Journal
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
Volume 1830, Issue 3, Pages 2600-2607Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbagen.2012.11.021
Keywords
Enamelysin; Amelogenin; Enamel; Tooth; Proteolysis; Enzyme kinetics
Categories
Funding
- NIH/NIDCR [DE-RO1-017529, RO1-017529S2, 2R01DE015821, 3R01DE015821S1]
- Sandler Family Foundation
- Gordon and Betty Moore Foundation
- NIH/NCI Cancer Center [P30 CA082103]
- Office of Science, Office of Biological and Environmental Research of the U.S. Department of Energy [DE-AC02-05CH11231]
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Background: Enamel synthesis is a highly dynamic process characterized by simultaneity of matrix secretion, assembly and processing during apatite mineralization. MMP-20 is the first protease to hydrolyze amelogenin, resulting in specific cleavage products that self-assemble into nanostructures at specific mineral compositions and pH. In this investigation, enzyme kinetics of MMP-20 proteolysis of recombinant full-length human amelogenin (rH174) under different mineral compositions is elucidated. Methods: Recombinant amelogenin was cleaved by MMP-20 under various physicochemical conditions and the products were analyzed by SDS-PAGE and MALDI-TOF MS. Results: It was observed that mineral ions largely affect cleavage pattern, and enzyme kinetics of rH174 hydrolysis. Out of the five selected mineral ion compositions. MMP-20 was most efficient at high calcium concentration, whereas it was slowest at high phosphate, and at high calcium and phosphate concentrations. In most of the compositions, N- and C-termini were cleaved rapidly at several places but the central region of amelogenin was protected up to some extent in solutions with high calcium and phosphate contents. Conclusion: These in vitro studies showed that the chemistry of the protein solutions can significantly alter the processing of amelogenin by MMP-20, which may have significant effects in vivo matrix assembly and subsequent calcium phosphate mineralization. General significance: This study elaborates the possibilities of the processing of the organic matrix into mineralized tissue during enamel development. (C) 2012 Elsevier B.V. All rights reserved.
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