Small-Angle X-Ray Scattering and Computer-Aided Molecular Modeling Studies of 20 kDa Fragment of Porcine Amelogenin: Does Amelogenin Adopt an Elongated Bundle Structure?

Abstract

Amelogenins, which are major matrix constituents in the developing tooth, play a regulatory role in the process of enamel crystal formation. Porcine amelogenin with 173 amino acid residues is rich in proline, glutamine, leucine, and histidine. We utilized the small-angle X-ray scattering (SAXS) technique to examine the solution structure of porcine amelogenin. Samples used were two porcine amelogenins with apparent molecular weights of 20 kDa (amino acids 1 to 148) and 13 kDa (amino acids 46 to 148) on SDS-PAGE. Prior to SAXS measurements, the protein samples were dissolved in 2% (v/v) acetic acid to give a concentration range up to 10 mg/ml. Comparison between Rg (the overall radius of gyration) and Rc (the cross-sectional radius of gyration) revealed that the 20 kDa amelogenin exists in this solution as asymmetric particles with a length of about 15 nm, presumably corresponding of dimers. Based on these experimental data and computer-aided molecular modeling studies, we propose that the 20 kDa amelogenin adopts an elongated bundle structure which mainly consists of extended structures similar to polyproline II and/or beta-strand, interspersed with beta-turn or loop.

The onset of amelogenin nanosphere aggregation studied by small-angle X-ray scattering and dynamic light scattering

“…Similar data were collected for rM179 at pH 4.5, and rM166 at pH < 6. The scattering curves for these samples were very similar to published synchrotron SAXS measurements of a native porcine amelogenin (P148) fragment of the full-length pig protein (P173) at low pH (Matsushima et al, 1998). They report a radius of gyration of about 3.5 nm which is perfectly consistent with the present data.…”

The leucine rich amelogenin protein (LRAP) adsorbs as monomers or dimers onto surfaces

“…This is consistent with a large number of studies of quaternary structures of the full-length amelogenin protein at pH values in the range of 6–8 using DLS (Aichmayer et al, 2005; Du et al, 2005; Moradian-Oldak et al, 1998b, 1994; Petta et al, 2006), small angle X-ray scattering (SAXS) (Aichmayer et al, 2005), and small angle neutron scattering (SANS) (Aichmayer et al, 2005). Small structures in the size range of monomers to dimers have been observed in solution, but primarily in acidic solutions less than pH 4 (Aichmayer et al, 2005; Matsushima et al, 1998; Petta et al, 2006) and from less polar solvents such as 60% acetonitrile in water (Du et al, 2005). We did a few adsorption studies from solutions of LRAP dissolved in 60% acetonitrile or acetic acid at pH 3 and observed monomer/dimer-sized structures, similar to the small structures found in this study at pH 7.4.…”

Protein Phosphorylation and Mineral Binding Affect the Secondary Structure of the Leucine-Rich Amelogenin Peptide

“…In an overlapped region to this, peaks observed between 1,610 and 1,640 cm −1 were attributed to β-sheet (Susi and Byler, 1983; Jackson and Mantsch, 1995). Random coil conformation was attributed to peaks between 1,640 and 1,650 cm −1 (Krimm and Bandekar, 1986; Barth and Zscherp, 2002; Elangovan et al, 2007), which have also been reported in amelogenin (Renugopalakrishnan et al, 1986; Goto et al, 1993; Matsushima et al, 1998; Elangovan et al, 2007; Yang et al, 2010). Also, peaks observed between 1,650 and 1,655 cm −1 were attributed to α-helix conformation (Susi and Byler, 1983; Surewicz et al, 1993; Roach et al, 2005).…”