The secretions of the salivary parotid and submandibular-sublingual (SMSL) glands constitute the main part of whole human saliva (WS) in which proline-rich proteins (PRPs) and mucins represent dominant groups. Although proteome analysis had been performed on WS, no identification of PRPs or mucins by 2-DE and MS was achieved in WS and no comprehensive analysis of both glandular secretions is available so far. The aim of this study was to compare the protein map of WS to parotid and SMSL secretions for the display of PRPs and mucins. WS and glandular secretions were subjected to 2-DE and spots were analyzed by MALDI-MS. New components identified in WS were cyclophilin-B and prolyl-4-hydroxylase. Also acidic and basic PRPs as well as the proline-rich glycoprotein (PRG) could now be mapped in WS. Acidic PRPs were found equally in parotid and SMSL secretions, whereas basic PRPs and PRG were found primarily in parotid secretion. Salivary mucin MUC7 was identified in SMSL secretion. Thus, the more abundant proteins of WS can be explained mainly by mixed contributions of parotid and SMSL secretions with only few components remaining that may be derived from local sources in the oral cavity.

The microaerophilic bacterium Helicobacter pylori is well established for its role in development of different gastric diseases. Bacterial adhesins and corresponding binding sites on the epithelial surface allow H. pylori to colonize the gastric tissue. In this investigation, the adhesion of H. pylori to dot blot arrays of natural glycoproteins and neoglycoproteins was studied. Adhesion was detected by overlay with fluorescence-labeled bacteria on immobilized (neo)glycoproteins. The results confirmed the interaction between the adhesin BabA and the H-1-, Lewis b-, and related fucose-containing antigens. In addition, H. pylori bound to terminal alpha2-3-linked sialic acids as previously described. The use of a sabA mutant and sialidase treatment of glycoconjugate arrays showed that the adherence of H. pylori to laminin is mediated by the sialic acid-binding adhesin, SabA. The adhesion to salivary mucin MUC5B is mainly associated with the BabA adhesin and to a lesser extent with the SabA adhesin. This agrees with reports, that MUC5B carries both fucosylated blood group antigens and alpha2-3-linked sialic acids. The adhesion of H. pylori to fibronectin and lactoferrin persisted in the babA/sabA double mutant. Because binding to these molecules was abolished by denaturation rather than by deglycosylation, it was suggested to depend on the recognition of unknown receptor moieties by an additional unknown bacterial surface component. The results demonstrate that the bacterial overlay method on glycoconjugate arrays is a useful tool for exploration and the characterization of unknown adhesin specificities of H. pylori and other bacteria.

Because gastric infection by Helicobacter pylori takes place via the oral route, possible interactions of this bacterium with human salivary proteins could occur. By using modified 1- and 2-D bacterial overlay, binding of H. pylori adhesins BabA and SabA to the whole range of salivary proteins was explored. Bound salivary receptor molecules were identified by MALDI-MS and by comparison to previously established proteome maps of whole and glandular salivas. By use of adhesin-deficient mutants, binding of H. pylori to MUC7 and gp-340 could be linked to the SabA and BabA adhesins, respectively, whereas binding to MUC5B was associated with both adhesins. Binding of H. pylori to the proline-rich glycoprotein was newly detected and assigned to BabA adhesin whereas the SabA adhesin was found to mediate binding to newly detected receptor molecules, including carbonic anhydrase VI, secretory component, heavy chain of secretory IgA1, parotid secretory protein and zinc-alpha(2)-glycoprotein. Some of these salivary glycoproteins are known to act as scavenger molecules or are involved in innate immunity whereas others might come to modify the pathogenetic properties of this organism. In general, this 2-D bacterial overlay technique represents a useful supplement in adhesion studies of bacteria with complex protein mixtures.

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