title: Structural analyses of the Slm1-PH domain demonstrate ligand binding in the non-canonical site.
abstract: BACKGROUND
  Pleckstrin homology (PH) domains are common membrane-targeting modules and their best characterized ligands are a set of important signaling lipids that include phosphatidylinositol phosphates (PtdInsPs). PH domains recognize PtdInsPs through two distinct mechanisms that use different binding pockets on opposite sides of the β-strands 1 and 2: i) a canonical binding site delimited by the β1-β2 and β3-β4loops and ii) a non-canonical binding site bordered by the β1-β2 and β5-β6loops. The PH domain-containing protein Slm1 from budding yeast Saccharomyces cerevisiae is required for actin cytoskeleton polarization and cell growth. We recently reported that this PH domain binds PtdInsPs and phosphorylated sphingolipids in a cooperative manner.
  
  
  RESULTS
  To study the structural basis for the Slm1-PH domain (Slm1-PH) specificity, we co-crystallized this domain with different soluble compounds that have structures analogous to anionic lipid head groups of reported Slm1 ligands: inositol 4-phosphate, which mimics phosphatidylinositol-4-phosphate (PtdIns(4)P), and phosphoserine as a surrogate for dihydrosphingosine 1-phosphate (DHS1-P). We found electron densities for the ligands within the so-called non-canonical binding site. An additional positively charged surface that contacts a phosphate group was identified next to the canonical binding site.
  
  
  CONCLUSIONS
  Our results suggest that Slm1-PH utilizes a non-canonical binding site to bind PtdInsPs, similar to that described for the PH domains of β-spectrin, Tiam1 and ArhGAP9. Additionally, Slm1-PH may have retained an active canonical site. We propose that the presence of both a canonical and a non-canonical binding pocket in Slm1-PH may account for the cooperative binding to PtdInsPs and DHS-1P.
authors.affiliation: European Molecular Biology Laboratory Heidelberg, Structural and Computational Biology Unit, Heidelberg, Germany. anand@embl.de
authors.forename: Kanchan
authors.initials: K
authors.lastname: Anand
authors.forename: Kenji
authors.initials: K
authors.lastname: Maeda
authors.forename: Anne-Claude
authors.initials: AC
authors.lastname: Gavin
mesh_terms: D000595:Amino Acid Sequence; D001059:Apoproteins / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D001798:Blood Proteins / Q000737:chemistry*; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D018360:Crystallography, X-Ray; D003598:Cytoskeletal Proteins; D008024:Ligands; D050356:Lipid Metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D001059:Apoproteins; D001798:Blood Proteins; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D008024:Ligands; D010750:Phosphoproteins; D029701:Saccharomyces cerevisiae Proteins; C502930:Slm1 protein, S cerevisiae; C039077:platelet protein P47