Shortest Amino Acid Sequence Required for Statherin to Reduce Demineralisation

Objectives: The salivary protein statherin (StN43) can reduce the rate of deminerlisation. Statherin and its shortened analogues containing only the N-terminal 21 amino acids (StN21) have been shown to significantly protect enamel against cariogenic challenges in vitro. Recent computational studies have shown that statherin's interactions with hydroxyapatite surfaces are almost all at its N-terminal residues, particularly Arg13 [Makrodimitris et al.: JACS 2007; 129:13713–13722]. The aim was to identify the minimum length of functional domain of statherin required for cariostatic function. The efficacy of decreasingly shorter length peptides containing only the N-terminal 21,15,10 and 5 amino acid residues of statherin in reducing the rate of porous hydroxyapatite pellet (HAp) mineral loss under simulated cariogenic challenges was measured using scanning microradiography (SMR).

Methods: Statherin-like peptides containing the N-terminal 5, 10, 15 and 21 residues respectively were prepared using FMOC synthesis, and dissolved in phosphate buffer at pH 7.4 (0.188 mM). HAp pellets (20% porosity) used as model substrates for enamel demineralisation studies were sectioned and mounted in scanning microradiography (SMR) environmental cells, and exposed to 0.1M acetic acid at pH4.0 for 120h. The sections were then treated with peptide solution for 24h, then demineralized for a further 120h. SMR was used to measure HAp demineralisation rates (RD_HAp) before and after exposure to peptide solution during simulated cariogenic challenges.

Results: Coating with StN21 resulted in a 40% reduction and StN15 a 35% reduction in RD_HAp. StN10, StN5 and buffer only coatings showed no cariostatic efficacy.

Conclusions: Computational chemistry studies of statherin binding to hydroxyapatite demonstrate that the N-terminal residues of statherin including Arg13 are required for binding of statherin to HAp. This may be a requirement for its cariostatic function. Statherin-like peptides containing its N-terminal residues of length greater than 15 amino acids are required for cariostatic function.