2008, Zheng et al., 2000). UbcH7 residues from loops 4 and 7 and helix 1 and HHARI RING1 residues from both Zn 2+ loops and the central helix make canonical E2/RING interactions (Figures 3A and S1A).…”

Structural basis of generic versus specific E2–RING E3 interactions in protein ubiquitination

“…When bound to UBE2D2, D2E3BR abolishes processive ubiquitination, via a mechanism similar to that seen in UBE2B‐RAD18 . Though, the biological significance of this is not yet known . On the other hand, D2E3BR binding to UBE2E3, which normally monoubiquitinates its substrates, promotes processivity by forcing UBE2E3~Ub in close‐state, thereby increasing UBE2E3~Ub:RNF25 reactivity …”

“…(a) Ribbon representation of a single RNF4 monomer from UBE2D1~Ub:RNF4 complex (PDB: 4AP4) showing the zinc ions as spheres, the residues involved in zinc‐coordination and the allosteric linchpin residue as ball‐and‐sticks, and demarcating the E2‐binding site. The ribbon diagram is colored to reflect the average pairwise positional shift of each overlapping C α atom between RNF4 and unique RING cores from all available E2–RING E3 complex structures including RNF146 (PDB: 4QPL), TRIM25 (PDB: 5FER), BIRC2 (PDB: 6HPR), BIRC3 (PDB: 3 EB6), BIRC7 (PDB: 4AUQ), RNF38 (PDB: 4V3K), RNF25 (PDB: 5D1M), RNF165 (PDB: 5D0M), MDM2 (PDB: 5MNJ), TRIM23 (PDB: 5VZW), RNF13 (PDB: 5ZBU), E4B (PDB: 3L1Z), RNF2 (PDB: 3RPG), GP78 (PDB: 2LXP), SIZ1 (PDB: 5JNE), c‐CBL (PDB: 1FBV), RBX1 (PDB: 4P5O), TRAF6 (PDB: 3HCT), TRIM5 (PDB: 4TKP), RNF8 (PDB: 4WHV), ZNRF1 (PDB: 5YWR), LNX1 (PDB: 5H7S), CHIP (PDB: 2C2V), and FANCL (PDB: 2CCG) . (b) UBE2D1~Ub:RNF4 complex highlighting position of the RING domain relative to the E2, with the allosteric linchpin residue coordinating E2 as well as the donor Ub shown as ball‐and‐sticks.…”

“…(b) Ribbon representation of UBE2B-RAD18 (PDB: 1YBF), 71 showing the zinc ions as spheres, residues involved the specialized E2-RING E3 interaction as ball-and-sticks, and hydrogen-bonding interactions as dashed lines. (c) Ribbon representation of UBE2D2-RNF25 (PDB: 5D1M), 73 showing the zinc ions as spheres, residues involved the specialized E2-RING E3 interaction as ball-and-sticks, and hydrogenbonding interactions as dashed lines degradation. 67,68 Toward this end GP78 sequester UBE2G2 to the ER via a specialized helix distal to its RING core, termed the UBE2G2-binding region (G2BR).…”

Mechanism and disease association of E2-conjugating enzymes: lessons from UBE2T and UBE2L3

“…In the UBE2D family, this surface supports non-covalent ubiquitin interactions that stimulate processivity of the E2 enzyme [68,69]. Several RING E3s also harbour ancillary E2 backside-binding regions that induce different allosteric effects, such as stabilization of the canonical RING–E2 interface, E2∼Ub charging, and processive ubiquitination [70–74]. Interestingly, in certain E2s, the backside interaction with E3s can regulate the type of ubiquitin signal generated on substrates [75,76].…”