Polyelectrolyte-Mediated Adsorption of Amelogenin Monomers and Nanospheres Forming Mono- or Multilayers

Abstract

We have applied optical waveguide lightmode spectroscopy combined with streaming potential measurements and Fourier transformed infrared spectroscopy to investigate adsorption of amelogenin nanospheres onto polyelectrolytes. The long term objective was to better understand the chemical nature of these assemblies and to gain further insight into the molecular mechanisms involved during self-assembly. It was found that monolayers of monomers and negatively charged nanospheres of a recombinant amelogenin (rM179) irreversibly adsorbed onto a positively charged polyelectrolyte multilayer films. Based on measurements performed at different temperatures it was demonstrated that intermolecular interactions for the formation of nanospheres were not affected by their adsorption onto polyelectrolytes. Consecutive adsorption of nanospheres resulting in the formation of multilayer structures was possible by using cationic poly(L-lysine) as mediators. NAcetyl-D-Glucosamine (GlcNac) did not disturb the nanosphere-assembled protein's structure and it only affected the adsorption of monomeric amelogenin. Infrared spectroscopy of adsorbed amelogenin revealed conformational differences between the monomeric and assembled forms of rM179. While there was a considerable amount of α-helices in the monomers, β-turn and β-sheet structures dominated the assembled proteins. Our work constitutes the first report on a structurally controlled in-vitro buildup of an rM179 nanosphere monolayer-based matrix. Our data support the notion that amelogenin self-assembly is mostly driven by hydrophobic interactions and that amelogenin/PEM interactions are dominated by electrostatic forces. We suggest that similar forces can govern amelogenin interactions with non-amelogenins or the mineral phase during enamel biomineralization.

Effects of phosphorylation on the self-assembly of native full-length porcine amelogenin and its regulation of calcium phosphate formation in vitro

“…Hence, these collective findings suggest that hydrophobic interactions between amelogenins play an important role in amelogenin self-assembly. A similar conclusion was drawn from studies on the adsorption of recombinant full-length mouse amelogenin (rM179) onto positively charged polyelectrolyte films [60]. Given the fact that amelogenin primarily contains hydrophobic residues, it is reasonable that hydrophobic interactions represent an important factor with respect to the regulation of protein-protein interactions.…”

Prospects and Pits on the Path of Biomimetics: The Case of Tooth Enamel

“…Recent combined small-angle X-ray scattering (SAXS) and dynamic light scattering (DLS) experiments have indicated that a certain ellipticity (with the aspect ratio in the range of 0.45℃0.5) may be attributed to amelogenin assemblies [39]. Limited proteolysis experiments and those based on polyelectrolyte multilayers have indicated that regions at both C- and N- termini are exposed on the surface of the nanospheres [40, 41]. Accordingly, the structure of amelogenin nanospheres is usually depicted as the one with both C- and N- terminal regions present at the nanosphere surface (Figure 5), with the hydrophilic C-terminal supposedly in contact with apatite [8].…”

Mammalian Enamel Formation

“…By applying the small-angle X-ray scattering (SAXS) technique a model of a dense hydrophobic core with a flexible hydrophilic region exposed on the surface of amelogenin molecules has also been proposed [120]. Further support for the notion that amelogenin nanospheres assemble to possess negatively charged surfaces was obtained by applying optical waveguide lightmode spectroscopy combined with streaming potential measurements [121]. Amelogenin nanospheres adsorbed on a positively charged polyeletrolyte multilayer film surface in a single monolayer while successive multilayer adsorption was possible using a cationic mediator.…”