title: Human DNA-Damage-Inducible 2 Protein Is Structurally and Functionally Distinct from Its Yeast Ortholog.
abstract: Although Ddi1-like proteins are conserved among eukaryotes, their biological functions remain poorly characterized. Yeast Ddi1 has been implicated in cell cycle regulation, DNA-damage response, and exocytosis. By virtue of its ubiquitin-like (UBL) and ubiquitin-associated (UBA) domains, it has been proposed to serve as a proteasomal shuttle factor. All Ddi1-like family members also contain a highly conserved retroviral protease-like (RVP) domain with unknown substrate specificity. While the structure and biological function of yeast Ddi1 have been investigated, no such analysis is available for the human homologs. To address this, we solved the 3D structures of the human Ddi2 UBL and RVP domains and identified a new helical domain that extends on either side of the RVP dimer. While Ddi1-like proteins from all vertebrates lack a UBA domain, we identify a novel ubiquitin-interacting motif (UIM) located at the C-terminus of the protein. The UIM showed a weak yet specific affinity towards ubiquitin, as did the Ddi2 UBL domain. However, the full-length Ddi2 protein is unable to bind to di-ubiquitin chains. While proteomic analysis revealed no activity, implying that the protease requires other factors for activation, our structural characterization of all domains of human Ddi2 sets the stage for further characterization.
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.|First Faculty of Medicine, Charles University in Prague, Katerinska 32, 121 08, Prague 2, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University, Hlavova 8, 128 00 Prague 2, Czech Republic.
authors.forename: Monika
authors.initials: M
authors.lastname: Sivá
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.|Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University, Hlavova 8, 128 00 Prague 2, Czech Republic.
authors.forename: Michal
authors.initials: M
authors.lastname: Svoboda
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Václav
authors.initials: V
authors.lastname: Veverka
authors.affiliation: Groupe de Recherche Axé sur la Structure des Protéines, Department of Pharmacology &Therapeutics, McGill University, Montreal, QC, H3G 1Y6, Canada.
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Trempe
authors.affiliation: Institute for Genetics, University of Cologne, Zülpicher Str. 47a, 50647 Cologne, Germany.
authors.forename: Kay
authors.initials: K
authors.lastname: Hofmann
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Milan
authors.initials: M
authors.lastname: Kožíšek
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Rozálie
authors.initials: R
authors.lastname: Hexnerová
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.|First Faculty of Medicine, Charles University in Prague, Katerinska 32, 121 08, Prague 2, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University, Hlavova 8, 128 00 Prague 2, Czech Republic.
authors.forename: František
authors.initials: F
authors.lastname: Sedlák
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University, Hlavova 8, 128 00 Prague 2, Czech Republic.
authors.forename: Jan
authors.initials: J
authors.lastname: Belza
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Jiří
authors.initials: J
authors.lastname: Brynda
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Pavel
authors.initials: P
authors.lastname: Šácha
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Martin
authors.initials: M
authors.lastname: Hubálek
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Jana
authors.initials: J
authors.lastname: Starková
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.
authors.forename: Iva
authors.initials: I
authors.lastname: Flaisigová
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University, Hlavova 8, 128 00 Prague 2, Czech Republic.
authors.forename: Jan
authors.initials: J
authors.lastname: Konvalinka
authors.affiliation: Gilead Sciences and IOCB Research Center, Institute of Organic Chemistry and Biochemistry of the Academy of Sciences of the Czech Republic, Flemingovo n. 2, 166 10 Prague 6, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University, Hlavova 8, 128 00 Prague 2, Czech Republic.
authors.forename: Klára Grantz
authors.initials: KG
authors.lastname: Šašková
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D057055:Aspartic Acid Proteases / Q000737:chemistry* / Q000378:metabolism*; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D019143:Evolution, Molecular; D057809:HEK293 Cells; D006801:Humans; D008958:Models, Molecular; D025821:Polyubiquitin / Q000378:metabolism; D011485:Protein Binding; D000072417:Protein Domains; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D059748:Proteolysis; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*; D053838:Scattering, Small Angle; D020539:Sequence Analysis, Protein; D012996:Solutions; D040681:Structural Homology, Protein
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D012996:Solutions; D025821:Polyubiquitin; D057055:Aspartic Acid Proteases; C000624278:DDI2 protein, human