UBE2D1

UBE2D1 (UbcH5A) is a promiscuous E2 ubiquitin-conjugating enzyme that occupies the midpoint of the E1–E2–E3 conjugation cascade and broadly supports ubiquitin-mediated protein degradation [PMID:bio_10.1101_2024.09.02.610852]. It engages E3 ligases and substrates through both its RING-binding and backside surfaces, and an engineered inhibitor blocking both sites simultaneously phenocopies UBE2D knockdown in HeLa cells, increasing the abundance of ~20% of detected proteins and sensitizing cells to cisplatin—consistent with impaired degradation and proteotoxic stress [PMID:bio_10.1101_2024.09.02.610852]. In ribosome-associated quality control, UBE2D1 charged with ubiquitin acts with the RING E3 Listerin, where the adaptor TCF25 binds Listerin's RING domain and the acceptor ubiquitin to orient its K48 for nucleophilic attack on the UBE2D1~Ub thioester, thereby imposing K48-linkage specificity on nascent-chain ubiquitination [PMID:bio_10.1101_2024.10.17.618946]. The gene was also shown to correspond to the previously reported iron-transport stimulator SFT, clarifying that those data reflect UbcH5A mRNA, although iron loading or chelation did not regulate UbcH5A in HepG2 cells (PMID:12480712).

1. 2002 Medium

   Resolved the molecular identity of the putative iron-transport factor SFT, establishing that the relevant sequence is UBE2D1 (UbcH5A) rather than a distinct iron transporter.

   Evidence Sequence analysis of SFT cDNA against the UbcH5A locus, RT-PCR, and iron loading/chelation in HepG2 cells

   PMID:12480712

   Open questions at the time

   • No demonstration that UbcH5A is regulated by iron status
   • Does not define UBE2D1's catalytic role in HIF-1α ubiquitination directly
   • Single-lab molecular identity assignment
2. 2024 Medium

   Defined how E3-adaptor context confers chain-linkage specificity on a promiscuous E2, showing TCF25 orients the acceptor ubiquitin's K48 toward the UBE2D1~Ub thioester during Listerin-mediated quality-control ubiquitination.

   Evidence In vitro biochemical reconstitution with AlphaFold3 modeling and binding studies of TCF25–Listerin RING and TCF25–acceptor Ub interactions (preprint)

   PMID:bio_10.1101_2024.10.17.618946

   Open questions at the time

   • Preprint, single lab; awaits peer review and experimental structure
   • Specificity mechanism shown for the Listerin/RQC context only
   • Cellular relevance of the modeled orientation not tested in vivo
3. 2024 Medium

   Established that simultaneous blockade of UBE2D1's RING- and backside-binding surfaces is sufficient to impair bulk ubiquitin-dependent degradation, validating the dual-surface engagement model of E2 function.

   Evidence Engineered multivalent protein inhibitor with defined binding sites; HeLa knockdown/inhibitor treatment with whole-cell proteomics and cisplatin sensitivity (preprint)

   PMID:bio_10.1101_2024.09.02.610852

   Open questions at the time

   • Preprint, single lab; not independently replicated
   • Cross-reactivity across the UBE2D family not fully resolved
   • Specific substrates driving the ~20% proteome increase not enumerated

• The full spectrum of E3 partners, substrates, and linkage outcomes that UBE2D1 mediates beyond the Listerin/RQC pathway remains undefined.
• No comprehensive E3 partner map in the corpus
• Chain-type preferences outside K48/Listerin context uncharacterized
• No experimental structure of UBE2D1 in the corpus