Structural basis for recognition of diubiquitins by NEMO.
NEMO is the regulatory subunit of the IkappaB kinase (IKK) in NF-kappaB activation, and its CC2-LZ region interacts with Lys63 (K63)-linked polyubiquitin to recruit IKK to receptor signaling complexes. In vitro, CC2-LZ also interacts with tandem diubiquitin. Here we report the crystal structure of CC2-LZ with two dimeric coiled coils representing CC2 and LZ, respectively. Surprisingly, mutagenesis and nuclear magnetic resonance experiments reveal that the binding sites for diubiquitins at LZ are composites of both chains and that each ubiquitin in diubiquitins interacts with symmetrical NEMO asymmetrically. For tandem diubiquitin, the first ubiquitin uses the conserved hydrophobic patch and the C-terminal tail, while the second ubiquitin uses an adjacent surface patch. For K63-linked diubiquitin, the proximal ubiquitin uses its conserved hydrophobic patch, while the distal ubiquitin mostly employs the C-terminal arm including the K63 linkage residue. These studies uncover the energetics and geometry for mutual recognition of NEMO and diubiquitins.
Lo
Yu-Chih
YC
Department of Biochemistry, Weill Cornell Medical College, New York, NY 10021, USA.
Lin
Su-Chang
SC
Rospigliosi
Carla C
CC
Conze
Dietrich B
DB
Wu
Chuan-Jin
CJ
Ashwell
Jonathan D
JD
Eliezer
David
D
Wu
Hao
H
D000595:Amino Acid Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D020022:Genetic Predisposition to Disease; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D051550:I-kappa B Kinase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D013329:Structure-Activity Relationship; D014452:Ubiquitins / Q000737:chemistry* / Q000378:metabolism
C491731:IKBKG protein, human; D016328:NF-kappa B; D011994:Recombinant Proteins; C109184:UBD protein, human; D014452:Ubiquitins; D051550:I-kappa B Kinase