Abstract:
:The maturation of dental enamel succeeds the degradation of organic matrix. Inhibition studies have shown that this degradation is accomplished by a serine-type proteinase. To isolate and characterize cDNA clones encoding this proteinase, we used two degenerate primer approaches to amplify part of the coding region using polymerase chain-reaction (PCR). First, we purified the proteinase from porcine transition-stage enamel matrix and characterized it by partial protein sequencing. The enzyme was isolated from the neutral soluble enamel extract by successive ammonium sulfate precipitations, hydroxyapatite HPLC, reverse-phase HPLC, DEAE ion exchange, and affinity chromatography with a Benzamidine Sepharose 6B column. The intact protein and lysylendopeptidase-generated cleavage products were characterized by amino acid sequence analyses. Degenerate oligonucleotide primers encoding two of the polypeptide sequences were synthesized. In a complementary strategy, degenerate oligonucleotide primers were designed against highly conserved active-site regions of chymotrypsin-like proteinases. Both approaches yielded PCR amplification products that served as probes for screening a porcine enamel organ epithelia-specific cDNA library. The longest full-length clone is 1133 nucleotides and encodes a preproprotein of 254 amino acids. We designate this protein enamel matrix serine proteinase 1 or EMSP1. The active protein has 224 amino acids, an isotope-averaged molecular mass of 24.1 kDa, and an isoelectric point of 6.0. Multiple-tissue Northern analysis indicates that EMSP1 is a tooth-specific protein. Gelatin enzymography shows a dramatic increase in EMSP1 activity in the transition-stage enamel matrix. EMSP1 is most homologous to kallikriens and trypsins.
journal_name
J Dent Resjournal_title
Journal of dental researchauthors
Simmer JP,Fukae M,Tanabe T,Yamakoshi Y,Uchida T,Xue J,Margolis HC,Shimizu M,DeHart BC,Hu CC,Bartlett JDdoi
10.1177/00220345980770020601subject
Has Abstractpub_date
1998-02-01 00:00:00pages
377-86issue
2eissn
0022-0345issn
1544-0591journal_volume
77pub_type
杂志文章abstract::The objective of this study was to calculate the stress distribution induced in anterior metal-ceramic crowns fabricated with either gold-alloy or nickel-alloy copings of reduced thickness using plane stress analyses. Two-dimensional finite element models of three crown designs were subjected to a simulated biting for...
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