Tissue inhibitors of metalloproteinases (TIMPs) comprise a family of four proteins in humans that modulate the turnover of the extracellular matrix by regulating the activities of endopeptidases that catalyze its degradation, especially the matrix metalloproteinases (MMP). In general, the four TIMPs are broad-spectrum tight binding inhibitors of MMPs with individual differences in specificity. In this study, we attempted to understand the basis of such variation by using membrane type-1 MMP (MT1-MMP) as a model, since it is inefficiently inhibited by TIMP-1 in contrast with the other TIMPs. We designed and engineered mutations in the N-domain of TIMP-1, based on current knowledge of TIMP interactions. By measuring inhibition levels of each mutant against several MMPs, including MT1-MMP, we were able to obtain a triple mutant with an vii improved affinity for MT1-MMP. / Our results, along with previous data, confirm that multiple residues in the critical interface segments between TIMPs and MMPs, namely at positions 2, 4, 5, 6, and 98, are key in determining the basic interaction between the two molecules. The second part of this work focused on naturally occurring mutations in TIMP-3 which cause an early form of macular degeneration called Sorsby's Fundus Dystrophy (SFD). The TIMP-3 mutants identified so far share certain features but the mechanism by which they result in macular disease is not yet understood. As an initial step, we expressed recombinant TIMP-3 carrying a truncation mutation, glutamic acid 139 to a stop codon (E139X), and assessed its activity towards representative MMPs and tumor necrosis factor-(Sa (Bconverting enzyme, another metalloproteinase normally inhibited by TIMP-3. Our results indicate that this mutation does not impair the inhibitory activity of TIMP-3. / Expression of this mutant in mammalian retinal cells revealed a difference in localization between wild-type and E139X mutant TIMP-3. Therefore, we concluded that the SFD mutations may actually influence the processing and/or binding properties of TIMP-3 in the retina. / by Asmaa Bilal Hamze. / Vita. / Thesis (Ph.D.)--Florida Atlantic University, 2008. / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2008. Mode of access: World Wide Web.
| Identifer | oai:union.ndltd.org:fau.edu/oai:fau.digital.flvc.org:fau_2863 |
| Contributors | Hamze, Asmaa Bilal., Charles E. Schmidt College of Science, Department of Biological Sciences |
| Publisher | Florida Atlantic University |
| Source Sets | Florida Atlantic University |
| Language | English |
| Detected Language | English |
| Type | Text, Electronic Thesis or Dissertation |
| Format | xiii, 135 p. : ill. (some col.)., electronic |
| Rights | http://rightsstatements.org/vocab/InC/1.0/ |
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