Neutron reflectometry studies of the adsorbed structure of the amelogenin, LRAP

Barbara J. Tarasevich, Ursula Perez-Salas, David L. Masica, John Philo, Paul Kienzle, Susan Krueger, Charles F. Majkrzak, Jeffrey L. Gray, Wendy J. Shaw

Research output: Contribution to journalArticle

Abstract

Amelogenins make up over 90% of the protein present during enamel formation and have been demonstrated to be critical in proper enamel development, but the mechanism governing this control is not well understood. Leucine-rich amelogenin peptide (LRAP) is a 59-residue splice variant of amelogenin and contains the charged regions from the full protein thought to control crystal regulation. In this work, we utilized neutron reflectivity (NR) to investigate the structure and orientation of LRAP adsorbed from solutions onto molecularly smooth COOH-terminated self-assembled monolayer (SAM) surfaces. Sedimentation velocity (SV) experiments revealed that LRAP is primarily a monomer in saturated calcium phosphate (SCP) solutions (0.15 M NaCl) at pH 7.4. LRAP adsorbed as ∼32 Å thick layers at ∼70% coverage as determined by NR. Rosetta simulations of the dimensions of LRAP in solution (37 Å diameter) indicate that the NR determined z dimension is consistent with an LRAP monomer. SV experiments and Rosetta simulations show that the LRAP monomer has an extended, asymmetric shape in solution. The NR data suggests that the protein is not completely extended on the surface, having some degree of structure away from the surface. A protein orientation with the C-terminal and inner N-terminal regions (residues ∼8-24) located near the surface is consistent with the higher scattering length density (SLD) found near the surface by NR. This work presents new information on the tertiary and quaternary structure of LRAP in solution and adsorbed onto surfaces. It also presents further evidence that the monomeric species may be an important functional form of amelogenin proteins.

Original languageEnglish (US)
Pages (from-to)3098-3109
Number of pages12
JournalJournal of Physical Chemistry B
Volume117
Issue number11
DOIs
StatePublished - Mar 21 2013

ASJC Scopus subject areas

  • Physical and Theoretical Chemistry
  • Surfaces, Coatings and Films
  • Materials Chemistry

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    Tarasevich, B. J., Perez-Salas, U., Masica, D. L., Philo, J., Kienzle, P., Krueger, S., Majkrzak, C. F., Gray, J. L., & Shaw, W. J. (2013). Neutron reflectometry studies of the adsorbed structure of the amelogenin, LRAP. Journal of Physical Chemistry B, 117(11), 3098-3109. https://doi.org/10.1021/jp311936j