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Host ligands and oral bacterial adhesion: studies on phosphorylated polypeptides and gp-340 in saliva and milk
Umeå University, Faculty of Medicine, Odontology, Cariology.
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Infectious diseases e.g. gastric ulcer, caries and perodontitis, are caused by bacteria in a biofilm. Adhesion of bacteria to host ligands e.g. proteins, polypeptides and glycoproteins, is a key event in biofilm formation and colonization of surfaces such as mucosa and tooth tissues. Thus, host ligands could contribute to the susceptibility to infectious diseases. The general aim of this doctoral thesis was to study the effect of phosphorylated polypeptides and gp-340 in saliva and milk on oral bacterial adhesion and aggregation.

Statherin is a non-glycosylated, phosphorylated polypeptide in saliva. The polypeptide inhibits precipitation and crystal growth of calcium phosphate and mediates adhesion of microorganisms. By using a hybrid peptide construct, the domain for adhesion of Actinomyces isolated from human infections and from rodents was found to reside in the C-terminal end, and the adhesion was inhibitable. With alanine substitution the peptide recognition epitope in the C-terminal end was delineated to Q and TF, where QAATF was an optimal inhibitory peptide. In contrast, human commensal Actinomyces bound to the middle region in a non-inhibitable fashion. Gp-340 is another protein in saliva, and it is a large, multifunctional glycoprotein. Four novel size variants (I-IV) of salivary gp-340 were distinguished within individuals, and their glycoforms were characterized. All four size variants were identical in the N-terminal amino acid sequence and shared core carbohydrates. Low-glyco lung gp-340, high-glyco saliva gp-340, and size variants I-III aggregated bacteria differently. Human milk, which shares many traits with saliva, could inhibit adhesion of Streptococcus mutans to saliva-coated hydroxyapatite (s-HA), a model for teeth, in an individually varying fashion. Human milk caseins, lactoferrin, secretory IgA, and IgG inhibited the binding avidly. By using synthetic peptides the inhibitory epitope in b-casein was mapped to a C-terminal stretch of 30 amino acids. Inhibition by human milk, secretory IgA and the b-casein-derived inhibitory peptide was universal among a panel of mutans streptococci.

The main conclusions are: (i) statherin mediates differential binding of commensal versus infectious Actinomyces strains with small conformation-dependent binding epitopes, (ii) salivary gp-340 has individual polymorphisms that at least affect binding of bacteria, (iii) human milk inhibits S. mutans adhesion to s-HA in an individually varying fashion, and the C-terminal end of human milk β-casein is one inhibitory component. Together these results suggest that the studied host ligands can influence the composition of the oral biofilm. Statherin may protect the host from colonization of bacteria associated with infections. Gp-340 size variants may affect functions related to host innate immune defences such as interactions with a wide array of bacteria, and human milk may have a protective effect in infants from colonization of mutans streptococci.

Place, publisher, year, edition, pages
Umeå: Umeå university , 2010. , 66 p.
Series
Umeå University odontological dissertations, ISSN 0345-7532 ; 112
Keyword [en]
Adhesion, statherin, gp-340, caseins, human milk, saliva, Streptococcus mutans, Actinomyces
National Category
Dentistry
Research subject
Cariology
Identifiers
URN: urn:nbn:se:umu:diva-32894ISBN: 978-91-7264-969-9 (print)OAI: oai:DiVA.org:umu-32894DiVA: diva2:306593
Public defence
2010-04-22, Sal D Tandläkarhögskolan, Umeå Universitet, Umeå, 09:00 (Swedish)
Opponent
Supervisors
Available from: 2010-03-31 Created: 2010-03-30 Last updated: 2010-03-31Bibliographically approved
List of papers
1. Salivary statherin peptide-binding epitopes of commensal and potentially infectious Actinomyces spp. delineated by a hybrid peptide construct
Open this publication in new window or tab >>Salivary statherin peptide-binding epitopes of commensal and potentially infectious Actinomyces spp. delineated by a hybrid peptide construct
2004 (English)In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 72, no 2, 782-787 p.Article, review/survey (Refereed) Published
Abstract [en]

Adhesion of microorganisms to host receptor molecules such as salivary statherin molecules is a common event in oral microbial colonization. Here we used a hybrid peptide construct (with both a hydroxyapatite-binding portion and a test peptide portion) to map the interaction of Actinomyces species (and Candida albicans) with statherin. Adhesion to hybrid peptides and truncated statherin variants revealed three binding types, types I to III. (i) Type I strains of rat, hamster, and human infection origins bound C-terminal-derived QQYTF and PYQPQY peptides. The QQYTF peptide inhibited statherin binding for some strains but not for others. (ii) Type II strains of human and monkey tooth origins bound middle-region-derived YQPVPE and QPLYPQ peptides. Neither strain was inhibited by soluble peptides. (iii) Type III strains of human infection origins (and C. albicans) did not bind to either statherin-derived peptides or truncated statherin. Moreover, the type I strains inhibited by QQYTF were also inhibited by TF and QAATF peptides and were detached from statherin by the same peptides. In conclusion, it is suggested that commensal and potentially infectious microorganisms bind middle or C-terminal statherin differently and that other microbes might require discontinuous epitopes.

National Category
Dentistry
Research subject
Cariology
Identifiers
urn:nbn:se:umu:diva-16969 (URN)10.1128/IAI.72.2.782-787.2004 (DOI)14742521 (PubMedID)
Available from: 2007-10-25 Created: 2007-10-25 Last updated: 2017-12-14Bibliographically approved
2. Variant size- and glycoforms of the scavenger receptor cysteine-rich protein gp-340 with differential bacterial aggregation
Open this publication in new window or tab >>Variant size- and glycoforms of the scavenger receptor cysteine-rich protein gp-340 with differential bacterial aggregation
Show others...
2007 (English)In: Glycoconjugate Journal, ISSN 0282-0080, E-ISSN 1573-4986, Vol. 24, no 2-3, 131-142 p.Article in journal (Refereed) Published
Abstract [en]

Glycoprotein gp-340 aggregates bacteria in saliva as part of innate defence at mucosal surfaces. We have detected size- and glycoforms of gp-340 between human saliva samples (n = 7) and lung gp-340 from a proteinosis patient using antibodies and lectins in Western blots and ELISA measurements. Western blots of saliva samples, and of gp-340 purified, from the seven donors using a gp-340 specific antibody distinguished four gp-340 size variants, designated I to IV (n = 2,2,2 and 1). While saliva gp-340 variants I to III had single bands of increasing sizes, variant IV and lung gp-340 had double bands. Purified I to IV proteins all revealed a N-terminal sequence TGGWIP upon Edman degradation. Moreover, purified gp-340 from the seven donors and lung gp-340 shared N-glycans, sialylated Galbeta1-3GalNAc and (poly)lactosamine structures. However, the larger size gp-340 grouping II/III (n = 4) and smaller size grouping I/IV correlated with a secretor, Se(+), and a non secretor, Se(-), dependent glycoform of gp-340, respectively (p = 0.03). The Se(+) glycoforms contained ABH, Le(b), Le(y) and polylactosamine structures, while the Se(-) glycoforms lacked ABH antigens but expressed Le(a), Le(x) and lactosamine structures. By contrast, lung gp-340 completely lacked ABH, Le(a/b), Le(x/y) or sLe(x) structures. Gp-340 and secretor typing of saliva from additional donors (n = 29) showed gp-340 glycoforms I to IV for 6, 16, 4 and 0 donors, respectively, and 3 non-typeable donors, and verified that gp-340 glycoforms I and II/III correlate with Se(-) and Se(+) phenotypes, respectively (p < 0.0001). The glycoforms of saliva and lung gp-340 mediated differential aggregation of Le(b)- (Helicobacter pylori), sialylpolylactosamine- (Streptococcus suis) or sialic acid- (Streptococcus mutans) binding bacteria. In conclusion, variant size- and glycoforms of gp-340 are expressed by different individuals and may modulate the biological properties of gp-340 pertinent to health and disease.

National Category
Dentistry
Research subject
Cariology
Identifiers
urn:nbn:se:umu:diva-17130 (URN)10.1007/s10719-006-9020-1 (DOI)17243023 (PubMedID)
Available from: 2007-11-01 Created: 2007-11-01 Last updated: 2017-12-14Bibliographically approved
3. Effects of human milk on adhesion of Streptococcus mutans to saliva-coated hydroxyapatite in vitro
Open this publication in new window or tab >>Effects of human milk on adhesion of Streptococcus mutans to saliva-coated hydroxyapatite in vitro
Show others...
2006 (English)In: Caries Research, ISSN 0008-6568, E-ISSN 1421-976X, Vol. 40, no 5, 412-417 p.Article, review/survey (Refereed) Published
Abstract [en]

Adhesion of bacteria to pellicle-coated tooth surfaces is the first step in biofilm formation on teeth. The aim of this study was to explore whether human milk prevents or promotes adhesion of cariogenic Streptococcus mutans to saliva-coated hydroxyapatite (HA) using an in vitro model system. S. mutans binding to HA coated with human parotid saliva (s-HA) or human milk was studied, in addition to binding inhibition to s-HA by human milk. S. mutans did not bind to HA coated with milk. S. mutans binding to s-HA was inhibited by milk from 15 (71 %) of 21 women, whereas milk from the remaining 6 mothers enhanced binding of S. mutans to s-HA. Inhibition of S. mutans binding correlated with bacterial binding to s-HA (r = 0.76). Human milk does not mediate adhesion of S. mutans to HA in vitro, but affects adhesion in an individually varying fashion. Phenotypic variations in milk and saliva glycosylation may explain the inhibitory capacity and possibly affect susceptibility to colonization by S. mutans in childhood. Copyright 2006 S. Karger AG, Basel.

National Category
Dentistry
Research subject
Cariology
Identifiers
urn:nbn:se:umu:diva-12189 (URN)10.1159/000094287 (DOI)
Available from: 2008-02-01 Created: 2008-02-01 Last updated: 2017-12-14Bibliographically approved
4. Human milk compounds inhibiting adhesion of mutans streptococci to host ligand-coated hydroxyapatite in vitro
Open this publication in new window or tab >>Human milk compounds inhibiting adhesion of mutans streptococci to host ligand-coated hydroxyapatite in vitro
2009 (English)In: Caries Research, ISSN 0008-6568, E-ISSN 1421-976X, Vol. 43, no 3, 171-178 p.Article in journal (Refereed) Published
Abstract [en]

Acquisition of mutans streptococci at an early age is a risk factor for later caries development. Following our recent finding that human milk may inhibit adhesion of Streptococcus mutans the aim of the present study was to identify compounds in human milk preventing adhesion of mutans streptococci to saliva- or gp340-coated hydroxyapatite (s-HA and gp340-HA) using an in vitro model system. Superdex 200 fractions of human milk and purified proteins were screened for binding inhibition of the S. mutans strain Ingbritt. Avid inhibition was seen to both s-HA and gp340-HA for caseins, lactoferrin, IgA and IgG, and moderate inhibition for alpha-lactalbumin and bile salt-stimulated lipase, whereas albumin and lysozyme had no effect. The inhibitory epitope in beta-casein was delineated to its C-terminal LLNQELLNPTHQIYPVTQPLAPVHNPISV stretch by use of synthetic peptides. Similarly, a peptide (SCKFDEYFSQSCA) corresponding to the human lactoferrin stretch that is highly homologous to the previously shown inhibitory stretch of bovine lactoferrin was found to inhibit S. mutans Ingbritt binding. Inhibition by human milk, IgA, and the inhibitory beta-casein peptide was universal among 4 strains of S. mutans (Ingbritt, NG8, LT11, JBP) and 2 strains of S. sobrinus (6715 and OMZ176). IgG inhibited 4, alpha-lactalbumin 3 and lactoferrin 2 of these 6 strains. It was also confirmed that none of the milk components coated on HA mediated S. mutans Ingbritt adhesion, which was consistent with the finding that no milk protein was recognized on Western blots by gp340/DMBT1 monoclonal antibodies.

Keyword
Adhesion of mutans streptococci, casein, human milk, saliva, Streptococcus mutans
National Category
Dentistry
Research subject
Cariology
Identifiers
urn:nbn:se:umu:diva-31698 (URN)10.1159/000213888 (DOI)19390191 (PubMedID)
Available from: 2010-02-15 Created: 2010-02-15 Last updated: 2017-12-12Bibliographically approved

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