Characterization of Two Polyubiquitin Binding Sites in the 26 S Protease Subunit 5a

• Published in: The Journal of biological chemistry Vol. 273; no. 10; pp. 5461 - 5467
• Main Authors: Young, Patrick, Deveraux, Quinn, Beal, Richard E., Pickart, Cecile M., Rechsteiner, Martin
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 06.03.1998; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Sequence; Binding Sites - physiology; Biopolymers - metabolism; Conserved Sequence - genetics; Humans; Molecular Sequence Data; Muramidase - metabolism; Mutagenesis, Site-Directed - genetics; Peptide Fragments - chemistry; Peptide Fragments - pharmacology; Peptide Hydrolases - chemistry; Peptide Hydrolases - physiology; Polyubiquitin; Proteasome Endopeptidase Complex; Protein Binding - physiology; Recombinant Proteins - chemistry; Recombinant Proteins - pharmacology; Sequence Deletion - genetics; Sequence Homology, Amino Acid; Ubiquitins - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.273.10.5461

Ubiquitylated proteins are degraded by the 26 S protease, an enzyme complex that contains 30 or more unique subunits. One of these proteins, subunit 5a (S5a), has been shown to bind ubiquitin-lysozyme conjugates and free polyubiquitin chains. Using deletional analysis, we have identified in the carboxyl-terminal half of human S5a, two independent polyubiquitin binding sites whose sequences are highly conserved among higher eukaryotic S5a homologs. The sites are approximately 30-amino acids long and are separated by 50 intervening residues. When expressed as small fragments or when present in full-length S5a molecules, the sites differ at least 10-fold in their apparent affinity for polyubiquitin chains. Each binding site contains 5 hydrophobic residues that form an alternating pattern of large and small side chains, e.g. Leu-Ala-Leu-Ala-Leu, and this pattern is essential for binding ubiquitin chains. Based on the importance of the alternating hydrophobic residues in the binding sites and previous studies showing that a hydrophobic patch on the surface of ubiquitin is essential for proteolytic targeting, we propose a model for molecular recognition of polyubiquitin chains by S5a.