title: Identification of the binding domain of Streptococcus oralis glyceraldehyde-3-phosphate dehydrogenase for Porphyromonas gingivalis major fimbriae.
abstract: Porphyromonas gingivalis forms communities with antecedent oral biofilm constituent streptococci. P. gingivalis major fimbriae bind to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) present on the streptococcal surface, and this interaction plays an important role in P. gingivalis colonization. This study identified the binding domain of Streptococcus oralis GAPDH for P. gingivalis fimbriae. S. oralis recombinant GAPDH (rGAPDH) was digested with lysyl endopeptidase. Cleaved fragments of rGAPDH were applied to a reverse-phase high-pressure liquid chromatograph equipped with a C18 column. Each peak was collected; the binding activity toward P. gingivalis recombinant fimbrillin (rFimA) was analyzed with a biomolecular interaction analysis system. The fragment displaying the strongest binding activity was further digested with various proteinases, after which the binding activity of each fragment was measured. The amino acid sequence of each fragment was determined by direct sequencing, mass spectrometric analysis, and amino acid analysis. Amino acid residues 166 to 183 of S. oralis GAPDH exhibited the strongest binding activity toward rFimA; confocal laser scanning microscopy revealed that the synthetic peptide corresponding to amino acid residues 166 to 183 of S. oralis GAPDH (pep166-183, DNFGVVEGLMTTIHAYTG) inhibits S. oralis-P. gingivalis biofilm formation in a dose-dependent manner. Moreover, pep166-183 inhibited interbacterial biofilm formation by several oral streptococci and P. gingivalis strains with different types of FimA. These results indicate that the binding domain of S. oralis GAPDH for P. gingivalis fimbriae exists within the region encompassing amino acid residues 166 to 183 of GAPDH and that pep166-183 may be a potent inhibitor of P. gingivalis colonization in the oral cavity.
authors.affiliation: Department of Preventive Dentistry, Osaka University Graduate School of Dentistry, 1-8 Yamadaoka, Suita, Osaka 565-0871, Japan. nagatah@dent.osaka-u.ac.jp
authors.forename: Hideki
authors.initials: H
authors.lastname: Nagata
authors.forename: Mio
authors.initials: M
authors.lastname: Iwasaki
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Maeda
authors.forename: Masae
authors.initials: M
authors.lastname: Kuboniwa
authors.forename: Ei
authors.initials: E
authors.lastname: Hashino
authors.forename: Masahiro
authors.initials: M
authors.lastname: Toe
authors.forename: Naoto
authors.initials: N
authors.lastname: Minamino
authors.forename: Hiromiki
authors.initials: H
authors.lastname: Kuwahara
authors.forename: Satoshi
authors.initials: S
authors.lastname: Shizukuishi
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000378:metabolism*; D018441:Biofilms; D002851:Chromatography, High Pressure Liquid; D036781:Fimbriae Proteins / Q000737:chemistry / Q000378:metabolism*; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating) / Q000737:chemistry / Q000378:metabolism*; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D016966:Porphyromonas gingivalis / Q000502:physiology*; D011485:Protein Binding; D019533:Streptococcus oralis / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; C063055:fimbrillin; D036781:Fimbriae Proteins; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating)