A binding motif for Siah ubiquitin ligase.
The Drosophila SINA (seven in absentia) protein and its mammalian orthologs (Siah, seven in absentia homolog) are RING domain proteins that function in E3 ubiquitin ligase complexes and facilitate ubiquitination and degradation of a wide range of cellular proteins, including beta-catenin. Despite these diverse targets, the means by which SINASiah recognize substrates or binding proteins has remained unknown. Here we identify a peptide motif (RPVAxVxPxxR) that mediates the interaction of Siah protein with a range of protein partners. Sequence alignment and mutagenesis scanning revealed residues that are important to this interaction. This consensus sequence correctly predicted a high-affinity interaction with a peptide from the cytoskeletal protein plectin-1 (residues 95-117). The unusually high-affinity binding obtained with a 23-residue peptide (K(Dapp) = 29 nM with SINA) suggests that it may serve as a useful dominant negative reagent for SINASiah proteins.
House
Colin M
CM
Trescowthick Research Laboratories, Peter MacCallum Cancer Institute, Melbourne 8006, Victoria, Australia.
Frew
Ian J
IJ
Huang
Huei-Luen
HL
Wiche
Gerhard
G
Traficante
Nadia
N
Nice
Edouard
E
Catimel
Bruno
B
Bowtell
David D L
DD
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000378:metabolism; D017124:Conserved Sequence; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D066298:In Vitro Techniques; D008025:Ligases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins
C491778:CACYBP protein, human; C111922:Cacybp protein, mouse; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D029721:Drosophila Proteins; D006360:Heat-Shock Proteins; D009687:Nuclear Proteins; D011994:Recombinant Proteins; C092070:phyl protein, Drosophila; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins; D008025:Ligases