Dcn1 functions as a scaffold-type E3 ligase for cullin neddylation.
Cullin-based E3 ubiquitin ligases are activated through modification of the cullin subunit with the ubiquitin-like protein Nedd8. Dcn1 regulates cullin neddylation and thus ubiquitin ligase activity. Here we describe the 1.9 A X-ray crystal structure of yeast Dcn1 encompassing an N-terminal ubiquitin-binding (UBA) domain and a C-terminal domain of unique architecture, which we termed PONY domain. A conserved surface on Dcn1 is required for direct binding to cullins and for neddylation. The reciprocal binding site for Dcn1 on Cdc53 is located approximately 18 A from the site of neddylation. Dcn1 does not require cysteine residues for catalytic function, and directly interacts with the Nedd8 E2 Ubc12 on a surface that overlaps with the E1-binding site. We show that Dcn1 is necessary and sufficient for cullin neddylation in a purified recombinant system. Taken together, these data demonstrate that Dcn1 is a scaffold-like E3 ligase for cullin neddylation.
Kurz
Thimo
T
Institute of Biochemistry, ETH Zürich, Schafmattstrasse 18, CH-8093 Zürich, Switzerland.
Chou
Yang-Chieh
YC
Willems
Andrew R
AR
Meyer-Schaller
Nathalie
N
Hecht
Marie-Lyn
ML
Tyers
Mike
M
Peter
Matthias
M
Sicheri
Frank
F
D000595:Amino Acid Sequence; D001665:Binding Sites; D002384:Catalysis; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D054875:Ubiquitination; D014452:Ubiquitins
C502135:Dcn1 protein, S cerevisiae; C111706:RUB1 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D014452:Ubiquitins; D044767:Ubiquitin-Protein Ligases