An ankyrin-repeat ubiquitin-binding domain determines TRABID's specificity for atypical ubiquitin chains

Abstract

Eight different types of ubiquitin (Ub) linkages are present in eukaryotic cells that regulate diverse biological processes. Proteins that mediate specific assembly and disassembly of atypical Lys6, Lys27, Lys29 and Lys33 linkages are largely unknown. We here reveal how the human Ovarian Tumor (OTU) domain deubiquitinase (DUB) TRABID specifically hydrolyzes both Lys29- and Lys33-linked diubiquitin (diUb). A crystal structure of the extended catalytic domain reveals an unpredicted Ankyrin repeat (Ank) domain that precedes an A20-like catalytic core. NMR analysis identifies the Ank domain as a new Ub binding fold termed AnkUBD, and DUB assays in vitro and in vivo show that this domain is crucial for TRABID efficiency and linkage-specificity. Our data are consistent with a role of the AnkUBD as an enzymatic S1' Ub binding site, which orients a Ub chain such that Lys29 and Lys33 linkages are cleaved preferentially.

HectD1 E3 Ligase Modifies Adenomatous Polyposis Coli (APC) with Polyubiquitin to Promote the APC-Axin Interaction

“…1) and HectD1 (data not shown) revealed a near majority of Lys-29 linkages, suggesting that these enzymes are primarily associated with Lys-29 polyubiquitin-specific DUB and E3 ligase activities, respectively. This would be consistent with the potent DUB activity of an extended Trabid catalytic domain toward Lys-29-linked diubiquitin in vitro (38). Nonetheless, we also detected significant amounts of Lys-48 linkages co-precipitating with Trabid, and these chains do not appear to be conjugated to Trabid itself; a conclusion based on examining the ubiquitylation of HA-Trabid in an anti-Lys-48 immunoprecipitation assay (data not shown).…”

“…2B) (24). Second, full-length, wild type Trabid hydrolyzes Lys-63-linked chains (24,(37)(38)(39). Third, although Lys-48 and Lys-29 chains were the most abundant ubiquitin polymers that co-precipitated with Trabid C443S, Lys-63 chains were also detected, and the majority of these were present in the HMW regions a and b (Fig.…”

TRPV4 channel activity is modulated by direct interaction of the ankyrin domain to PI(4,5)P2

“…The ARD is present in more than 500 human proteins, with diverse binding specificities and affinities 49 . This characteristic of ARDs is thought to be based on the high diversity of their surfaces, because of various numbers and combinations of ankyrin repeats 50 . Consistently, artificial ankyrin repeat proteins (DARpin) can be selected for targeting to specific proteins from a library of artificial ARDs, but have not been rationally designed 51 .…”

Deubiquitylases From Genes to Organism

“…Remarkably, the OTU family catalytic domains display a wide diversity in chain preferences, despite a high degree of structural conservation. For example, OTUB1 and OTUB2 show opposite proclivities for Lys63-and Lys48-linked chains, respectively, while Cezanne/OTU7B and TRA-BID are most active towards Lys11-and Lys29-linked chains, respectively (27,60,154,268,270 …”