Chemistry Reference
In-Depth Information
weight approximately 27 kDa. The molecule has essentially three domains. The first 50
amino-acid terminal sequence of the secreted protein is highly conserved in most species
and contains an internal YxxxxYxxxxxxWYxxxxxxxYxxYxY motif, where W is
tryptophane and Y is tyrosine as in the human sequence:
MPLPPHPGHPG
Y
INFS*
Y
EVLTPLKW
Y
QSMIRPP
Y
SS
Y
G
Y
EMPG
GW
LHHQ-
The underlined GW is a conserved proteolytic cleavage site between G45 and W46, The
S* represents a single phosphorylation site. The cleaved N-terminal 45-mer peptide is
designated as the TRAP peptide. Another highly conserved sequence is at the C-terminus.
The final 29 residues, corresponding to exon 6d and 7,
PLPPMLPDLTEAWP
STDKTKREEVD
is polar and acidic. This acidic tail is a very important feature of the intact amelogenin
molecule. The splice product peptide comprised from the amino acids encoded by exons
2, 3, 5, 6d,7, 59 amino acids in length, is known as the “leucine rich amelogenin peptide”
LRAP. The hydrophilic 14 residue peptide encoded by exon 4 is usually not expressed,
the long peptide sequence encoded by exon 6a,b,c in the large molecular weight
amelogenin is quite hydrophobic.
Ameloblastin (also called amelin or sheathlin) is the second most abundant protein in
the secretory enamel matrix. The nascent AMBN has a M
r
68 kDa but this is rarely
found in the enamel matrix since, like the amelogenin, it is proteolytically processed
rapidly to a series of peptides with apparent molecular weights of 52, 40, 37, 19, 17, 16,
15, 14 and 13 kDa, all of which have been detected on gels (Brookes et al. 2001). In the
rat incisor, the 68, 52, 40, 37 and 13 kDa forms are readily soluble and can be extracted
in simulated enamel fluid. The 19, 17 and 16 kDa proteins were only partly soluble in the
enamel fluid and required phosphate buffers to wash them from the mineral surface. The
15 and 14 kDa peptides and the final part of the 17 and 16 kDa peptides were obtained
only after dissociative extraction in SDS-containing buffer, apparently because they
formed an insoluble aggregate
in vivo
and were bound to the mineral phase.
In vivo
the
AMBN proteins were completely absent from mature enamel. The function of the AMBN
is not known, but the aggregated peptides may be accumulated at the growing prism
surfaces and prism boundaries, hence the name sheathlin.
Enamelin is present in still smaller amount in the secretory enamel, but it is strongly
bound and represents the major protein of the mature enamel, bound to the enamel
crystals. It cannot be dissolved without dissolution of the mineral phase (Termine et al.
1980). Mouse enamelin has been cloned (Hu et al. 1998) and is postulated to have a
sequence of 1236 amino acids, a pI of 9.4 and a calculated molecular mass of 137 kDa
without consideration of any post-translational modifications. It is immediately processed
upon secretion into the Tomes' process compartment to lower mass fragments. It is
processed stepwise from the carboxyl terminus to products of 155, 142, and 89 k apparent
molecular mass, each of which retains the amino-terminus (Fukae et al. 1996). The 89
kDa form accumulates but is further processed to 32 and 25 kDa products. The 32 kDa
form is glycosylated and phosphorylated and has an isoelectric point of pH 6.4, and
accounts for about 1% of the enamel matrix protein (Tanabe et al. 1990). The enamelin
degradation products are present in the secretory enamel, but disappear as the enamel
matures. The enamelin is not present in the prism sheaths but is restricted to the rod and
interrod crystallites. The functions of the enamelin cleavage products are not known
(Uchida et al 1998).
The final enamel matrix protein of note is tuftelin, a protein localized to the DEJ,
potentially nucleating crystallization of the enamel rods. There have been several
≈
