title: Structures of Rpn1 T1:Rad23 and hRpn13:hPLIC2 Reveal Distinct Binding Mechanisms between Substrate Receptors and Shuttle Factors of the Proteasome.
abstract: Three receptors (Rpn1/S2/PSMD2, Rpn10/S5a, Rpn13/Adrm1) in the proteasome bind substrates by interacting with conjugated ubiquitin chains and/or shuttle factors (Rad23/HR23, Dsk2/PLIC/ubiquilin, Ddi1) that carry ubiquitinated substrates to proteasomes. We solved the structure of two such receptors with their preferred shuttle factor, namely hRpn13(Pru):hPLIC2(UBL) and scRpn1 T1:scRad23(UBL). We find that ubiquitin folds in Rad23 and Dsk2 are fine-tuned by residue substitutions to achieve high affinity for Rpn1 and Rpn13, respectively. A single substitution in hPLIC2 yields enhanced interactions with the Rpn13 ubiquitin contact surface and sterically blocks hRpn13 binding to its preferred ubiquitin chain type, K48-linked chains. Rpn1 T1 binds two ubiquitins in tandem and we find that Rad23 binds exclusively to the higher-affinity Helix28/Helix30 site. Rad23 contacts at Helix28/Helix30 are optimized compared to ubiquitin by multiple conservative amino acid substitutions. Thus, shuttle factors deliver substrates to proteasomes through fine-tuned ubiquitin-like surfaces.
authors.affiliation: Protein Processing Section, Structural Biophysics Laboratory, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA.
authors.forename: Xiang
authors.initials: X
authors.lastname: Chen
authors.affiliation: Protein Processing Section, Structural Biophysics Laboratory, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA.
authors.forename: Leah
authors.initials: L
authors.lastname: Randles
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA.
authors.forename: Ke
authors.initials: K
authors.lastname: Shi
authors.affiliation: Biophysics Resource, Structural Biophysics Laboratory, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA.
authors.forename: Sergey G
authors.initials: SG
authors.lastname: Tarasov
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA.
authors.forename: Hideki
authors.initials: H
authors.lastname: Aihara
authors.affiliation: Protein Processing Section, Structural Biophysics Laboratory, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA. Electronic address: kylie.walters@nih.gov.
authors.forename: Kylie J
authors.initials: KJ
authors.lastname: Walters
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000071183:Autophagy-Related Proteins; D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013816:Thermodynamics; D014452:Ubiquitins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: C500781:ADRM1 protein, human; D048868:Adaptor Proteins, Signal Transducing; D000071183:Autophagy-Related Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; C490846:RAD23 protein, S cerevisiae; C499663:RPN1 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C405662:UBQLN2 protein, human; D014452:Ubiquitins; D046988:Proteasome Endopeptidase Complex
keywords: Dsk2; Rad23; Rpn1; Rpn13; proteasome; ubiquitin