Dental enamel is an excellent example of a highly mineralised tissue, composed of hierarchically organised apatite mineral. This unique organisation gives enamel superior mechanical properties. However, when mature, enamel becomes acellular and unable to repair itself during traumatic or carious damage. The lack of self-repair requires dental intervention, where the common treatment of decayed enamel is to remove the affected and healthy tissue, and replace with restorative materials. The restorative materials, currently used, can cause further complications in the form of secondary caries or failure due to thermal and mechanical property mismatch with enamel. Problems associated with current restorative materials have driven researchers to explore biomimetic enamel treatment routes. To mimic the natural enamel formation, we can explore how proteins can guide mineral growth, in order to form enamel-like ordered mineral structures. In this thesis, the use of a synthetic, recombinant protein called an elastin-like protein (ELP) containing the analogue of the N-terminal of statherin (STNA15) was under investigation. Statherin is a protein present in saliva that is said to aid in the remineralisation of enamel. ELP with STNA15 (STNA15-ELP) has already shown promise in biomimetic mineralisation. This thesis investigated how conformation and structure of STNA15-ELP can be affected and manipulated by different chemical environments, surface constraint and crosslinking. The STNA15-ELP characteristics were related to formation of fluorapatite. STNA15-ELP conformation changed due to presence if salts in solution and whether or not it was constrained. We linked the conformational changes within STNA15-ELP, in solution versus on the surface, to two different routes of mineral formation. FAp formed in an uncontrolled manner with free STNA15-ELP. Ordered FAp formed via a precursor when STNA15-ELP was constrained on a surface. This work leads to an understanding of biomimetic mineralisation using STNA15-ELP. This information can aid in the design of novel biomimetic, enamel-like therapeutics.

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.

Aim: The aim of this study is to determine the varying levels of salivary statherin production in patients with varying levels of risk for diabetes. The goal is to identify a causal relationship and thus, statherin could be used as a preliminary biomarker for identifying patients with diabetes. Materials and Methods: Saliva from 47 participants were collected in order to quantify the levels of statherin production via western blot analysis. Participants were also asked to fill out self-reported questionnaires regarding risk factors for type 2 diabetes. The questionnaire consisted of 7 questions regarding age, sex, history of diabetes, hypertension, level of physical activity, and weight class. Each individual factor as well as total risk for type two diabetes was compared to levels of salivary statherin levels via the unpaired t-test and one way ANOVA testing. Results: Risk factors for type two diabetes such as; age, sex, history of diabetes, hypertension, level of physical activity, and weight class showed no correlation to levels of salivary statherin secretion. All risk factors combined as a total risk level for type two diabetes also did not show a correlation to levels of salivary statherin secretion. Conclusions: It can be concluded in this study that salivary statherin protein does not show a correlation for risk or status of type two diabetes. Salivary statherin does not act as a useful biomarker for detection of type two diabetes.

Diss. (sammanfattning) Umeå : Umeå universitet, 2010.