Use of the photoaffinity cross-linking agent N-hydroxysuccinimidyl-4-azidosalicylic acid to characterize salivary-glycoprotein-bacterial interactions.
The present study has utilized the iodinatable cross-linking agent N-hydroxysuccinimidyl-4-azidosalicylic acid (ASA) to examine the specific interaction between the proline-rich glycoprotein (PRG) of human parotid saliva and Streptococcus sanguis G9B. The binding of 125I-ASA-PRG to Streptococcus sanguis G9B displayed saturation kinetics, reversibility and was inhibited by unlabelled PRG. Inhibition studies with other glycoproteins and saccharides indicated that binding was mediated by a bacterial adhesin with specificity towards N-acetylneuraminic acid, galactose, and N-acetylgalactosamine. After cross-linking, the 125I-ASA-PRG-adhesin complex could be extracted with SDS and separated from uncoupled 125I-ASA-PRG by gel filtration on Sepharose CL-6B. Approx. 1% of the 125I-ASA-PRG was cross-linked to the bacterial surface. Examination of the 125I-ASA-PRG-adhesin complex by SDS/polyacrylamide-gel electrophoresis/fluorography on 5% -(w/v)-polyacrylamide gels revealed that PRG was bound to two bacterial components. These findings support our previous suggestion that human salivary glycoproteins can specifically interact with oral streptococci and that these interactions occur between the glycoprotein's carbohydrate units and lectin(s) on the bacterial cell surface.
Bergey
E J
EJ
Levine
M J
MJ
Reddy
M S
MS
Bradway
S D
SD
Al-Hashimi
I
I
D000328:Adult; D000345:Affinity Labels; D001386:Azides; D002241:Carbohydrates / Q000494:pharmacology; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000494:pharmacology; D006801:Humans; D008297:Male; D010455:Peptides / Q000378:metabolism*; D055232:Proline-Rich Protein Domains; D011485:Protein Binding / Q000187:drug effects; D012459:Salicylates; D012471:Salivary Proteins and Peptides / Q000378:metabolism*; D013298:Streptococcus sanguis / Q000378:metabolism*
D000345:Affinity Labels; D001386:Azides; D002241:Carbohydrates; D003432:Cross-Linking Reagents; D006023:Glycoproteins; D010455:Peptides; D012459:Salicylates; D012471:Salivary Proteins and Peptides; C046019:4-azidosalicylic acid N-hydroxysuccinimide ester