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BMC Biology (1)
The Biochemical journal (1)
Brzovic, Peter S. (1)
KLEVIT, Rachel E. (1)
Klevit, Rachel E (1)
Klevit, Rachel E. (1)
Lissounov, Alexei (1)
STOLL, Kate E. (1)
WENZEL, Dawn M. (1)
Wenzel, Dawn M (1)
Wenzel, Dawn M. (1)
Year of Publication
UbcH7 reactivity profile reveals Parkin and HHARI to be RING/HECT hybrids
Brzovic, Peter S.
Klevit, Rachel E.
Although the functional interaction between ubiquitin conjugating enzymes (E2s) and ubiquitin ligases (E3s) is essential in ubiquitin (Ub) signaling, the criteria that define an active E2–E3 pair are not well-established. The human E2 UbcH7 (Ube2L3) shows broad specificity for HECT-type E3s1, but often fails to function with RING E3s in vitro despite forming specific complexes2–4. Structural comparisons of inactive UbcH7/RING complexes with active UbcH5/RING complexes reveal no defining differences3,4, highlighting a gap in our understanding of Ub transfer. We show that, unlike many E2s that transfer Ub with RINGs, UbcH7 lacks intrinsic, E3-independent reactivity with lysine, explaining its preference for HECTs. Despite lacking lysine reactivity, UbcH7 exhibits activity with the RING-In Between-RING (RBR) family of E3s that includes Parkin and human homologue of ariadne (HHARI)5,6. Found in all eukaryotes7, RBRs regulate processes such as translation8 and immune signaling9. RBRs contain a canonical C3HC4-type RING, followed by two conserved Cys/His-rich Zn2+-binding domains, In-Between-RING (IBR) and RING2 domains, which together define this E3 family7. Here we show that RBRs function like RING/HECT hybrids: they bind E2s via a RING domain, but transfer Ub through an obligate thioester-linked Ub (denoted ‘~Ub’), requiring a conserved cysteine residue in RING2. Our results define the functional cadre of E3s for UbcH7, an E2 involved in cell proliferation10 and immune function11, and suggest a novel mechanism for an entire class of E3s.
Following Ariadne's thread: a new perspective on RBR ubiquitin ligases
Klevit, Rachel E
Ubiquitin signaling pathways rely on E3 ligases for effecting the final transfer of ubiquitin from E2 ubiquitin conjugating enzymes to a protein target. Here we re-evaluate the hybrid RING/HECT mechanism used by the E3 family RING-between-RINGs (RBRs) to transfer ubiquitin to substrates. We place RBRs into the context of current knowledge of HECT and RING E3s. Although not as abundant as the other types of E3s (there are only slightly more than a dozen RBR E3s in the human genome), RBRs are conserved in all eukaryotes and play important roles in biology. Re-evaluation of RBR ligases as RING/HECT E3s provokes new questions and challenges the field.
E2s: Structurally Economical and Functionally Replete
STOLL, Kate E.
KLEVIT, Rachel E.
The Biochemical journal
Ubiquitination is a post-translational modification pathway involved in myriad cellular regulation and disease pathways. The ubiquitin (Ub) transfer cascade requires three enzyme activities: a Ub-activating (E1) enzyme, a Ub-conjugating (E2) enzyme, and a Ub ligase (E3). Because the E2 is responsible both for E3 selection and substrate modification, E2s function at the heart of the Ub transfer pathway and are responsible for much of the diversity of Ub cellular signaling. There are currently over ninety three-dimensional structures of E2s, both alone and in complex with protein binding partners, providing a wealth of information regarding how E2s are recognized by a wide variety of proteins. In this review, we describe the prototypical E2/E3 interface and discuss limitations of current methods to identify cognate E2/E3 partners. We present non-canonical E2-protein interactions and highlight the economy of E2s in their ability to facilitate many protein-protein interactions at nearly every surface on their relatively small, compact catalytic domain. Lastly, we compare the structures of conjugated E2~Ub species, their unique protein interactions, and the mechanistic insights provided by species that are poised to transfer Ub.
ubiquitin; ubiquitin-conjugating enzyme; ubiquitin ligase enzyme; ubiquitination; ubiquitylation
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