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ENAMELYSIN (MMP20) AND KALLIKREIN 4 (KLK4)
FUNCTIONS DURING ENAMEL FORMATION
by
Zhi Sun
A Dissertation Presented to the
FACULTY OF THE GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(CRANIOFACIAL BIOLOGY)
May 2008
Copyright 2008 Zhi Sun
Object Description
| Title | Enamelysin (MMP20) and Kallikrein 4 (KLK4) functions during enamel formation |
| Author | Sun, Zhi |
| Author email | zhisun@usc.edu |
| Degree | Doctor of Philosophy |
| Document type | Dissertation |
| Degree program | Biology |
| School | School of Dentistry |
| Date defended/completed | 2008-02-25 |
| Date submitted | 2008 |
| Restricted until | Unrestricted |
| Date published | 2008-03-28 |
| Advisor (committee chair) | Oldak, Janet |
| Advisor (committee member) |
Paine, Michael Minkin, Cedric Markland, Francis Shuler, Charles |
| Abstract | Introduction: Dental enamel formation is a dynamic process involving crystal maturation concomitant with matrix proteolysis. Gene mutations of enamel proteases were found to cause amelogenesis imperfecta (AI), a hereditary disease affecting dental enamel. Two types of enamel proteases have been previously identified and reported to cleave amelogenin. However, the molecular mechanisms and function of enamel proteases during amelogenesis still remains unclear. I designed a series of in vitro experiments to systematically study the function of enamel proteases on amelogenin processing, assembly and interactions between amelogenin and apatite following proteolytic activities.; Methods and materials: Three amelogenin proteins were used: recombinant full length pig amelogenin (rP172), recombinant homolog of its major cleavage product (rP148), and a native amelogenin lacking both N- and C-termini (13k). Recombinant pig MMP20 (rpMMP20) and recombinant human kallikrein 4 (rhKLK4) were incubated with each of these three amelogenins with and without HAP crystals. Proteolytic products were detected by SDS-PAGE, HPLC, and mass spectrometry. Assembly properties of these three amelogenins and their proteolytic products were detected by dynamic light scattering (DLS) and atomic force microscopy (AFM). Affinity of thethree amelogenins to apatite was systematically evaluated by means of Langmuir model for protein adsorption.; Results: rpMMP20 cleaved rP172 more efficiently than it cleaved rP148 and it didn't process the "13k". rhKLK4 processed all of the above with an exponential reduction. These three amelogenins had different assembly properties. The cleavage of Cterminus led to aggregation of amelogenin; and the N-terminal cleavage resulted in the dis-assembly of amelogenin molecules. During proteolysis amelogenin substrates had higher binding affinity to HAP than their proteolytic products. HAP did not change the cleavage pattern of amelogenin by proteases. Comparative adsorption isotherms indicated the binding affinity of amelogenins in the order, rP172>rP148>13k. Conclusions: Processing of amelogenin by MMP20 and KLK-4 is stepwise, progressive, and with different patterns. Enamel proteinases cause alterations of amelogenin assembly with consequence of dis-assembly. Cleavages of C- and N-termini of amelogenin reduce its apatite binding affinity. We suggest that such controlled proteolysis will dictate the function of amelogenin in controlling crystal morphology and orientation. |
| Keyword | MMP20; KLK4; enamel; amelogenin; self-assembly; apatite |
| Language | English |
| Part of collection | University of Southern California dissertations and theses |
| Publisher (of the original version) | University of Southern California |
| Place of publication (of the original version) | Los Angeles, California |
| Publisher (of the digital version) | University of Southern California. Libraries |
| Type | texts |
| Legacy record ID | usctheses-m1063 |
| Rights | Sun, Zhi |
| Repository name | Libraries, University of Southern California |
| Repository address | Los Angeles, California |
| Repository email | http://www.usc.edu/isd/libraries/services/ask_a_librarian/email/ |
| Filename | etd-Sun-20080328 |
| Archival file | uscthesesreloadpub_Volume26/etd-Sun-20080328.pdf |
Description
| Title | Page 1 |
| Full text | ENAMELYSIN (MMP20) AND KALLIKREIN 4 (KLK4) FUNCTIONS DURING ENAMEL FORMATION by Zhi Sun A Dissertation Presented to the FACULTY OF THE GRADUATE SCHOOL UNIVERSITY OF SOUTHERN CALIFORNIA In Partial Fulfillment of the Requirements for the Degree DOCTOR OF PHILOSOPHY (CRANIOFACIAL BIOLOGY) May 2008 Copyright 2008 Zhi Sun |
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