Abstract
Objectives: To study the role of amelotin in enamel formation, purified recombinant human amelotin was used in a study of amelotin self assembly and its role in new HAP crstal structure formation in a laboratory setting. Methods: The full-length human amelotin gene was cloned from developing tooth buds and recombinant amelotin was expressed and purified. Morphological changes during amelotin self-assembly and the role of amelotin in promoting new minieralization structure is studied using Transmission electronic microscopy and Scanning electron microscopy. Results: The measured molecular weight of expressed human amelotin was 28 kDa that matches the predicted values. Amelotin self-assembles to form nano-spheres 10 - 30 nm in size. The amelotin self-assembly process further produces higher 3-D structure in the forms of chain-like fabric arranged in parallel and perpendicular direction. The neonatal pearl crystals were arranged side by side to form a string of small rods and small parallel rods extend from the demineralized glaze on the surface of teeth that were pre-treated with amelotin.
Highlights
The primary unit of tooth enamel is tightly packed hydroxyapatite (HAP) nanocrystals with a rough cross section of 33 - 65 nm and a c-axis length of 100 1000 nm [1] [2]
Expression Optimization of Recombinant Full-Length hAMTN Protein pET-28a (+)-AMTN was transformed into the competent E. coli strain BL21 (DE3) and the clones containing hAMTN gene were selected by colony PCR (Figure 1(C))
The 28 kDa band was weak in the supernatant, indicating that the hAMTN protein was mainly expressed in the form of inclusion bodies (Figure 1(D))
Summary
The primary unit of tooth enamel is tightly packed hydroxyapatite (HAP) nanocrystals with a rough cross section of 33 - 65 nm and a c-axis length of 100 1000 nm [1] [2]. The formation of dental enamel is initiated enamel organ and dental papilla [3]. The ameloblasts secreted extracellular matrix proteins (EMPs), which play a major role in the following biomineralization p [4]. Three main structural proteins in the EMPs are amelogenin (AMEL) enamelin (ENAM) and amphiphilic ameloblastin (AMBN) [5]. We subcloned the human recombinant AMTN gene and analyzed the crystal morphology and mineral phase of the surface of the nanocomposite in the presence of the recombinant human full-length AMTN using field emission scanning electron microscopy (FE-SEM) and X-ray Diffraction (XRD)
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