Altered Amelogenin Self-assembly Based on Mutations Observed in Human X-linked Amelogenesis Imperfecta (AIH1)

Michael L Paine,Ya-Ping Lei,Kenneth Dickerson,Malcolm L Snead

doi:10.1074/jbc.m110473200

Michael L Paine, Ya-Ping Lei + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m110473200

Copy DOI

Abstract

A hallmark of biological systems is a reliance on protein assemblies to perform complex functions. We have focused attention on mammalian enamel formation because it relies on a self-assembling protein complex to direct mineral habit. The principle protein of enamel is amelogenin, a 180-amino acid hydrophobic protein that self-assembles to form nanospheres. We have used independent technical methods, consisting of the yeast two-hybrid (Y2H) assay and surface plasmon resonance (SPR), to demonstrate the importance of amelogenin self-assembly domains. In addition, we have analyzed mutations in amelogenin observed in patients with amelogenesis imperfecta who demonstrate defects in enamel formation. Assessments of self-assembly of these mutant amelogenins by either SPR or Y2H assay yield concordant data. These data support the conclusion that the amelogenin amino-terminal self-assembly domain is essential to the creation of an enamel extracellular organic matrix capable of directing mineral formation. It also suggests that a pathway through which point mutations in the amelogenin protein can adversely impact on the formation of the enamel organ is by disturbing self-assembly of the organic matrix. These data support the utilization of the Y2H assay to search for protein interactions among extracellular matrix proteins that contribute to biomineralization and provide functional information on protein-protein and protein-mineral interactions.

Highlights

The self-assembly of a protein complex is a hallmark of biological systems
We have focused attention on mammalian enamel formation because it relies on a self-assembling protein complex to direct mineral habit
We have focused attention on mammalian enamel as an example of the self-assembly of an extracellular matrix protein complex

Summary

Introduction

The self-assembly of a protein complex is a hallmark of biological systems. From processive enzymatic reactions to extracellular matrices, cells achieve control over their microenvironment through the assembly of a mixture of proteins. The Y2H assay and in vitro assembly experiments demonstrated that the amino acid substitution interfered with the ability of the mutated collagen molecules to engage in trimerization (1). In this example, confirmation of a valid animal model for a human genetic disease has been possible in part by the Y2H assay. Amelogenesis imperfecta is a hereditary disease of enamel and, when linked to the amelogenin locus, severely reduces the ability of amelogenin to self-assembly as measured by either the Y2H assay or by SPR. Enhanced understanding of the physicochemical guidelines governing protein self-assembly and the ability to manipulate these interactions will contribute to the studies of nanotechnology and biomimetics (10)

Methods

Results

Conclusion