Unusual binding properties of the SH3 domain of the yeast actin-binding protein Abp1: structural and functional analysis.
Abp1p is an actin-binding protein that plays a central role in the organization of Saccharomyces cerevisiae actin cytoskeleton. By a combination of two-hybrid and phage-display approaches, we have identified six new ligands of the Abp1-SH3 domain. None of these SH3-mediated novel interactions was detected in recent all genome high throughput ... protein interaction projects. Here we show that the SH3-mediated association of Abp1p with the Ser/Thr kinases Prk1p and Ark1p is essential for their localization to actin cortical patches. The Abp1-SH3 domain has a rather unusual binding specificity, because its target peptides contain the tetrapentapeptide +XXXPXXPX+PXXL with positive charges flanking the polyproline core on both sides. Here we present the structure of the Abp1-SH3 domain solved at 1.3-A resolution. The peptide-binding pockets in the SH3 domain are flanked by two acidic residues that are uncommon at those positions in the SH3 domain family. We have shown by site-directed mutagenesis that one of these negatively charged side chains may be the key determinant for the preference for non-classical ligands.
Mesh Terms:
Actins, Amino Acid Motifs, Amino Acid Sequence, Binding Sites, Cytoskeleton, DNA-Binding Proteins, Endocytosis, Enzyme-Linked Immunosorbent Assay, Gene Library, Ligands, Models, Biological, Models, Molecular, Molecular Sequence Data, Mutagenesis, Site-Directed, Peptide Library, Peptides, Plant Proteins, Plasmids, Protein Binding, Protein Conformation, Protein Structure, Tertiary, Receptor Protein-Tyrosine Kinases, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Schizosaccharomyces pombe Proteins, Structure-Activity Relationship, Transcription Factors, Two-Hybrid System Techniques, src Homology Domains
Actins, Amino Acid Motifs, Amino Acid Sequence, Binding Sites, Cytoskeleton, DNA-Binding Proteins, Endocytosis, Enzyme-Linked Immunosorbent Assay, Gene Library, Ligands, Models, Biological, Models, Molecular, Molecular Sequence Data, Mutagenesis, Site-Directed, Peptide Library, Peptides, Plant Proteins, Plasmids, Protein Binding, Protein Conformation, Protein Structure, Tertiary, Receptor Protein-Tyrosine Kinases, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Schizosaccharomyces pombe Proteins, Structure-Activity Relationship, Transcription Factors, Two-Hybrid System Techniques, src Homology Domains
J. Biol. Chem.
Date: Feb. 15, 2002
PubMed ID: 11668184
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