PMID- 26130806 OWN - NLM STAT- MEDLINE DCOM- 20151030 LR - 20181113 IS - 1091-6490 (Electronic) IS - 0027-8424 (Print) IS - 0027-8424 (Linking) VI - 112 IP - 28 DP - 2015 Jul 14 TI - Structural characterization of the interaction of Ubp6 with the 26S proteasome. PG - 8626-31 LID - 10.1073/pnas.1510449112 [doi] AB - In eukaryotic cells, the 26S proteasome is responsible for the regulated degradation of intracellular proteins. Several cofactors interact transiently with this large macromolecular machine and modulate its function. The deubiquitylating enzyme ubiquitin C-terminal hydrolase 6 [Ubp6; ubiquitin-specific protease (USP) 14 in mammals] is the most abundant proteasome-interacting protein and has multiple roles in regulating proteasome function. Here, we investigate the structural basis of the interaction between Ubp6 and the 26S proteasome in the presence and absence of the inhibitor ubiquitin aldehyde. To this end we have used single-particle electron cryomicroscopy in combination with cross-linking and mass spectrometry. Ubp6 binds to the regulatory particle non-ATPase (Rpn) 1 via its N-terminal ubiquitin-like domain, whereas its catalytic USP domain is positioned variably. Addition of ubiquitin aldehyde stabilizes the binding of the USP domain in a position where it bridges the proteasome subunits Rpn1 and the regulatory particle triple-A ATPase (Rpt) 1. The USP domain binds to Rpt1 in the immediate vicinity of the Ubp6 active site, which may effect its activation. The catalytic triad is positioned in proximity to the mouth of the ATPase module and to the deubiquitylating enzyme Rpn11, strongly implying their functional linkage. On the proteasome side, binding of Ubp6 favors conformational switching of the 26S proteasome into an intermediate-energy conformational state, in particular upon the addition of ubiquitin aldehyde. This modulation of the conformational space of the 26S proteasome by Ubp6 explains the effects of Ubp6 on the kinetics of proteasomal degradation. FAU - Aufderheide, Antje AU - Aufderheide A AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; FAU - Beck, Florian AU - Beck F AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; FAU - Stengel, Florian AU - Stengel F AD - Department of Biology, University of Konstanz, 78457 Konstanz, Germany; FAU - Hartwig, Michaela AU - Hartwig M AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; FAU - Schweitzer, Andreas AU - Schweitzer A AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; FAU - Pfeifer, Gunter AU - Pfeifer G AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; FAU - Goldberg, Alfred L AU - Goldberg AL AD - Department of Cell Biology, Harvard Medical School, Boston, MA 02115. FAU - Sakata, Eri AU - Sakata E AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; FAU - Baumeister, Wolfgang AU - Baumeister W AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; baumeist@biochem.mpg.de foerster@biochem.mpg.de. FAU - Forster, Friedrich AU - Forster F AD - Department of Molecular Structural Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; baumeist@biochem.mpg.de foerster@biochem.mpg.de. LA - eng SI - PDB/5A5B PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20150630 PL - United States TA - Proc Natl Acad Sci U S A JT - Proceedings of the National Academy of Sciences of the United States of America JID - 7505876 RN - 0 (Saccharomyces cerevisiae Proteins) RN - EC 3.4.- (Endopeptidases) RN - EC 3.4.25.1 (Proteasome Endopeptidase Complex) RN - EC 3.4.99.- (ATP dependent 26S protease) RN - EC 3.4.99.- (UBP6 protein, S cerevisiae) SB - IM CIN - Nat Struct Mol Biol. 2015 Sep;22(9):652-3. PMID: 26333712 MH - Catalytic Domain MH - Cryoelectron Microscopy MH - Endopeptidases/chemistry/*metabolism MH - Proteasome Endopeptidase Complex/chemistry/*metabolism MH - Protein Binding MH - Protein Conformation MH - Saccharomyces cerevisiae/metabolism MH - Saccharomyces cerevisiae Proteins/chemistry/*metabolism PMC - PMC4507206 OTO - NOTNLM OT - conformational switching OT - proteolysis OT - proteostasis OT - quality control COIS- The authors declare no conflict of interest. EDAT- 2015/07/02 06:00 MHDA- 2015/10/31 06:00 PMCR- 2015/06/30 CRDT- 2015/07/02 06:00 PHST- 2015/07/02 06:00 [entrez] PHST- 2015/07/02 06:00 [pubmed] PHST- 2015/10/31 06:00 [medline] PHST- 2015/06/30 00:00 [pmc-release] AID - 1510449112 [pii] AID - 201510449 [pii] AID - 10.1073/pnas.1510449112 [doi] PST - ppublish SO - Proc Natl Acad Sci U S A. 2015 Jul 14;112(28):8626-31. doi: 10.1073/pnas.1510449112. Epub 2015 Jun 30.