Protein Interaction between Ameloblastin and Proteasome Subunit α Type 3 Can Facilitate Redistribution of Ameloblastin Domains within Forming Enamel

J Biol Chem. 2015 Aug 21;290(34):20661-20673. doi: 10.1074/jbc.M115.640185. Epub 2015 Jun 12.


Enamel is a bioceramic tissue composed of thousands of hydroxyapatite crystallites aligned in parallel within boundaries fabricated by a single ameloblast cell. Enamel is the hardest tissue in the vertebrate body; however, it starts development as a self-organizing assembly of matrix proteins that control crystallite habit. Here, we examine ameloblastin, a protein that is initially distributed uniformly across the cell boundary but redistributes to the lateral margins of the extracellular matrix following secretion thus producing cell-defined boundaries within the matrix and the mineral phase. The yeast two-hybrid assay identified that proteasome subunit α type 3 (Psma3) interacts with ameloblastin. Confocal microscopy confirmed Psma3 co-distribution with ameloblastin at the ameloblast secretory end piece. Co-immunoprecipitation assay of mouse ameloblast cell lysates with either ameloblastin or Psma3 antibody identified each reciprocal protein partner. Protein engineering demonstrated that only the ameloblastin C terminus interacts with Psma3. We show that 20S proteasome digestion of ameloblastin in vitro generates an N-terminal cleavage fragment consistent with the in vivo pattern of ameloblastin distribution. These findings suggest a novel pathway participating in control of protein distribution within the extracellular space that serves to regulate the protein-mineral interactions essential to biomineralization.

Keywords: 20S proteosome; ameloblastin; biomineralization; enamel microstructure pattern; extracellular matrix protein; proteasome subunit α type 3; protein-protein interaction; tooth; yeast two-hybrid.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Ameloblasts / cytology
  • Ameloblasts / metabolism*
  • Animals
  • Cytoplasm / chemistry
  • Cytoplasm / metabolism
  • Dental Enamel / cytology
  • Dental Enamel / growth & development
  • Dental Enamel / metabolism*
  • Dental Enamel Proteins / genetics
  • Dental Enamel Proteins / metabolism*
  • Extracellular Matrix / chemistry
  • Extracellular Matrix / metabolism
  • Gene Expression Regulation, Developmental
  • Gene Library
  • Glutamate Carboxypeptidase II / genetics
  • Glutamate Carboxypeptidase II / metabolism*
  • Humans
  • Incisor / cytology
  • Incisor / growth & development
  • Incisor / metabolism*
  • Membrane Glycoproteins / genetics
  • Membrane Glycoproteins / metabolism*
  • Mice
  • Mutation
  • Odontogenesis / genetics*
  • Proteasome Endopeptidase Complex / metabolism
  • Protein Binding
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Protein Structure, Tertiary
  • Protein Transport
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Signal Transduction
  • Two-Hybrid System Techniques


  • Ambn protein, mouse
  • Dental Enamel Proteins
  • Membrane Glycoproteins
  • Protein Isoforms
  • Recombinant Fusion Proteins
  • Glutamate Carboxypeptidase II
  • PSMA protein, mouse
  • Proteasome Endopeptidase Complex