Structural basis of sugar-recognizing ubiquitin ligase.
SCF(Fbs1) is a ubiquitin ligase that functions in the endoplasmic reticulum (ER)-associated degradation pathway. Fbs1/Fbx2, a member of the F-box proteins, recognizes high-mannose oligosaccharides. Efficient binding to an N-glycan requires di-N-acetylchitobiose (chitobiose). Here we report the crystal structures of the sugar-binding domain (SBD) of Fbs1 alone and in complex with chitobiose. The SBD is composed of a ten-stranded antiparallel beta-sandwich. The structure of the SBD-chitobiose complex includes hydrogen bonds between Fbs1 and chitobiose and insertion of the methyl group of chitobiose into a small hydrophobic pocket of Fbs1. Moreover, NMR spectroscopy has demonstrated that the amino acid residues adjoining the chitobiose-binding site interact with the outer branches of the carbohydrate moiety. Considering that the innermost chitobiose moieties in N-glycans are usually involved in intramolecular interactions with the polypeptide moieties, we propose that Fbs1 interacts with the chitobiose in unfolded N-glycoprotein, pointing the protein moiety toward E2 for ubiquitination.
Mizushima
Tsunehiro
T
Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012, Japan.
Hirao
Takeshi
T
Yoshida
Yukiko
Y
Lee
Soo Jae
SJ
Chiba
Tomoki
T
Iwai
Kazuhiro
K
Yamaguchi
Yoshiki
Y
Kato
Koichi
K
Tsukihara
Tomitake
T
Tanaka
Keiji
K
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002236:Carbohydrate Conformation; D050260:Carbohydrate Metabolism; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000737:chemistry; D003001:Cloning, Molecular; D004721:Endoplasmic Reticulum / Q000201:enzymology; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011134:Polysaccharides / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000378:metabolism*
D002241:Carbohydrates; D011134:Polysaccharides; D011994:Recombinant Proteins; D044767:Ubiquitin-Protein Ligases