Prediction of glycolipid-binding domains from the amino acid sequence of lipid raft-associated proteins: application to HpaA, a protein involved in the adhesion of Helicobacter pylori to gastrointestinal cells.
Protein-glycolipid interactions mediate the attachment of various pathogens to the host cell surface as well as the association of numerous cellular proteins with lipid rafts. Thus, it is of primary importance to identify the protein domains involved in glycolipid recognition. Using structure similarity searches, we could identify a common glycolipid-binding domain in the three-dimensional structure of several proteins known to interact with lipid rafts. Yet the three-dimensional structure of most raft-targeted proteins is still unknown. In the present study, we have identified a glycolipid-binding domain in the amino acid sequence of a bacterial adhesin (Helicobacter pylori adhesin A, HpaA). The prediction was based on the major properties of the glycolipid-binding domains previously characterized by structural searches. A short (15-mer) synthetic peptide corresponding to this putative glycolipid-binding domain was synthesized, and we studied its interaction with glycolipid monolayers at the air-water interface. The synthetic HpaA peptide recognized LacCer but not Gb3. This glycolipid specificity was in line with that of the whole bacterium. Molecular modeling studies gave some insights into this high selectivity of interaction. It also suggested that Phe147 in HpaA played a key role in LacCer recognition, through sugar-aromatic CH-pi stacking interactions with the hydrophobic side of the galactose ring of LacCer. Correspondingly, the replacement of Phe147 with Ala strongly affected LacCer recognition, whereas substitution with Trp did not. Our method could be used to identify glycolipid-binding domains in microbial and cellular proteins interacting with lipid shells, rafts, and other specialized membrane microdomains.
Fantini
Jacques
J
Laboratoire de Biochimie et Physicochimie des Membranes Biologiques, Faculté des Sciences de St-Jérôme, Université Paul Cézanne, Marseille, France. jacques.fantini@univ-cezanne.fr
Garmy
Nicolas
N
Yahi
Nouara
N
D018829:Adhesins, Bacterial / Q000737:chemistry* / Q000378:metabolism*; D000465:Algorithms; D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000378:metabolism; D001422:Bacterial Adhesion; D001665:Binding Sites; D002240:Carbohydrate Sequence; D041981:Gastrointestinal Tract / Q000382:microbiology; D006017:Glycolipids / Q000378:metabolism*; D016480:Helicobacter pylori / Q000472:pathogenicity; D006801:Humans; D007790:Lactosylceramides / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism
D018829:Adhesins, Bacterial; D015703:Antigens, CD; D006017:Glycolipids; D007790:Lactosylceramides; D011506:Proteins; D011956:Receptors, Cell Surface; C068556:Shiga-like toxin receptor; C095977:adhesin A, Helicobacter; C009744:CDw17 antigen