DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways

BACKGROUND: RBBP6 is a 250 kDa splicing-associated protein that has been identified as an E3 ligase due to the presence of a RING finger domain. In humans and mice it interacts with both p53 and Rb, and plays a role in the induction of apoptosis and regulation of the cell cycle. RBBP6 has recently been shown to be highly up-regulated in oesophageal cancer, and to be a promising target for immunotherapy against the disease. RESULTS: We show here using heteronuclear NMR that the N-terminal 81 amino acids of RBBP6 constitute a novel ubiquitin-like domain, which we have called the DWNN domain. The domain lacks conserved equivalents of K(48 )and K(63), although the equivalents of K(6 )and K(29 )are highly, although not absolutely, conserved. The di-glycine motif that is characteristic of proteins involved in ubiquitination is found in the human and mouse form of the domain, although it is not present in all organisms. It forms part of a three-domain form of RBBP6 containing the DWNN domain, a zinc knuckle and a RING finger domain, which is found in all eukaryotic genomes so far examined, in the majority of cases at single copy number. The domain is also independently expressed in vertebrates as a single domain protein. CONCLUSION: DWNN is a novel ubiquitin-like domain found only at the N-terminus of the RBBP6 family of splicing-associated proteins. The ubiquitin-like structure of the domain greatly increases the likelihood that RBBP6 functions through some form of ubiquitin-like modification. Furthermore, the fact that the DWNN domain is independently expressed in higher vertebrates leads us to propose that the domain may itself function as a novel ubiquitin-like modifier of other proteins

DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways-4

<p><b>Copyright information:</b></p><p>Taken from "DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways"</p><p>BMC Structural Biology 2006;6():1-1.</p><p>Published online 5 Jan 2006</p><p>PMCID:PMC1360078.</p><p></p>nge (

DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways-0

<p><b>Copyright information:</b></p><p>Taken from "DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways"</p><p>BMC Structural Biology 2006;6():1-1.</p><p>Published online 5 Jan 2006</p><p>PMCID:PMC1360078.</p><p></p> eukaryotic genomes analysed to date, including the single celled parasite , in which it is very much reduced in size. In vertebrates and insects the protein includes a long C-terminal extension containing p53 and Rb-interaction domains in human and mouse. A short form consisting of the DWNN domain and a poorly conserved C-terminal tail is also found in vertebrates

DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways-3

<p><b>Copyright information:</b></p><p>Taken from "DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways"</p><p>BMC Structural Biology 2006;6():1-1.</p><p>Published online 5 Jan 2006</p><p>PMCID:PMC1360078.</p><p></p>and secondary structure. The backbone adopts a ubiquitin-like β-grasp fold in which the central α-helix packs against a five-stranded β-sheet comprised of strands β, β, β, βand β. Unlike ubiquitin, DWNN contains an additional double-stranded β-sheet at the N-terminal end of the central α-helix, comprising strands βand β. (C) Superposition of the backbone traces of the DWNN domain (in blue) and ubiquitin (1UBI, in yellow). The RMSD over structurally aligned regions between the two structures is 1.88 Å. (D) Structural alignment of the primary sequences of DWNN and ubiquitin, determined using the Dali server. The structurally equivalent regions comprise the following residues of DWNN: 2–9, 11–20, 22–40, 44–50, 53–57, 59–64, 66–75. Molecular fitting, calculation of RMSD's and generation of figures were performed using MOLMOL [26]

DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways-2

<p><b>Copyright information:</b></p><p>Taken from "DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways"</p><p>BMC Structural Biology 2006;6():1-1.</p><p>Published online 5 Jan 2006</p><p>PMCID:PMC1360078.</p><p></p>condary structural elements are derived from the final family of structures. Coupling constants were extracted from a N-HMQC-J spectrum. Half-circles correspond to J< 4 Hz; filled circles correspond to J> 8 Hz, except in the case of residues 56-7, which have Jvalues of 7.4 and 7.7 Hz respectively. These residues form part of strand β, which is the least stable of the five strands making up the main β-sheet. The presence of a β-sheet comprised of strands βand β, which does not occur in ubiquitin, is confirmed by large values of J, negative Cshifts and αN(i,i+1) NOE's. The diagram was generated using CYANA [45]

DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways-1

<p><b>Copyright information:</b></p><p>Taken from "DWNN, a novel ubiquitin-like domain, implicates RBBP6 in mRNA processing and ubiquitin-like pathways"</p><p>BMC Structural Biology 2006;6():1-1.</p><p>Published online 5 Jan 2006</p><p>PMCID:PMC1360078.</p><p></p>t single copy number, but not in prokaryotes. The alignment was performed using ClustalX [39] and the diagram produced using Pfaat [49]