Ubiquitin recognition by the DNA repair protein hHR23a.
Ubiquitin is a prominent regulatory protein in numerous biological processes, including targeted protein degradation, endocytic sorting, transcriptional control, intranuclear localization, and retroviral virion budding. Ubiquitin-associated (UBA) domains, ubiquitin interacting motifs (UIM), and coupling of ubiquitin conjugation to ER degradation (CUE) motifs have been identified as ubiquitin receptors. The DNA repair protein hHR23a has two UBA domains that can each bind ubiquitin in addition to an N-terminal UBL domain that binds S5a and S2, two components of the 26S proteasome. Here we reveal hHR23a recognizes ubiquitin through a predominately hydrophobic surface formed by residues within alpha1 and alpha3 of each of its UBA domains. These two UBA surfaces bind a region on ubiquitin that includes K48. These findings have implications for published studies revealing that hHR23a inhibits K48-linked polyubiquitin chain formation. In addition, by using (15)N NMR relaxation experiments, we find that binding ubiquitin requires a structural change in hHR23a. HHR23 proteins are hypothesized to link ubiquitin to S5a, and we provide direct evidence that hHR23 could form a ternary complex with ubiquitin and S5a.
Wang
Qinghua
Q
Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Goh
Amanda M
AM
Howley
Peter M
PM
Walters
Kylie J
KJ
D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D004260:DNA Repair; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008239:Lysine / Q000378:metabolism; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D055672:Static Electricity; D025801:Ubiquitin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C498506:PSMD4 protein, human; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D025801:Ubiquitin; C086985:RAD23A protein, human; D046988:Proteasome Endopeptidase Complex; D045643:DNA Repair Enzymes; D008239:Lysine