PNNL

Abstract

The organic matrix in forming enamel consists largely of the amelogenin protein, self-assembled into nanospheres that are necessary to guide the formation of the unusually long and highly ordered hydroxyapatite (HAP) crystallites that constitute enamel. Despite its ability to direct crystal growth, the interaction of the amelogenin protein with HAP is unknown, though the charged C-terminus is often implicated in this function. To elucidate if the C-terminus is important in the binding and orientation of amelogenin with HAP, we used solid state NMR to determine the orientation of the C-terminus of an amelogenin splice variant, LRAP, which contains the charged C-terminus of the full protein, on the HAP surface. These experiments demonstrate that the sidechain methyl labeled residue, A46, is 8 Å from the HAP surface, under hydrated conditions, for the protein with and without phosphorylation. Modeling results are consistent with experimental measurements, resulting in an average 13Cala46-31P distance of 7.62 ± 1.8 Å for LRAP orientation onto the 100 face of HAP. The computational model also reveals that with 13Cala46 at this distance, the negatively charged residues are energetically favored to align themselves with the Ca2+ in the HAP lattice. The experimental results and supporting computational models provide direct evidence orienting the charged C-terminal region of the amelogenin protein on the HAP surface, optimized to exert control on developing enamel crystals.