PROSITE documentation PDOC52053

Novel E3 ligase (NEL) domain profile

The ubiquitin system regulates diverse cellular processes in all eukaryotes through the covalent attachment of ubiquitin to acceptor lysine residues on target substrates, thereby dictating protein stability, activity, and localization. Ubiquitination involves one ubiquitin-activating enzyme (E1), a limited number of ubiquitin-conjugating enzymes (E2s), and a large number of ubiquitin-ligating enzymes (E3s). E3 ligases determine the substrate specificity of ubiquitination and have been classified into two main classes, the HECT (homologous with E6-associated protein C-terminus) and RING (really interesting new gene) finger families. RING (see <PDOC00449>) and U-box (a modified RING motif) (see <PDOC51698>) domain-containing E3s act as adaptor-like molecules by bringing a ubiquitinated E2 and the substrate into sufficiently close proximity to promote the ubiquitination of the substrate. In contrast, HECT domain-containing E3s possess an essential cysteine residue that acts as an acceptor for the ubiquitin carried by the E2 before its transfer to the substrate [1,2,3,4].

Although ubiquitin is absent from bacteria, some species secrete effectors displaying E3 activity, which are able to subvert the ubiquitination pathway of the eukaryotic host cell. A family of bacterial effectors with a novel E3 ubiquitin ligase (NEL) domain, which is structurally distinct from the RING- and HECT-type E3 ubiquitin ligase domains has been identified from several animal- and plant-pathogenic bacteria, and several plant-symbiotic bacteria, such as Shigella spp., Salmonella enterica, Yersinia spp., P. syringae and Rhizobium spp.. The ~300 residue NEL domain bears a cysteine residue that mediates transfer of ubiquitin to substrates through the formation of a thioester intermediate, analogous to the structurally unrelated HECT domain in eukaryotes. In almost all NEL-domain-containing effectors, a leucine-rich repeat (LRR) domain (see <PDOC51450>) is paired with the NEL domain. The LRR domain of NEL effectors is required not only for substrate recognition by binding to target proteins, but also for the autoinhibition of its E3 ubiquitin ligase activity by sequestering the catalytic cysteine residue of the NEL domain in the absence of target proteins [1,2,3,5].

The NEL domain adopts a complete α-helical fold consisting of 12 helices and bearing no resemblance to previously defined E3 ubiquitin ligases (see <PDB:3CVR>). The core structure is the two antiparallel helix pairs (a8-a9 and a9-a10), with other helices wrapping around them. The catalytic cysteine residue located within a conserved CXD motif of the NEL domain acts as an essential catalytic nucleophile in the ubiquitination reaction and functions analogously to the catalytic cysteine in HECT-type E3 ligases. The catalysis of ubiquitin transfer mediated by the NEL domain also requires the conserved aspartic acid within the CXD motif in addition to the cysteine residue functioning as a thiol nucleophile [3,4,6].

The profile we developed covers the entire NEL domain.