The native 67-kilodalton minor fimbria of Porphyromonas gingivalis is a novel glycoprotein with DC-SIGN-targeting motifs.
We recently reported that the oral mucosal pathogen Porphyromonas gingivalis, through its 67-kDa Mfa1 (minor) fimbria, targets the C-type lectin receptor DC-SIGN for invasion and persistence within human monocyte-derived dendritic cells (DCs). The DCs respond by inducing an immunosuppressive and Th2-biased CD4(+) T-cell response. We have now purified the native minor fimbria by ion-exchange chromatography and sequenced the fimbria by tandem mass spectrometry (MS/MS), confirming its identity and revealing two putative N-glycosylation motifs as well as numerous putative O-glycosylation sites. We further show that the minor fimbria is glycosylated by ProQ staining and that glycosylation is partially removed by treatment with beta(1-4)-galactosidase, but not by classic N- and O-linked deglycosidases. Further monosaccharide analysis by gas chromatography-mass spectrometry (GC-MS) confirmed that the minor fimbria contains the DC-SIGN-targeting carbohydrates fucose (1.35 nmol/mg), mannose (2.68 nmol/mg), N-acetylglucosamine (2.27 nmol/mg), and N-acetylgalactosamine (0.652 nmol/mg). Analysis by transmission electron microscopy revealed that the minor fimbria forms fibers approximately 200 nm in length that could be involved in targeting or cross-linking DC-SIGN. These findings shed further light on molecular mechanisms of invasion and immunosuppression by this unique mucosal pathogen.
Zeituni
Amir E
AE
Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, NY 11794-8703, USA.
McCaig
William
W
Scisci
Elizabeth
E
Thanassi
David G
DG
Cutler
Christopher W
CW
D000116:Acetylgalactosamine / Q000032:analysis; D000117:Acetylglucosamine / Q000032:analysis; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D036781:Fimbriae Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010861:Fimbriae, Bacterial / Q000648:ultrastructure; D005643:Fucose / Q000032:analysis; D008401:Gas Chromatography-Mass Spectrometry; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D037181:Lectins, C-Type / Q000378:metabolism*; D008358:Mannose / Q000032:analysis; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016966:Porphyromonas gingivalis / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism*; D020539:Sequence Analysis, Protein; D053719:Tandem Mass Spectrometry; D037521:Virulence Factors / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin
D000116:Acetylgalactosamine; D000117:Acetylglucosamine; D001426:Bacterial Proteins; D015815:Cell Adhesion Molecules; D036781:Fimbriae Proteins; D005643:Fucose; D006023:Glycoproteins; D037181:Lectins, C-Type; D008358:Mannose; D011956:Receptors, Cell Surface; D037521:Virulence Factors; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin; C545438:Mfa1 protein, Porphyromonas gingivalis