Characterization of Binding of Streptococcus oralis Glyceraldehyde-3-Phosphate Dehydrogenase to Porphyromonas gingivalis Major Fimbriae

Abstract

Binding of Streptococcus oralis glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to Porphyromonas gingivalis fimbriae was characterized via a biomolecular interaction analysis system. The interaction was specific, and the association constant value was 4.34 ؋ 10 7 M ؊1 , suggesting that S. oralis GAPDH functions as a dominant receptor for P. gingivalis and contributes to P. gingivalis colonization.Interaction of Porphyromonas gingivalis, which is a predominant periodontal pathogen, with early plaque-forming bacteria plays an important role with respect to colonization in periodontal pockets (4,12,13). P. gingivalis interacts with a variety of other oral gram-positive bacteria, including Actinomyces naeslundii (16), Streptococcus gordonii (5), Streptococcus oralis (7), and Streptococcus sanguinis (14); these intergeneric coaggregations may lead to the initial colonization of P. gingivalis in the oral cavity.A series of studies demonstrated that P. gingivalis 381 strongly coaggregated with S. oralis ATCC 9811; moreover, its fimbriae were primarily responsible for the interaction, in which several domains of the carboxy terminus of fimbrillin participated (1). A recent paper regarding coadhesin of S. oralis by Maeda et al. (6) revealed that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of S. oralis ATCC 9811 mediated binding to P. gingivalis 381 major fimbriae. GAPDH, which is a tetrameric enzyme of the glycolytic pathway, is responsible for the phosphorylation of glyceraldehyde-3-phosphate, which leads to generation of 1,3-bisphosphoglycerate (15). In addition to its enzymatic activity, GAPDH reportedly possesses multiple binding activities. Pancholi and Fischetti (11) found that the GAPDH of group A streptococci binds to fibronectin, lysozyme, and the cytoskeletal proteins myosin and actin; they noted that it may function in the colonization of those bacteria. In the present study, kinetic interaction of GAPDH of S. oralis ATCC 9811 with P. gingivalis 381 fimbriae was analyzed based on surface plasmon resonance spectroscopy with a biomolecular interaction analysis system (BIAcore).S. oralis ATCC 9811 was maintained as frozen stocks and was cultured in brain heart infusion broth (BBL Microbiology Systems, Cockeysville, Md.) for 15 h at 35°C in air. Bacterial cells were harvested by centrifugation (High Speed Refrigerated Centrifuge SRX-201; Tomy Seiko Co. Ltd., Tokyo, Japan) at 5,000 ϫ g for 30 min at 4°C; they were subsequently washed three times with 20 mM phosphate buffer supplemented with 0.15 M NaCl (phosphate-buffered saline [PBS] [pH 6.0]) and were suspended in the same buffer.S. oralis ATCC 9811 GAPDH was purified by mild ultrasonication, 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate treatment, ammonium sulfate precipitation, and chromatography with an affinity column coupled with P. gingivalis recombinant fimbrillin (rFimA) as reported previously (6). P. gingivalis 381 rFimA and S. oralis ATCC 9811 recombinant GAPDH (rGAPDH) were prepared in accordance with the methods previously report...

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