title: Unusual binding properties of the SH3 domain of the yeast actin-binding protein Abp1: structural and functional analysis.
abstract: 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.
authors.affiliation: Department of Biology, University of Rome Tor Vergata, Via della Ricerca Scientifica, 00133 Roma, Italy.
authors.forename: Barbara
authors.initials: B
authors.lastname: Fazi
authors.forename: M Jamie T V
authors.initials: MJ
authors.lastname: Cope
authors.forename: Alice
authors.initials: A
authors.lastname: Douangamath
authors.forename: Silvia
authors.initials: S
authors.lastname: Ferracuti
authors.forename: Katja
authors.initials: K
authors.lastname: Schirwitz
authors.forename: Adriana
authors.initials: A
authors.lastname: Zucconi
authors.forename: David G
authors.initials: DG
authors.lastname: Drubin
authors.forename: Matthias
authors.initials: M
authors.lastname: Wilmanns
authors.forename: Gianni
authors.initials: G
authors.lastname: Cesareni
authors.forename: Luisa
authors.initials: L
authors.lastname: Castagnoli
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites; D003599:Cytoskeleton; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D004705:Endocytosis; D004797:Enzyme-Linked Immunosorbent Assay; D015723:Gene Library; D008024:Ligands; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry; D010940:Plant Proteins; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary; D020794:Receptor Protein-Tyrosine Kinases / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D013329:Structure-Activity Relationship; D014157:Transcription Factors; D020798:Two-Hybrid System Techniques; D018909:src Homology Domains
chemical_list: C061886:ABF1 protein, S cerevisiae; D000199:Actins; D004268:DNA-Binding Proteins; D008024:Ligands; D019151:Peptide Library; D010455:Peptides; D010940:Plant Proteins; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D014157:Transcription Factors; C097691:cbp1 protein, S pombe; C088479:PRK1 protein, Petunia inflata; D020794:Receptor Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases